Scientific Malaysian Issue 12

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The Vaccine & Immunology Edition

CONTENTS

EDITOR’S FOREWORD W

Editors Adaikalavan Ramasamy, PhD Andrew Chan, PhD Beatrice Chin Hui Tze Jaa Yien New, PhD Juliana Ariffin, PhD Kok Onn Kwong Lee Khai Ern, PhD Valerie Soo, PhD Lee Hooi Ling, PhD

J U N E 2 0 1 6 ISSUE NUMBER 12

Featuring:

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The Top 10 Finalists of SciMy’s Writing Competition

EDITOR’S FOREWORD

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SciMy Ask Me Anything (AMA):

Betty Kim Lee Sim The Founder of Protein Potential LLC & Executive Vice-President at Sanaria Inc. lets us in on her thoughts and experiences.

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Gems From Our Web Articles

Vaccines: An Immunological Revolution 1 By Juliana Ariffin as a Cure for Cancer?

8 Viruses

By Suet Lin Chia

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Candice Lim

A ‘Trp’ to the Land of Kyns

By Felicita Fedelis Jusof Adli Ali

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The Imitation

By Nur Atikah Abdullah, Charles George Gajim & Seti Faezah Rosli

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Teaching Your Immune System to Fight Cancer

By Litt-Yee Hiew

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Illustration Alle Chun Kong Yink Heay Mohd. Arshad Yusoff

Suet Lin Chia

Snake Antivenoms: Science, Values & Challenges

By Tan Choo Hock

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Designer Xinhui Yap

Superhero

By Candice Lim Childhood Vaccine Controversies: the Myths, the Facts and the Uncertainties

By Adli Ali

Felicita Fedelis Jusof

Tan Choo Hock

Nur Atikah Abdullah Charles George Gajim

Seti Faezah Rosli

Litt-Yee Hiew Adaikalavan

Current Progress on Malaria Vaccines

By Adaikalavan Ramasamy

Ramasamy

Nor Ilham Ainaa

The Dengue Vaccine Dilemma: Route to Prevention - Are We There Yet?

By Nor Ilham Ainaa

Cover Illustration

The cover image for this issue symbolises the focus of our theme; depicting the combat against diseases through the development and enlistment of vaccines and immunology. Adapted from an illustration by Mohd Arshad; Scientific Malaysian Magazine Issue 12 (June 2016) ©

Editor-in-Chief Hwong Yi Ling

Hwong Yi Ling Editor-in-Chief

SCIENTIFIC MALAYSIAN MAGAZINE is published in a web format (http://magazine. scientificmalaysian.com) and in a downloadable digital magazine format (PDF).

Our digital magazines are distributed to Malaysian societies around the world.

The Scientific Malaysian Magazine is published biannually and it is FREE of charge.

DISCLAIMER Opinions and articles published in Scientific Malaysian Magazine do not necessarily reflect those of the editors/staff and members of SciMy. All articles and original photos/illustrations cannot be reproduced without prior permission from the author(s) and Scientific Malaysian.

CONTACT DETAILS Email: magazine@ scientificmalaysian.com Website: http://www. scientificmalaysian.com Facebook: http://facebook. com/ScientificMalaysian Twitter: @ScientificMsian

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Vaccines: An immunological Revolution

By Juliana Ariffin

Did you know that if you went back in time, most of you would be immune to some of the most devastating diseases that have ravaged humankind in the past? We owe this invisible armour that we possess to the hard work and sacrifice of our predecessors who discovered… you guessed it, vaccines!

LOGY O N U IMM K S BAC T H G I F

The World’s First Vaccine Vanquishes Smallpox; The Killer of Kings

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pproximately 3,000 years ago in ancient Egypt, there lived a young pharaoh by the name of Ramses V. Not much is known about his reign, except that almost 4 years after he was anointed as the ‘living god’ of Egypt, Ramses V probably felt unusually lethargic and feverish.

He might also have thrown up a few times, had a sore throat or a bad headache. Initially, these symptoms might have been attributed to a minor malady, such as the flu or stress.

However, these symptoms were just a prelude.

Within the next few days, a bumpy rash appeared all over his face and body, and sores developed in his mouth, throat and nose.

These rashes then swelled further and leaked infectious pus.

Finally in 1157 B.C., approximately two weeks after he fell sick, Ramses V died, leaving only his scarred and mummified remains as his legacy.

These would be discovered in 1910 A.D., and would become the earliest physical evidence of smallpox [2], one of the deadliest, and now extinct, diseases to have plagued humankind.

Of variolation and vaccines Unfortunately for Ramses V, no reliable way to guard against smallpox existed during his time. In fact, since 10,000 B.C., humans have been vulnerable to smallpox epidemics that trailed the pattern of human migration, resulting in a marked impact on human history—smallpox is known to have been more influential in bringing down the Aztec and Incan empires than the invading Spanish Conquistadors [3]. By the end of the 18th century, smallpox had killed five European kings, and by the 20th century 300-500 million people worldwide had died from smallpox, a number far exceeding the combined fatalities from the two world wars [3].

The fight against smallpox, and thus the discovery of vaccines, began in 430 B.C. when the Athenian, Thucydides, observed that survivors of smallpox became immune to it. In 910 A.D., the Persian alchemistphysician-philosopher, Abu Bakr Muhammad Bin Zakariya Ar-Razi (Rhazes), also noted the immunity of survivors and person-to-to-person transmission of smallpox, leading him to propose the first theory of acquired immunity [3, 4].

Thucydides and Rhazes were not alone in their observations. By 600 A.D., physicians in China were grinding up dried smallpox scabs

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with musk to inoculate the noses of healthy people, while people in India wore shirts from infected patients and slept alongside smallpox victims [3, 5]. In Africa and the Middle East, scrapings from smallpox pustules from a mildly infected individual were applied onto a scratch or a vein of a healthy person. These methods of transmitting smallpox are called variolation, and were conducted with the hope that a minor infection with smallpox would occur, stimulating the immune response and conferring immunity against the disease [3, 5].

Variolation was practised for hundreds of years although it was not a foolproof method, due to variations in the amount of virus transmitted, and the fact that inoculated individuals could still transmit the disease. Still, it saved the lives of many, including Catherine the Great and her son [5], and was used by George Washington to safeguard the American army during the War of Independence against Great Britain [6].

In 1795, Edward Jenner observed that milkmaids who had contracted cowpox were immune to smallpox. He experimented using the pus from a cowpox pustule on a milkmaid, Sarah Nelmes, to inoculate scratches on the arm of an 8-year-old boy, James Phipps.

Luckily for James, he developed cowpox, but like the milkmaids, was immune to smallpox.

Jenner then developed the world’s first vaccine using cow serum containing the cowpox virus [7]. This method of preventing smallpox was strongly supported by the then U.S President, Thomas Jefferson, and Napoleon Bonaparte, who had his entire army vaccinated in 1805 and ordered all French civilians to be vaccinated a year later [8]. Over the next few centuries, immunity against smallpox became widespread, resulting in global eradication of the disease in 1980 [9].

“physicians in China were grinding up

Dried smallpox scabs ...to inoculate the noses of

healthy people”

How vaccines work

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ince Edward Jenner’s discovery, vaccines have been developed for many other diseases including polio, chicken pox, hepatitis B, human papillomavirus (HPV) and influenza.

These vaccines work by introducing an agent (Infobox 1) from a disease-causing microorganism that is recognised by the immune system as a threat. This agent stimulates an immune reaction that persists for years in the form of ‘immunological memory’, enabling a quicker and more effective immune response upon future encounters with the microorganism.

INFOBOX 1

This system of speedy recognition and the launching of a specific defence is termed adaptive immunity, and is what enables the body to destroy invading microorganisms before they enter cells, or to destroy infected cells before the microorganism can multiply to huge numbers within the body. When developing a vaccine, the biggest concern is how best to induce an immune reaction while keeping the risk of developing the disease low.

While vaccination protects against a disease, it doesn’t always confer lasting protection.

Just as normal memories fade, immunological

Types of vaccines and the diseases they protect against [18, 19]

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memory can also decline, resulting in lost immunity years after the initial vaccination.

This can be detected by measuring antibody levels agains the vaccinated agent. In cases where immunity has declined, administration of a ‘booster’ shot, or an extra administration of a vaccine following the earlier dose, helps

to regain immunity. Example of vaccines that normally require follow-up booster shots include the oral polio vaccine (OPV) which persists for only 6 months and tetanus, which requires a booster shot every 10 years.

FIG. 1

Herd Immunity (image credit to Bioninja [20])

Herd immunity and pox parties

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hildren are usually vaccinated as soon as their immune system is sufficiently developed, and many countries have a schedule of recommended vaccinations.

However, the large number of injections for vaccinations and booster shots, as well as the fear of side effects often leads to problems in compliance with the schedule.

One of the biggest hindrances for vaccination was the fraudulent claim by Andrew Wakefield, a former British surgeon and medical researcher who published a paper that described a link between the measles, mumps and rubella (MMR) vaccine, and the occurrence of autism and bowel disease [10]. Although his research was not reproducible, was shown to be financially motivated and was also revealed to be conducted without ethical approval [10], mass media disseminated his findings reporting a link between vaccination and autism. As a result, many parents have chosen to ‘protect’ their children from autism by not allowing them to be vaccinated.

Instead, some parents have resorted to holding pox parties where healthy children are exposed to a child infected with a disease such as chickenpox, measles, or rubella. Similarly to variolation, this hopefully allows for an infection and acquisition of natural immunity. If the majority of the public were vaccinated, and

if vaccinations conferred lasting protection, this would not be such a major concern for everyone. Indeed, the biggest threat would be unvaccinated children succumbing to severe disease and being at risk of fatality or deformity.

However, as mentioned above, vaccinations do not confer lasting protection. Also, individuals who are too young, too old, pregnant, or immunocompromised are unable to receive vaccinations. These individuals are at high risk of contracting disease if they come in contact with an infected individual.

Normally, the number of vaccinated individuals is high enough that it confers what is termed ‘herd immunity’ (Fig. 1) to a population, where the incidence of contracting the disease is so low that even unvaccinated individuals will be protected. However, if the number of unvaccinated individuals increases, such as an increase in immunocompromised individuals or when children are intentionally not vaccinated (even if they do not attend pox parties), herd immunity is no longer effective and the population becomes at risk of a disease epidemic [11].

About the author: Juliana Ariffin

is a postdoctoral fellow researching liver inflammation at Beth Israel Deaconess, Medical Center, Harvard Medical School. Prior to this, she studied human immune responses at The Institute for Molecular Biosciences, The University of Queensland. In her spare time she reads and writes fiction, dabbles in photography and considers genetically engineering a zombie propagating virus to repopulate the earth.

Find out more about Juliana Ariffin by visiting her Scientific Malaysian profile: http://www.scientificmalaysian.com/members/julianna/

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REFERENCES 1) Ramesses V, Wikipedia entry. (https:// en.wikipedia.org/wiki/Ramesses_V) 2) Erik Hornung. (1997). The Pharaoh, p.292 in The Egyptians (ed.) Sergio Donadoni and Robert Bianchi, University of Chicago Press.

3) Epidemics of the Past, Smallpox: 12,000 Years of Terror. (http://www.infoplease.com/ cig/dangerous-diseases-epidemics/smallpox12000-years-terror.html) 4) SC Ashtiyani, A Amoozandeh. (2010).

Rhazes Diagnostic Differentiation of Smallpox and Measles, Iranian Red Crescent Medical Journal. (http://ircmj.com/291.fulltext) 5) Variolation, Wikipedia entry en.wikipedia.org/wiki/Variolation)

(https://

6) Benjamin A. Drew. (2015). George Washington and Smallpox, A Revolutionary Hero and Public Health Activist, JAMA Dermatology. (http://archderm.jamanetwork.com/article. aspx?articleID=2169316) 7) Stefan Riedel. (2005). Edward Jenner and the history of smallpox and vaccination, Proc (Bayl Univ Med Cent). (http://www.ncbi.nlm.nih. gov/pmc/articles/PMC1200696/) 8) Sheryl Persson. (2010). Smallpox, Syphilis and Salvation: Medical Breakthroughs that Changed the World.

9) World Health Organisation, Smallpox. (http:// www.who.int/csr/disease/smallpox/en/) 10) Andrew Wakefield, Wikipedia entry. (https:// en.wikipedia.org/wiki/Andrew_Wakefield) 11) Vaccines.gov, Community Immunity. (http:// www.vaccines.gov/basics/protection/) 12) Centers for Disease Control and Prevention, Vaccines and Immunisations (http://www.cdc. gov/vaccines/vac-gen/whatifstop.htm).

13) Centers for Disease Control and Prevention, Emergency Preparedness and Response, Smallpox. (http://www.bt.cdc.gov/agent/ smallpox/vaccination/faq.asp).

14) Penny Sarchet. (2016). Use vaccines as a weapon against antibiotic-resistant bacteria, Newscientist. (https://www.newscientist.com/ article/2077157-use-vaccines-as-a-weaponagainst-antibiotic-resistant-bacteria/)

Viruses as a Cure for Cancer?

By Suet Lin Chia

15) Michael Brooks. (2016). Booster shots: the accidental advantages of vaccines, Newscientist. (https://www.newscientist.com/ article/dn24027-booster-shots-the-accidentaladvantages-of-vaccines/) 16) Lindsey et al., (2016). Assessment of the Safety and Immunogenicity of 2 Novel Vaccine Platforms for HIV-1 Prevention: A Randomised Trial, Annals of Internal Medicine. (http://annals. org/article.aspx?articleid=2484873) 17) Abhishek et al., (2016). Dendritic cell vaccines based on immunogenic cell death elicit danger signals and T cell-driven rejection of high-grade glioma, Science Translational Medicine. (http://stm.sciencemag.org/ content/8/328/328ra27) 18) Vaccine, Wikipedia entry. en.wikipedia.org/wiki/Vaccine)

At a Glance: Cancer has been haunting the human population for centuries. Despite the availability of various current treatments, the number of cancer patients is on the rise. This article discusses an alternative treatment for cancer – cancer virotherapy, a concept of using virus to kill cancer cells. It has been the subject of investigation for decades but has only recently been approved by the US Food and Drug Administration (FDA) as a therapeutic agent for cancer. A brief description of the history of virotherapy, how the virus kill the cancer

(https://

19) World Health Organisation, Vaccine Safety Basics, Types of Vaccine. (http://vaccinesafety-training.org/types-of-vaccine.html) 20) Bioninja, Vaccination (http://ib.bioninja.com. au/higher-level/topic-11-animal-physiology/111antibody-production-and/vaccination.html)

In today’s society,

cancer is no longer a foreign word. The chances that we know someone who has been diagnosed with or died from cancer are on the rise. In 2012 alone, the World Health Organisation (WHO) reported 32.6 million people living with cancer, 14.1 million new cases and 8.2 million deaths from cancer. Most fatalities result from late diagnosis or treatment.

In many of these cases, the cancer cells have metastasised

(or spread) to other organs and are beyond curability with current technologies. In cases of early detection, surgical procedures can be performed to remove the cancer, followed by chemotherapy and radiotherapy to kill any remaining “escaped” cells. These treatments have become so commonplace that they are considered obligatory for any cancer treatment regimen. Less recognised is that, after surgical excision and chemotherapy, 40-60%

of cancers detected even in early stages of the disease would occur again, with a much higher rate of recurrence in late stage cancers. Recurrence arises from the “escape” of some cancer cells from surgery or chemotherapy which subsequently develop resistance towards chemotherapy drugs.

And when they come back, they come back with a vengeance.

These cancer cells continue to grow, forming a more aggressive cancerous cell population that

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Progress in science and technology has made it possible to engineer viruses to kill cancer cells in patients. Viruses are very small particles, invisible even under a normal microscope. In order to replicate, viruses must

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Issue 12/2016 the host. For example, hepatitis viruses infect liver cells but not brain cells. Similarly, a virus that infects cancer cells will not infect normal cells. Even if normal cells are infected by accident, defence proteins present in

known as “oncolysis” (“onco”: cancer; “lysis”: killing by disrupting the cell membrane). reported in the early 19th century in a cancer patient virus. Curiously, the cancer shrank after infection with the virus. Although scientists did not fully understand the phenomenon, they suggested that the virus might have contributed to the remission.

In the years that followed, substantial research focused on identifying existing viruses with similar cancer-killing properties. Surprisingly, many viruses were found to be very promising in destroying cancer cells. Some viruses, such as coxsackieviruses, reoviruses, herpes simplex viruses and adenoviruses, are

“Oncolysis” Onco: Cancer

Lysis:

host (such as plants, animals Killing by disrupting the cell or bacteria).

Infection membrane occurs via proteins on the surface of the virus that act as a “key” that inserts into a these cells, but not in their “lock”, represented by receptor cancerous counterparts, will proteins on cell surface. The interaction between the two replication. proteins helps the virus to enter the cell and begin to replicate. sores, and gastrointestinal The key-and-lock system is so Viruses kill cancer diseases in humans. Others, cells through a process like Newcastle disease virus and

vaccinia virus, are responsible for animal diseases. Whilst a number of viruses, including Newcastle disease virus and reoviruses, possess a natural ability to selectively kill cancer cells, others can be genetically spare normal cells.

But how do viruses

kill cancer cells? Generally, cancer cells were once normal dying through a process called programmed cell death. Cancer cells, however, may acquire mutations in genes responsible for cell death. This prevents cancer cells from undergoing normal programmed cell death and allows them to continue growing unchecked.

This uncontrolled growth eventually leads to a lump of cancer cells and the creation of a favourable “habitat” for oncolytic viruses, which exploit the immortal properties of cancer cells as virus production factories. As the viruses mature, they burst open the cell and spread to surrounding cancer cells. In

addition, as cancer cells are burst open and killed, the body’s immune cells are attracted

approximately 15-30 patients to determine the safe dosage, route of administration of the

destroyed cells. This, in turn, educates our immune system to recognise distinct “traits” or cancer antigens, which act like a “cancer vaccine”, and start attacking remaining cancer cells including those at a distant site that display these antigens on their surface. As the cancer cell population is killed, the oncolytic viruses run out of host cells to grow in and are cleared by the body’s immune system.

new treatment on the human body; phase II involves less than 100 patients to determine the

Despite studies

numerous

indicating that oncolytic viruses are relatively safe for use in humans (see for example, [1]), the general public remains sceptical. Such scepticism is the very reason that therapeutic agents undergo many stages of preclinical trials on animals, followed by very closely monitored clinical trials in humans. Clinical trials commonly consist of three phases: phase I involves

for various cancers and also the body; phase III involves hundreds to thousands of patients to the new treatment compared to the current standard treatment.

Many oncolytic viruses are currently in clinical trial phases I, II or III. OncoVEX (herpes simplex virus) and Reolysin® (reovirus) are examples of oncolytic viruses currently being studied in phase III clinical trials. been demonstrated, they can be approved by the Food and Drug Administration (FDA) for use as cancer treatment in hospitals. In fact, 2015 was a historic year for cancer virotherapy. IMLYGIC™ (talimogene laherparepvec), oncolytic virus to be licenced by the FDA for the treatment

“Despite numerous studies indicating that oncolytic viruses are relatively safe for use in humans... the general public remains sceptical” Illustration by Kong Yink Heay

11 Issue 12/2016 of melanoma. In addition, Oncorine (adenovirus) has also been approved in China for use in cancer treatments.

So far, patients’ outcomes have been

very promising, and clinical successes were reproducible [2,3]. Solid cancer masses in patients have been shown to shrink or at least cease growing [4]. Importantly, these therapies may also give patients a better quality of life; virus treatments are associated with much fewer

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Issue 12/2016 to those from chemotherapy and radiotherapy [1,5]. Despite promising results in clinical trials, however, virotherapy does have its limitations. Though these viruses are used therapeutically, they are still recognised as foreign particles or “antigens” to our body. This alerts the body’s immune response to detect and eliminate the viruses from the body. In addition, the administration of viral therapies requires trained professionals and close monitoring of the patients during treatment.

These requirements pose major obstacles and explain, at least in

part, why only a few countries in the world currently provide this treatment. Nevertheless, with international teams of scientists currently working on genetically modifying oncolytic viruses to improve their safety and under investigation with the hope of isolating more potent variants, the future is bright for virotherapy. One day, we hope to be able to treat cancer like a available oncolytic virus pills and eliminate the public fear associated with cancer.

SUPERHERO ...By Candice Lim

Illustrations by Alle Chun

“Mommy,

Hi, I’m Leila!

I don’t wanna take

the shot,” whimpered the little girl in my clinic this morning.

About The Author: Suet Lin Chia is a senior lecturer at the Department of Microbiology in the Faculty

of Biotechnology and Biomolecular Sciences at the Universiti Putra Malaysia. He is currently a postdoctoral research fellow working on oncolytic virotherapy in the laboratory of Professor Len Seymour at the University of Oxford. The author has also virotherapy. Dr. Chia aims to contribute to the development of cancer therapeutics and welcomes collaborations with researchers with the same goal. The author can be contacted at suetlin@upm.edu.my

She was dressed all in pink and didn’t look more than five years old.

and it’s killed many people...” She paused, then continued. “Daddy’s one of them.”

“Please be a good girl, Leila. It’s not gonna hurt,” coaxed her mother. She looked tired and had two black circles under her baggy eyes. Like the rest of the patients milling in the waiting room, she had come with her daughter to get her flu shot.

I nodded and swallowed the lump in my throat.

The virus was strong and had claimed more lives than scientists had predicted it would. The nation was in a panic. When the Food and Drug Administration (FDA) gave the green light for the new vaccine, everybody started to queue up at the nearest clinics to get their shots. “Do you know what that means?”

“It means we’re gonna die.”

/

References: 1. Zeh HJ, Downs-Canner S, McCart JA, Guo ZS, Rao UN, Ramalingam L, Thorne SH, Jones HL, Kalinski P, Wieckowski E, O’Malley ME, Daneshmand M, Hu K, Bell JC, Hwang TH, Moon A, Breitbach CJ, Kirn DH, Bartlett DL (2015) First-in-man study of Western Reserve strain oncolytic vaccinia virus: safety, systemic spread, and antitumor activity. Mol Ther 23(1): 202–214.

2. Russell SJ, Peng KW, Bell JC (2012) Oncolytic virotherapy. Nat Biotechnol 30(7): 658–670.

3. Chiocca EA, Rabkin SD (2014) Oncolytic viruses and their application to cancer immunotherapy. Cancer Immunol Res 2(4): 295–300.

Leila

Today is the day to get my flu shot, but I don’t want to, because it’s gonna hurt!

“No, Leila.” I took her hands and smiled at her.

“You’re not going to die.”

4. Carroll J (2013) Amgen trumpets T-Vec oncolytic virus results from Ph III melanoma study. Chicago, IL: FierceBiotech; 2013 [cited 2016 Feb 11]. Available from trumpets-t-vec-oncolytic-virus-results-phiii-melanomastudy/2013-06-01.

I sat down next to Leila and took her hand.

She looked at me with her two big eyes.

Cancer in: Miner BR (ed.) Cancer management in Man: Chemotherapy, Biological Therapy, Hyperthermia and Supporting Measures. Springer Science+Business Media B.V. pp.295-316.

“Leila.” She took my hand hesitantly and quickly let go.

“Nice to meet you, Leila. Now, can you tell me what’s happening?”

“Hello, my name’s Doctor Hershey. What’s your name?”

Leila looked around her for a while, then stared at her feet. “There’s a new virus out there

“Then… then, why do we have to take the shots?”

“We’re going to make you a Superhero.”

Doctor Hershey

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“A superhero?” Leila’s eyes lit up as she grinned, baring her two missing front teeth. It’s been a while since I saw a smile, especially from a child.

Your body’s strong enough to withstand the vaccine. But, what about the newborns, pregnant women, and those with life-threatening illnesses?”

I nodded. “Yes, the shot will give you the superpower to protect yourself and others who can’t have their own injection against the virus.”

“But.” Leila turned to look at Mommy. “Mommy told me the shot contains viruses. I don’t wanna have viruses in my body.”

“Yes, they are, but do you know what they do?”

Leila shook her head.

“These viruses are inactivated, meaning they cannot make copies of themselves or cause harm to your body. They’re safe. There’s nothing to worry about. Your body will still see them as a threat and when it does, it will produce antibodies to attack them. Then your body will remember the virus. If you happen to come into contact with the flu, your body will immediately recognise the same virus and produce the antibodies to fight them. This is called ‘active immunity’ [1-2].”

good viruses

bad viruses

“Against the flu virus, yes.”

“But, how can I protect others?”

“Look, Leila. You’re a strong and healthy girl.

Science can make them superheroes. They don’t know that day came a long time ago.” Removing the syringe from the metal tray and pushing the air out, I beamed at the little girl. Leila and her mother looked so much happier and confident than when they first came in.

“They are. They just fight a different kind of bad guy. Many people anticipate the day

“So, Leila, are you ready to receive your superpower?”

I shook my head. “Their bodies cannot resist even the inactivated viruses. They’ll have to depend on the people around them to be their human shield. If no one has the immunity, the flu can spread around and get to them easily.

But when most people get their shots, there’s little chance for an outbreak. In this way, the vulnerable ones are protected from the disease.

This is called ‘herd’ or ‘community immunity’ [34].”

Leila bit her lip and bent her head. “It’s gonna hurt a lot, right?”

Leila stared at me for a couple of seconds and nodded confidently. “Yes, I am. I can do this!”

“Cool!” exclaimed the little girl. “That means I’m invincible!”

“I can’t believe it. Superheroes are real!” Leila smiled brightly at her mother. “I’ve always thought they were only in the movies.”

“They cannot be vaccinated?” Leila frowned.

“Well, this is the test you have to go through to become a true superhero. Do you think you’re brave enough to be a superhero?”

POWER

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I turned to the mother as she ran her hand over her daughter’s hair. “You’re so lucky, ma’am. You have a brave daughter. I’m sure she’s going to be a great superhero.”

“Thank you so much, Doctor. Leila was so worried when I told her we’re going to get our flu shots,” said Leila’s mother thankfully.

“Don’t mention it, ma’am. That’s our job.”

Leila

ABOUT THE AUTHOR

Candice Lim Wan Chi is a science writer by day and a science fiction and

fantasy (SFF) novelist by night. Graduated with a biotech degree, her research interests lie in genetics, tissue culture, transgenesis, and of course cyborgs, androids, blasters, and spaceships. She’s working to publish her SFF trilogies “Outbreak” and “Hell Break” under pseudonym James Levin. She hopes to pursue an entrepreneurial venture in science.

Find out more about Candice by visiting her profile at: http://www.scientificmalaysian.com/members/candistic/

References 1)Burton, D. R. (2002). Antibodies, viruses and vaccines. Nature Reviews Immunology, 2(9), 706-713.

2)Pulendran, B., & Ahmed, R. (2011). Immunological mechanisms of vaccination. Nature Immunology, 12(6), 509–517.

3)Fine, P., Eames, K., & Heymann, D. L. (2011). “Herd immunity”: A rough guide. Clinical Infectious Diseases, 52(7), 911-916.

4) Valleron, A. J. (2012). Can the modeling of herd immunity help design influenza immunization policy?. Preventive Medicine, 55(1), 7879.

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Childhood Vaccine Controversies: myths, the facts the uncertainties

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CONTROVERSY 1:

Vaccines cause autism Verdict: MYTH This “myth” was based on an article published in 1998 in The Lancet, written by Andrew Wakefield, a British surgeon, suggesting that the measles, mumps and rubella (MMR) vaccine increased autism incidences among British children [1]. This “infamous” article and his claim on the causal-relationship between vaccines and autism that ended up being discredited created a widespread fear among the public. Up until now, none of the numerous subsequent studies conducted were able to prove Dr. Wakefield’s claim and no scientific evidence has proven any possible link between the MMR vaccine and autism [2, 3, 4]. The article was eventually deemed flawed and retracted by The Lancet based on the serious procedural errors, undisclosed financial conflicts of interest, and ethical violations [5]. This controversy will always be a major myth in the vaccine world, hence, is officially referred to as the most damaging medical hoax of the century.

By Adli Ali

Vaccine controversies and anti-vaccine movements are not something new. They began some 90 years ago in the early 20th century, even before the term “vaccination” was formally used. Let us now revisit what we may consider to be the 7 most scientifically relevant controversial issues relating to “vaccines” and “vaccination” and explore the facts and verdicts for each of these issues.

Figure 1: Reduction of measles cases and death upon measles vaccine introduction in 1968 in England and Wales (Data source: Public Health of England [17])

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CONTROVERSY 2:

Vaccines contain mercury and are damaging to the body, leading to autism Verdict: MYTH Vaccines did contain Thiomersal, an organic mercury-containing compound, which was used in small amounts previously as antifungal preservative in some multi-doses vaccines. The amount used was significantly below the acceptable level considered tolerable by the World Health Organisation (WHO), and had evidently proven to be safe. Nonetheless, as a precautionary principle, Center for Disease Control (CDC) had directed removal of thiomersal from all childhood vaccines in 1999 [6]. Concerns on Thiomersal could have been responsible for autism was disapproved based on the fact that no causal relationship was ever documented, and despite current removal of thiomersal-based vaccines, the steady increase in incidences of autism is maintained [7].

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CONTROVERSY 4:

Human foetuses are aborted to produce vaccines Verdict: PARTIAL FACT Research development and production of vaccines, especially in virus-based-vaccines, require culturing of the virus in specific human cells. The statement that human foetuses were aborted to produce vaccine is untrue, as in the development process of vaccines, commercial-laboratory produced human cell-line culture are being used [9]. Nonetheless, historically, these commercially grown cells were harvested through intentional medically-sacrificed human fetuses, approved and monitored strictly by ethical bodies, and a few decades ago, this was done purely for the need of the scientific research [10, 11]. No subsequent abortion was performed thereafter for any other scientific purposes. The usage of these foetus cell lines is extensive, extending to many different fields of the sciences, not just vaccine research and development.

CONTROVERSY 3:

Some vaccines are not Halal Verdict: PARTIAL FACT Some vaccines use porcine-based-enzymes in their production, creating concerns on the “halal”ness of those vaccines. However, despite the fact that these enzymes are used in some of the processes of the vaccine manufacturing, the Islamic fatwa committee has allowed the usage of these vaccines. This is based on several Islamic principles in dissecting the issue. On the “halal”ness of the vaccine product itself, the concept of the “tiny amount” of enzymes added to the huge quantity of vaccine solution produced, in line with the concept of “suci” or “pure element” in Islam. Secondly, the removal of the used enzymes in the final production is also parallel to the concept of “cleanliness and pure element” in Islam, hence making the final product as “not containing” any of the “non-halal” elements through the principle of look, smell and taste. Finally, based on the crucial need of using the vaccine in disease prevention in situations whereby we do not have other “purely halal” alternatives, Islamic scholars have ruled that the use of this vaccine may be allowed in accordance to a ruling based on the concept of “dharurah” or emergency situations. However there exist some differences of opinion on this matter, whereby the more strict scholars cannot accept this new “law” based on “ijma” or “agreements” of scholars, particularly in relation to the “halal” concept [8].

Illustration

by Mohd Arsh a

d

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CONTROVERSY 5:

Natural protection is adequate and safer, vaccination is just redundant Verdict: PARTIAL FACT Natural infection does provide lifelong immunity against certain infections. However, in the case of most vaccine-preventable diseases, the idea that natural infection is safer and adequate is dangerously untrue. Vaccine-preventable infections are proven to cause significant health problems, such as neurological side effects (or sequelae) from Hemophilus influenza meningitis and birth defects from maternal rubella infection, and fatality. In some organisms, a single naturally occurring infection may not be adequate to prevent the next infection and may lead to more severe disease, such as in the case of Dengue fever. Vaccination does not only prevent infection, but also prevents the disease associated with natural infection, i.e. cervical cancer through Human Papillomavirus (HPV) vaccination and hepatitis through Hepatitis B vaccination. As many organisms also have several serotypes, vaccination is not redundant in naturally protected individuals, as the vaccine may provide protection against other serotypes and enhance immune response towards the naturally protected serotype.

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CONTROVERSY 7:

Vaccination is not without side effects Verdict: FACT As with any other drugs or interventions, vaccination is, of course, not without side effects. Most of them are mild and tolerable side effects such as fever, flu-like illness, and pain and/or redness at local injection site pain. These known side effects are clearly stated in the information sheets and well-tolerated in many. Development of vaccines, as with most drugs, involves several phases, and safety is among the first phase requirement before the candidate vaccine even goes to the clinical trial phase. Nonetheless, there were incidences where serious and unexpected complications occurred and only recognised after the wide usage of the vaccine within the population. One example is in the case of the rotavirus vaccine, Rotashield. This vaccination causes an increased incidence of intussusception among toddlers, leading to its retraction from usage soon after it was noticed [13]. Tight surveillance is in place to monitor the safety of any drug and vaccine, as experience in different wider population might reveal new entities not discovered during the clinical trials.

CONTROVERSY 6:

Too many childhood vaccinations, immune system overloaded Verdict: MYTH Our immune system is just remarkable. Without us even realising it, the immune system is constantly exposed and able to prevent infections competently. Giving several vaccines at the same time is shown to be safe, immunologically effective, logistically and economically efficient.

Concerns on overloading the immune system are false and based on a non-immunological assumption. It is crucial that protection is achieved by a certain point of time in a child’s life. This is based on the epidemiological knowledge on the prevalence and likeliness to encounter the infection. Combination vaccines also means less injections which lead to less parental anxiety and stress on the child [12].

There are several other arguments, not discussed in this article, that are being exploited by the anti-vaccine movements which are based on sentiment and false-beliefs [14]. Faulty arguments that we do not need vaccination for measles since the infection is no longer a major problem, has led to the current measles outbreak in several countries [15, 16]. It is important for scientists, physicians and patients to know the advantages and the limitation of vaccines. Vaccination has come a long way since its discovery and had changed its primary role in infection prevention, to even therapeutic and disease prevention now. The new technology embedded in vaccine development, using an antigenic part of the pathogen (subunit vaccine) or the pathogen’s DNA materials (DNA vaccine) may not just come with less side effects, but possibly a more effective vaccine and also the answer to many more diseases that need to be tackled where a vaccine is yet to be developed.

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About The Author: Dr Adli Ali is a medical academician and clinical scientist, passionate about exploring the mysterious and fascinating world of immunology. A paediatrician at heart and by training, he also loves the science of fantastic gourmet and the art of travelling the world. Currently furthering his sub-specialisation at University of Oxford, he is ambitious and determined to develop and enhance the translational clinical and collaborative medical research in the region.

To find out more about the author, visit his Scientific Malaysian profile at http://www.scientificmalaysian.com/members/adliali/

References 1.

Wakefield A, Murch S, Anthony A; et al. (1998).

“Ileal-lymphoid-nodular hyperplasia, non-specific colitis, and pervasive developmental disorder in children”.

Lancet 351 (9103) : 637–41.

2.

Godlee Fiona, Smith Jane, Marcovitch Harvey.

Wakefield’s article linking MMR vaccine and autism was fraudulent BMJ 2011; 342 :c7452 3.

Immunization Safety Review Committee, Board on Health Promotion and Disease Prevention, Institute of Medicine (2004). Immunization Safety Review: Vaccines and Autism. Washington, DC: The National Academies Press. ISBN 0-309-09237-X 4.

Doja A, Roberts W (2006). “Immunizations and autism: a review of the literature”.Can J Neurol Sci 33 (4): 341–6. doi:10.1017/s031716710000528x.

5.

Lancet. 2010;375(9713):445.

6.

“Thimerosal in vaccines”. Center for Biologics Evaluation and Research, U.S. Food and Drug Administration. 2007-09-06. Retrieved 2007-10-01 7.

Baker JP (2008). “Mercury, Vaccines, and Autism: One Controversy, Three Histories”. Am J Public Health 98 (2): 244–53. doi:10.2105/AJPH.2007.113159.

PMC 2376879.

8.

Abdullah, Ahmad Badri. “Halal vaccine and the Ethical Dimension of vaccination Programmes.” Islam and Civilisational Renewal (ICR) 5.3 (2014).

9. Genzel, Y. (2015), Designing cell lines for viral vaccine production: Where do we stand?. Biotechnology Journal, 10: 728–740.

10. Hayflick, Leonard (March 1965). “The Limited in vitro Lifetime of Human Diploid Cell Strains”. Experi-

mental Cell Research 37: 614–636. doi:10.1016/00144827(65)90211-9 11. Jacobs, Characteristics of a human diploid cell designated MRC-5 (1970). Nature 277:168:247-56 1970 12. Addressing Parents’ Concerns: Do Multiple Vaccines Overwhelm or Weaken the Infant’s Immune System? Paul A. Offit, Jessica Quarles, Michael A. Gerber, Charles J. Hackett, Edgar K.Marcuse, Tobias R. Kollman, Bruce G. Gellin, Sarah Landry Pediatrics Jan 2002, 109 (1) 124-129.

13. Rothman, Kenneth J., Yinong Young-Xu, and Felix Arellano. “Age dependence of the relation between reassortant rotavirus vaccine (RotaShield) and intussusception.” Journal of Infectious Diseases 193.6 (2006): 898-898.

14. Zipprich, Jennifer, et al. “Measles outbreak—California, December 2014–February 2015.” MMWR Morb Mortal Wkly Rep 64.6 (2015): 153-154.

15. Takahashi, Saki, et al. “Reduced vaccination and the risk of measles and other childhood infections post-Ebola.” Science 347.6227 (2015): 1240-1242.

16. Blume, Stuart. “Anti-vaccination movements and their interpretations.”Social science & medicine 62.3 (2006): 628-642.

17. https://www.gov.uk/government/publications/ measles-confirmed-cases/measles-notifications-andconfirmed-cases-by-quarter-in-england-2013-to-2015

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D

A ‘Trp’ to the Land of Kyns

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espite being implicated in immune responses of a number of normal body functions as well as disease processes, the kynurenine (Kyn) pathway is known amongst researchers only if one has worked on it, or topics related to it. Worse still, if one has not done any immunology-related research, one may never hear of it although its implication in medicine is transdisciplinary in nature, extending beyond a single disease. The Kyn pathway is essentially a metabolic pathway involving the amino acid, tryptophan (Trp), which is an essential building block for proteins (see Figure 1). Through this

metabolic pathway, more than 95% of Trp is catabolized [1].

This pathway is a curious one as it has the capacity to elicit immune responses that may both protect or harm the body depending on the extent of the activation of the pathway as well as the cells it is activated in. One of the earliest documented roles of the Kyn pathway was in bacterial infections [2]. Its role in other infectious diseases such as HIV/AIDS and hepatitis, as well as malaria, has since been established [3-5]. The role of the pathway in the pathogenesis of dengue infections has also been suggested [6]. Other pathological conditions in

Felicita Fedelis Jusof

At a Glance: Breakdown of the amino acid tryptophan through the kynurenine pathway impacts a plethora of immunological processes, ranging from the starvation of invading bacteria to the induction of immunotolerance and the modulation of neurological diseases.

Figure 1: Diagram of the Kyn pathway

which the Kyn pathway has been implicated include neurological disorders such as Alzheimer’s, AIDS-related dementia and schizophrenia [7-8], as well as autoimmune disorders such as cancer, allergy, arthritis and asthma in which it has been gaining traction [9-13].

The Kyn pathway also contributes to the immunotolerance observed in pregnancy, where the activation of this pathway suppresses the mother’s local immunity in the placenta, preventing the mother’s immunity from rejecting her foetus, which would otherwise be recognised as a foreign

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Different Journeys:

body [14]. This discovery was monumental in contributing to our understanding of how the pathway modulates immunity.

A similar mechanism of immunotolerance has been documented in cases of tra ns pla ntati o n , wh e re induction of Trp-catabolic enzyme rendered protection from organ rejection in cases of liver transplantation [15].

The ability of this pathway to induce immunosuppression also contributes to a poorer outcome in several types of cancers as it allows the tumour to persist in the host [16-18].

One of the first mechanisms proposed through which the Kyn pathway exerts its immunological response involves the manipulation of Trp availability to cells and pathogens in the m i c ro e n v i ro n m e n t .

As Trp is a food source for pathogens in infections, the activation of this pathway during microbial infections strategically depletes Trp, effectively sta rvi ng a nd eventually eliminating the pathogens [2]. The alternative

mechanism through which the Kyn pathway exerts an i m m u n o m o d u l ato r y effect is either through the suppression of immune cells known as T cells or through the generation of metabolites that can act on neurons [1921]. These metabolites may either be neuroprotective or neurotoxic.

The first step in this pathway is the conversion of Trp to kynurenine (Kyn). This step of the pathway can be catalysed by three enzymes, tryptophan 2,3 dioxygenase (TDO), indoleamine 2,3 dioxygenase 1 (IDO1) and the most recently discovered IDO2. While the three enzymes catalyse the same reaction, their distribution and role in tissues differ. The longest known enzyme of the three, TDO is expressed constitutively in high amounts in the mammalian liver and its best-understood role is in the regulation of dietary Trp [22].

However, as it also is expressed constitutively albeit at low levels in other tissues such as epididymis, testis, pancreas

and heart, an alternative role for TDO in these tissues has been considered [23,24]. IDO1, on the other hand, which is expressed constitutively in relatively low levels in the epididymis, intestine and placenta, is induced in various cells and tissue types in the presence of inflammatory stimuli, namely interferon gamma (IFN ), TNF and LPS [14, 25-27].

The least understood of the three is the isozyme of IDO1, IDO2. Based on the most recent phylogenetic study, it is shown that IDO1 is more similar to the ancestral IDO gene and that IDO2 arose from duplication of the ancestral gene that occurred in early vertebrate evolution with IDO1 being eventually lost in a number of the lower vertebrates [28]. Despite both IDO homologues in mammals possessing striking genomic structural similarities and Trp-catabolic properties, the enzymatic activity of IDO2 is relatively low and its substrate specificity differs from IDO1, leading to speculations that it

Pathogens, Take a Hike!

is a food source for pathogens in infections, “ Trp the activation of this pathway ... depletes Trp, effectively starving and eventually eliminating the pathogens

“

all cell types express enzymes “ not downstream in the [kyns] pathway

may possess functions aside from its ability to metabolise Trp. The expression of IDO2 protein has been confirmed only in the mammalian liver while its presence in other tissues are ambiguous, as its RNA has been detected in kidney, brain, colon as well as in epididymis [29,30]. IFN inducible IDO2 was detected in antigen-presenting dendritic cells, macrophages, astrocytes and mesenchymal stem cells [31-33]. In terms of clinical significance, IDO2 has been implicated in some forms of cancer [32-34]. However, more recently, it was also reported to be involved in the pathogenesis of rheumatoid arthritis and contact hypersensitivity [35,36].

The generation of neuroactive metabolites through the Kyn pathway are known to contribute to the progression of some of the diseases it is implicated in.

These metabolites include 3-hydroxykynurenine,

kynurenic acid, quinolinic acid and picolinic acid. Of these, kynurenic acid (KA) has been reported to be neuroprotective [37,38], whereas downstream metabolites of the pathway, 3-hydroxykynurenine, quinolinic acid (QA) and picolinic acid (PA) have been shown to exert neurotoxic effects [19,39].

Fascinatingly, not all cell types express enzymes downstream in the pathway.

In cells where enzymes downstream in the pathway are absent or present in very minute levels, the Kyn pathway often does not metabolise substrates beyond kynurenine.

For example, in endothelial cells, Trp is catabolised to kynurenic acid constitutively while generating both kynurenine and kynurenic acid when primed with IFN [40,41].

However, endothelial cells do not possess the capacity to synthesise kynurenine metabolites downstream

in the pathway, such as quinolinic or picolinic acid, as the downstream enzymes of the pathway are either inactive, absent or present in levels too low to exert an effect.

Human foetal brain cultures and lung cells also exhibited similar features [42,42]. On the other hand, macrophages, monocytes, microglia and liver cells were reported to have a high level of quinolinic acid, indicating the capacity of these cells to metabolise substrates downstream of the pathway [40, 42,43].

H

ow is this pathway relevant to the Malaysian context, one may ask.

The involvement of this pathway in immune responses of va ri o us d iseases reflects the potential of this pathway to help us better understand more than just one immuneand inflammation-related disease. The major health

“

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burden of Malaysia according to the National Strategic Plan for Non-communicable Disease, Ministry of Health Malaysia, is non-communicable diseases which include cardiovascularrelated illness, diabetes and cancer [44]. In addition to these non-communicable diseases, infectious diseases, namely dengue and HIV/AIDS, rounds up the eight leading disease burdens of Malaysia.

Strikingly, the Kyn pathway has been implicated in all of the above mentioned diseases.

It has been suggested that the balance between the neuroprotective and neurotoxic metabolites generated by the pathway could be used for early diagnosis, prognosis and intervention in diseases in which Kyn pathway is involved.

It would be interesting to investigate if the balance between the neuroprotective and neurotoxic metabolites influences the progression of these diseases and wh eth er i n h i biti ng th e generation of

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the neurotoxic metabolites, or administering neuroprotective metabolites could be a possible approach to intervention. The most exciting thing about deepening our understanding of this pathway is the potential to apply the knowledge in more than just one disease.

T h e re m a r k a b l e t h i n g about research is that the solutions are never quite straightforward and often enough, the direction a study takes can be unpredictable and surprising. The best of research projects begins with a very simple yet logical and exciting idea which, with further persistent probing, yields valuable information for our understanding of the human body and disease processes.

Perhaps this is the vision that the understanding of kynurenine pathway and the key players in it calls us to. That one day, a better u n d e rsta n d i n g of

th is pathway may help us with the diagnosis and treatment of more than one physiological or pathological condition.

About The Author Dr. Felicita Fedelis Jusof considers herself a newbie in the academic world. Having completed her doctoral degree in the University of Sydney in the year 2015, she returned to serve the very people who funded her doctoral degree overseas (Malaysian taxpayers) through her service to the University of Malaya. Although she loves the Big Bang Theory, she sees herself as an ordinary person who enjoys normal things like The Voice, amazing Asian food and Ed Sheeran. Despite this normalcy, she can sometimes be accused of being an adrenaline junkie who enjoys bungee jumping, bridge climbing and slingshot rides to name a few.

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Sanni LA, Thomas SR, Tattam BN, Moore DE, Chaudhri G, Stocker R, et al. Dramatic changes in oxidative tryptophan metabolism along the kynurenine pathway in experimental cerebral and noncerebral malaria.

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10. Uyttenhove C, Pilotte L, Theate I, Stroobant V, Colau D, Parmentier N, et al. Evidence for a tumoral immune resistance mechanism based on tryptophan degradation by indoleamine 2,3-dioxygenase. Nature medicine. 2003;9(10):1269-74.

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12. Hayashi T, Beck L, Rossetto C, Gong X, Takikawa O, Takabayashi K, et al. Inhibition of experimental asthma by indoleamine 2,3-dioxygenase.

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A tryptophan metabolite, kynurenine, promotes mast cell activation through aryl hydrocarbon receptor. Allergy.

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14. Munn DH, Zhou M, Attwood JT, Bondarev I, Conway SJ, Marshall B, et al. Prevention of allogeneic fetal rejection by tryptophan catabolism.

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15. Lin YC, Goto S, Tateno C, Nakano T, Cheng YF, Jawan B, Kao YH, Hsu lW, Lai CY, Yoshizato K & Chen C l. 2008. Induction of indoleamine 2,3-dioxygenase in livers following hepatectomy prolongs survival of allogeneic hepatocytes after transplantation. Transplant Proc, 40, 2706-8.

16. Ino K, Yoshida N, Kajiyama H, Shibata K, Yamamoto E, Kidokoro K, et al. Indoleamine 2,3-dioxygenase is a novel prognostic indicator for endometrial cancer. British journal of cancer. 2006;95(11):1555-61.

17. Okamoto A, Nikaido T, Ochiai K, Takakura S, Saito M, Aoki Y, et al.

Indoleamine 2,3-dioxygenase serves as a marker of poor prognosis in gene expression profiles of serous ovarian cancer cells. Clinical cancer research : an official journal of the American Association for Cancer Research. 2005;11(16):6030-9.

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Godin-Ethier J, Hanafi LA, Piccirillo CA, Lapointe R. Indoleamine 2,3-dioxygenase expression in human cancers: clinical and immunologic perspectives. Clinical

cancer research : an official journal of the American Association for Cancer Research. 2011;17(22):6985-91.

19. Heyes MP, Saito K, Crowley JS, Davis LE, Demitrack MA, Der M, et al. Quinolinic acid and kynurenine pathway metabolism in inflammatory and non-inflammatory neurological disease. Brain. 1992;115 ( Pt 5):124973.

20. Mellor AL, Munn DH. Tryptophan catabolism and T-cell tolerance: immunosuppression by starvation?

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Mellor AL, Munn DH. IDO expression by dendritic cells: tolerance and tryptophan catabolism.

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22. Knox WE, Auerbach VH. The hormonal control of tryptophan peroxidase in the rat. The Journal of biological chemistry. 1955;214(1):30713.

23. Britan A, Maffre V, Tone S, Drevet JR. Quantitative and spatial differences in the expression of tryptophan-metabolizing enzymes in mouse epididymis. Cell and tissue research. 2006;324(2):301-10.

24. Suzuki S, Tone S, Takikawa O, Kubo T, Kohno I, Minatogawa Y. Expression of indoleamine 2,3-dioxygenase and tryptophan 2,3-dioxygenase in early concepti.

The Biochemical journal. 2001;355(Pt 2):425-9.

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Hayaishi O. Induction of indoleamine 2,3-dioxygenase in mouse lung during virus infection. Proceedings of the National Academy of Sciences of the United States of America.

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26. Dai X, Zhu BT. Indoleamine 2,3-dioxygenase tissue distribution and cellular localization in mice: implications for its biological functions. J Histochem Cytochem.

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27. Higuchi K, Hayaishi O. Enzymic formation of D-kynurenine from D-tryptophan.

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28. Ball HJ, Jusof FF, Bakmiwewa SM, Hunt NH, Yuasa HJ. Tryptophancatabolizing enzymes - party of three.

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Fukunaga M, Yamamoto Y, Kawasoe M, Arioka Y, Murakami Y, Hoshi M, et al. Studies on tissue and cellular distribution of indoleamine 2,3-dioxygenase 2: the absence of IDO1 upregulates IDO2 expression in the epididymis. J Histochem Cytochem. 2012;60(11):854-60.

31. Croitoru-Lamoury J, Lamoury FM, Caristo M, Suzuki K, Walker D, Takikawa O, et al. Interferon-gamma regulates the proliferation and differentiation of mesenchymal stem cells via activation of indoleamine

2,3 dioxygenase (IDO). PLoS One. 2011;6(2):e14698.

32. Lob S, Konigsrainer A, Zieker D, Brucher BL, Rammensee HG, Opelz G, et al. IDO1 and IDO2 are expressed in human tumors: levobut not dextro-1-methyl tryptophan inhibits tryptophan catabolism.

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Metz R, Duhadaway JB, Kamasani U, Laury-Kleintop L, Muller AJ, Prendergast GC. Novel tryptophan catabolic enzyme IDO2 is the preferred biochemical target of the antitumor indoleamine 2,3-dioxygenase inhibitory compound D-1-methyl-tryptophan.

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34. Witkiewicz AK, Costantino CL, Metz R, Muller AJ, Prendergast GC, Yeo CJ, et al. Genotyping and expression analysis of IDO2 in human pancreatic cancer: a novel, active target. J Am Coll Surg.

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35. Merlo LM, Pigott E, DuHadaway JB, Grabler S, Metz R, Prendergast GC, et al. IDO2 is a critical mediator of autoantibody production and inflammatory pathogenesis in a mouse model of autoimmune arthritis.

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36. Metz R, Rust S, Duhadaway JB, Mautino MR, Munn DH, Vahanian NN, et al. IDO inhibits a tryptophan sufficiency signal that stimulates mTOR: A novel IDO effector pathway targeted by D-1-methyl-tryptophan.

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Snake antivenoms: Science, Values & Challenges

40. Owe-Young R, Webster NL, Mukhtar M, Pomerantz RJ, Smythe G, Walker D, et al. Kynurenine pathway metabolism in human blood-brainbarrier cells: implications for immune tolerance and neurotoxicity. J Neurochem. 2008;105(4):1346-57.

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42. Heyes MP, Achim CL, Wiley CA, Major EO, Saito K, Markey SP. Human microglia convert l-tryptophan into the neurotoxin quinolinic acid. The Biochemical journal. 1996;320 ( Pt 2):595-7.

43. Heyes MP, Chen CY, Major EO, Saito K. Different kynurenine pathway enzymes limit quinolinic acid formation by various human

Written by Tan Choo Hock

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At a glance ites from venomous snakes can lead to snakebite envenomation (not poisoning), and antivenom is the only definitive therapy to date.

Antivenoms used in current clinical practices are derived from antibodies of animals (e.g. horses) that have been immunised with one or a mix of snake venoms. However, the production of these biologics is highly costly, and there is no

B

eared or loved, reviled or revered, the snake is a significant yet mystical subject throughout human civilisations. There are more than 3,000 snake species in the world. About 600 species of these are venomous and over 200 are considered medically important – their bites can cause envenomation, a condition characterised by the development of systemic or local toxicity due to the venom’s effect [1]. Snakebite envenomation is prevalent in many tropical and subtropical countries, affecting mainly the poor rural populations.

Worldwide, the annual mortality rate has been estimated to be around 100,000 deaths [2]. Unfortunately, the persistent underreporting/underestimation of its true epidemiology has made it the most neglected disease condition, and ironic enough, this is further aggravated by its removal from the WHO-list of neglected tropical diseases in 2015.

F

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universal antivenom, as the effectiveness is limited by the different snake species and their geographical locality.

The production and use of antivenoms can be optimised by unravelling the complexity of venoms, especially their immunogenicity and the dynamics-kinetics of venomantivenom interplay.

anaging snakebite envenomation takes on multiple steps; antivenom remains the definitive and etiological therapy. Antivenom is perhaps one of the oldest “antibody-based treatments”, pioneered by Albert Calmette (recalling: BCG vaccine) in the late 19th century, which principle still applies today in the treatment of snakebites – while immunotherapy against infections (e.g. diphtheria) has basically been replaced by vaccination. Vaccination against snake venom is still controversial and has not been proven effective in the earlier trial.

Antivenoms thus remain important, relevant, and life-saving to date. They are derived from animals (typically horses) that have been hyperimmunised with venom(s) of a single species (thus raising monovalent/monospecific antivenom) or several species (thus raising polyvalent/ polyspecific antivenom).

Current technologies process antivenoms in three main forms: whole immunoglobulin, the antigen-binding fragment (Fab), or the dimeric form F(ab’)2. They act by binding

to toxins in the venom, forming inactive immunocomplexes that can be eliminated through phagocytosis. Clinically, this neutralises the toxicity caused by venom, either by reversing or halting the progression of venom toxic effect. wo all-time pertinent challenges surrounding the use of antivenom are: (1) the effectiveness or quality of antivenom; (2) the availability of antivenom [3]. There is no universal antivenom or antidote, despite claims by some traditional medicine providers. The use of antivenom is species-specific

T

to ensure effective treatment, and this is attributed to the vast variations in the composition of venoms from different or even a same species. Venom variations can manifest as differences in the subtypes of toxins, their relative abundances, peptide sequences and even the epitopes.

The ramification of venom variation is far-reaching as it can lead to discrepancies in clinical presentation and syndrome progress, as well as suboptimal response to antivenom especially when the antivenom used was produced for a different species or from a distant country [3-5].

This explains the importance

M

Figure 1: A marine (left) and an arboreal (right) snake were identified and carefully

milked for their venoms by the author. Snake species identity and their habitat location are important information for Dr Tan’s research, as venom toxins can vary significantly among snakes.

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of species recognition in envenomation cases, as inappropriately administered antivenom would not only be futile in rescuing the victim but also increases the medical cost and exposing the patient to unnecessary risk of adverse effects of antivenom, for instance, hypersensitive reactions that can be fatal. Nevertheless, certain venoms share similar toxin profiles and hence may be cross-neutralised by a specific antivenom - for example, the monocled cobra antivenom manufactured in Thailand is effective in treating neurotoxic envenomation by the equatorial spitting cobra in Malaysia [6]. For pit viper envenomations, venoms of the Malayan pit viper and the Asiatic lance-headed pit vipers of Trimeresurus complex , although exhibiting similar hemotoxic effects, have substantial antigenic differences in toxins that the respective specific antivenom does not cross-neutralise each other well, therefore warranting the use of different specific antivenoms [6]. n the other hand, antivenom manufacturing is highly costly and technically demanding. Several antivenom plants closed in recent years ostensibly for the high production cost and limited market demand by region. This

O

further aggravates the already inadequate supply of antivenoms in many regions of the world. Note, although antivenoms are generally in high demand globally, unlike any other generic medicines, their formulations are limited by species and locality of concern, practically making each product some kind of an orphan drug – so, to produce or not to produce an expensive treatment to cater for a small market (limited by different snake species and geography) for consumers who are mostly poor – this is a serious but realistic question to answer. In Malaysia, the local antivenom production facility has long been closed down, and the country has been relying on imported antivenoms for the past few decades. This is a solution that many countries have adopted, however, the quality and effectiveness of these imported antivenoms, which were raised from different species (although may be closely related) from foreign lands, must be first rigorously tested at the laboratory level to provide insights into the suitability for use in local cases. In recent years, with the available emerging laboratory evidence and favourable clinical observations, Malaysia is moving towards the use of some antivenoms produced in Thailand for the common and closely

Figure 2: Feared or loved, but rarely understood

– venomous snakes do not prey on humans and envenomations are usually results of unpleasant encounters between humans and snakes – feeling threatened, the fangs (in this picture, a pit viper with its front fangs shown) and the venom channelled through them become the best bio-weapon for the snake. Venoms leave an impact on human lives in a paradox: the side that can kill and destroy, and the other side that serves as a rich pool of novel bioactive compounds, from which drug discovery can be made.

Figure 3: Examples of local “haemotoxic” snakes: Malayan pit viper (left) and Cameron

Highland pit viper (right). Envenomation by these species can cause similar pattern of bleeding disorder, but requires different kind of monovalent antivenom for effective treatment due to the differences in the molecular makeup of their venom toxins.

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related species shared by the two countries in close proximity [5-7]. Patients envenomed by some of the species in Malaysia can benefit from cross-neutralisation conferred by certain Thai antivenoms, due to the conserved antigenicity of principal toxins in the different lineages. n this context, it should be noted that Professor Tan Nget Hong from the University of Malaya (UM) has led venom research in Malaysia for almost 40 years. Established recently, the Venom and Toxin Research Laboratory (VTRL, Faculty of Medicine, UM), which the author of this article is co-leading, has been continuously researching on snake venoms and antivenoms to unravel the complexity of snake venom toxins and the myth of venomantivenom interactions. Findings that have been published include the transcriptomics and proteomics of several major or exotic species in the region [4, 8-11]. Coupled with functional, in vitro/in vivo characterisation of the venoms and their antivenom neutralisation profiles, the group is determined to provide detailed insights into the pathophysiology of envenomation and how treatment can be optimised - this aspect

I

concerns the community most. Recently, the group has also identified some of the limiting factors in governing the immunogenicity of venom used in the production of antivenoms; research is now on-going to strategise an approach that can overcome the limitation of antivenom effectiveness [12-15]. It is hoped that the findings will be translated into a trans-border, pan-region collaborative research that will eventually seek the production of an affordable, “broadspectrum” antivenom with high potency and multiple species coverage. In addition, the data generated by the group may be useful for future studies in drug discovery, biodiversity and wildlife conservation. Thus far, the group has unveiled the functional venom proteomes (with or without venomgland transcriptomes) of several important lineages, including the monocled cobras (from Malaysia, Thailand and Vietnam), king cobra (Malaysia), hump-nosed pit viper and Russell’s viper (Sri Lanka), equatorial spitting cobra (Malaysia), beaked sea snake (Malaysia) and the exotic Malayan blue coral snake (Malaysia) [4; 8-11].

Ab o ut The Au thor : Dr. Tan Choo Hock

(MBBS, PhD) is a Senior Medical Lecturer in Pharmacology from UM. His research interests span a wide range of topics in toxinology, including molecular and functional characterisation of venoms, antivenom pharmacology, and the “-omics” of venomous species. He enjoys passionately his research activities, from collecting samples in the wild to experiments conducted in the laboratory. He can be reached at tanchoohock@gmail.com. Find out more about the author by visiting his profile at: http://www.scientificmalaysian.com/members/tanchoohock/

REFERENCES [1] WHO. (2010). Guidelines for the Management of Snake-bites. World Health Organization: Regional Office for South-East Asia.

[2] Kasturiratne, A., Wickremasinghe, A. R., de Silva, N., Gunawardena, N. K., Pathmeswaran, A., Premaratna, R., . . . de Silva, H. J. (2008). The global burden of snakebite: a literature analysis and modelling based on regional estimates of envenoming and deaths. PLOS Medicine, 5(11):e218.

[3] Williams, D. J., Gutierrez, J. M., Calvete, J. J., Wuster, W., Ratanabanangkoon, K., Paiva, O., . . . Warrell, D. A. (2011). Ending the drought: new strategies for improving the flow of affordable, effective antivenoms in Asia and Africa. Journal of Proteomics, 74(9):1735-1767.

[4] Tan, K. Y., Tan, C. H., Fung, S. Y., & Tan, N. H. (2015). Venomics, lethality and neutralization of Naja kaouthia (monocled cobra) venoms from three different geographical regions of Southeast Asia. Journal of Proteomics, 120:105-125.

[5] Tan, K. Y., Tan, C. H., Sim, S. M., Fung, S. Y., & Tan, N. H. (2016).

Geographical Venom Variations of the Southeast Asian Monocled Cobra (Naja kaouthia): Venom-Induced Neuromuscular Depression and Antivenom Neutralization. Comparative Biochemistry and Physiology - Part C: Toxicology and Pharmacology, (In press). doi: 10.1016/j.cbpc.2016.03.005.

[6] Tan, C. H., Tan, N.H. (2015).

Toxinology of Snake Venoms: The Malaysian Context. In P. Gopalakrishnakone, H. Inagaki, A. K. Mukherjee, T. R. Rahmy, & C.-W. Vogel (Eds.), Snake Venoms (pp. 1-37): Springer Netherlands.

[7] Leong, P. K., Tan, C. H., Sim, S. M., Fung, S. Y., Sumana, K., Sitprija, V., & Tan, N. H. (2014). Cross neutralization of common Southeast Asian viperid venoms by a Thai polyvalent snake antivenom (Hemato Polyvalent Snake Antivenom).

Acta Tropica, 132:7-14.

[8] Tan, C. H., Tan, K.Y., Fung, S.F., Tan, N.H. . (2015). Venom-gland

transcriptome and venom proteome of the Malaysian king cobra (Ophiophagus hannah). BMC Genomics, 16:687 [9] Tan, C. H., Tan, K. Y., Lim, S. E., & Tan, N. H. (2015). Venomics of the beaked sea snake, Hydrophis schistosus: A minimalist toxin arsenal and its cross-neutralization by heterologous antivenoms. Journal of Proteomics, 126:121-130.

[10] Tan, N. H., Fung, S.Y., Tan, K.Y., Yap, M.K.K., Gnanathasan, C.A., Tan, C.H. (2015). Functional venomics of the Sri Lankan Russell’s viper (Daboia russelii) and its toxinological correlations.

Journal of Proteomics, 128:403-423.

[11] Tan, C. H., Fung, S. Y., Yap, M. K., Leong, P. K., Liew, J. L., & Tan, N. H. (2016). Unveiling the elusive and exotic: Venomics of the Malayan blue coral snake (Calliophis bivirgata flaviceps). Journal of Proteomics, 132:1-12.

[12] Tan, C. H., Tan, N. H., Tan, K. Y., & Kwong, K. O. (2015). Antivenom crossneutralization of the venoms of Hydrophis schistosus and Hydrophis curtus, two common sea snakes in Malaysian waters.

Toxins (Basel), 7(2):572-581.

[13] Wong, K. Y., Tan, C.H., Tan, N.H. (2016). Venom and Purified Toxins of the Spectacled Cobra (Naja naja) from Pakistan: Insights into Toxicity and Antivenom Neutralization. The American Journal of Tropcal Medicine and Hygiene, (In press). doi:doi:10.4269/ajtmh.15-0871.

[14] Tan, K. Y., Tan, C. H., Fung, S. Y., Tan, N. H. (2016). Neutralization of the principal toxins from the venoms of Thai Naja kaouthia and Malaysian Hydrophis schistosus: Insights into toxin-specific neutralization by two different antivenoms. Toxins (Basel), 8:86. doi:10.3390/toxins8040086.

[15] Ratanabanangkoon, K., Tan, K. Y., Eursakun, S., Tan, C. H., Simsiriwong, P., Pamornsakda, T., Wiriyarat, W., Klinpayom, C., Tan, N. H. (2016) A Simple and Novel Strategy for the Production of a Pan-specific Antiserum against Elapid Snakes of Asia.

PLOS Neglected Tropical Diseases, 10(4):e0004565. doi: 10.1371/journal. pntd.0004565.

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Disclaimer:

In a wasteland, and at a time where vaccination was illegal, a boy named Mikhail was destined to save the last piece of humanity.

The following article is a fictional piece that may or may not be scientifically accurate. Reading with discretion is advised.

The Imitation

By Nur Atikah Abdullah, Charles George Gajim & Seti Faezah Rosli

At a Glance: The idea of vaccination has always been heavily debated throughout the past few decades. Back in the year of 2016, more people started to reject the idea of vaccination, claiming that immunised children were unhealthy. By the year 2050, the vaccination was permanently banned by governments all over the world. However, a little boy named Mikhail was raised and secretly immunised by his parents with a vaccine called AFOV (Allfor-One-Vaccine). It is the only cure against a new emerging disease called SIC (Severe Intravascular Croatoan). By extracting the antibodies from his blood, replicating it and then silently distributing it to those in need, Mikhail vowed to save the last of mankind.

It started in 2016, when more and more parents refused to get their children vaccinated, claiming that immunised children were unhealthy and at risk of developing chronic diseases later in life. In a mere three months, the propaganda had spread to Asia, including Malaysia. In the year 2050, 98% of parents worldwide were against vaccination. Petitions were signed globally and governments were forced to delegalise vaccines in the name of democracy.

By 2051, epidemics began to occur.

Born to a genius scientist, and a remarkable ex-navy Lieutenant, Mikhail’s parents were a part of the 2% of the human population that still practiced vaccination.

Knowing what would happen, Mikhail’s father invented a super vaccine — AFOV, also known as ‘All-for-One-Vaccine’ — to fight off all communicable diseases. When Mikhail was born in 2050, he was vaccinated with AFOV by his father, making him immune to the worldwide epidemics that had begun to plague the world’s population.

Twenty years later, the world had changed dramatically. Things had gone from bad to worse and many people who were born after the year 2050 had succumbed to epidemics. The deadliest of them all was a new plague called Severe Intravascular Croatoan or famously known as SIC, which was wiping out the world population every day. By the time someone realised what was happening, it was too late. There just weren’t adequate resources to formulate a cure for it, let alone a vaccine.

Every nation in the world was hit by the plague. There was no food to eat, no jobs to

go to, and no entertainment to cheer anyone up. The only entertainment available was the news, which only existed to inform the citizens about the current death toll. People were quarantined inside their own homes.

It was Armageddon.

“I’m next!” Mikhail was frantic. He had just witnessed his neighbour’s son, a boy around his age dropping dead right before his eyes through his bedroom window.

“We will not let anything happen to you,” consoled Diana, Mikhail’s mother.

“Everyone I know is dead,” Mikhail gasped.

“I’m going to die soon, just like them.”

“Son, there is something you should know,” Mikhail’s father said calmly, seemingly unaffected by his son’s outburst.

Learning the truth about himself was by far the most shocking event in Mikhail’s life.

According to his father, the SIC plague was the least of his worries. Thanks to a vaccine called AFOV, created in secret by Mikhail’s father and given to him as a child, Mikhail was resistant to the plague and other diseases as well. He was not immortal by any means, but at least, he was safe.

Nevertheless, he was restless. He was certainly safe, but what about others? The plague began with fever, but the symptoms progressed dramatically in the victims. In a matter of time, the blood vessels would burst, mimicking a haemorrhagic stroke.

Some took months to reach the final stage, while the unlucky ones only had days.

Those who were infected never made it to another year. He couldn’t bear seeing any more people die. He couldn’t just sit and do nothing while others fought for their lives each day.

He had to think of something.

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He had to do something.

“Are you sure you want to do this?” Diana appeared hesitant.

“This is the only way we can stop this, Mom. We can end this once and for all,” replied Mikhail.

“You have to be cautious. If you were to be caught, you would be their lab rat, you would be a precious test subject for their everlasting experiments,” said his Dad, voicing his doubts.

“With all the physical activity training I’ve done with mom since I was four to keep me fit and athletic, and the homeschooling from you, Dad, I will be fine. I am the ultimate combination of brain and brawn, aren’t I?” Mikhail said confidently.

“What is your plan?”

“Dad, you can extract the antibodies from my blood, right? We can synthesise it and distribute it to everyone,” Mikhail said. “Carefully and stealthily,” he added to reassure his parents.

Mikhail began his mission in plain sight. He didn’t use any costume to do his job. He wasn’t planning to be a superhero wannabe but he did do his best to conceal his identity. Every day after sundown, he would visit those who were infected and give them a vial of ready-made antibodies.

With it, people were progressively getting better. They started calling him a vigilante since he wasn’t exactly a law-abiding citizen. He didn’t mind being a vigilante for the rest of his life as long as he could help people.

It took years before the SIC plague was completely eradicated.

Nonetheless, people finally learned their lesson about the importance of vaccination.

ABOUT THE AUTHORS: Nur Atikah Abdullah, Charles George Gajim, and Seti Faezah Rosli are Bachelor

of Biomedical Science final year students at Management and Science University (MSU), Shah Alam. All three of them have also previously studied Diploma in Medical Laboratory Technology. Their final year research projects comprise many research fields such as anatomy, pharmacology, and microbiology. Nur Atikah enjoys reading and writing fiction, Charles is into sports especially basketball, while Seti Faezah likes cooking. Find out more about the authors by visiting their profiles at: http://www.scientificmalaysian.com/members/abdullahika/ http://www.scientificmalaysian.com/members/c90kidz04/ http://www.scientificmalaysian.com/members/iradiana/

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Our immune system is naturally gifted with remarkable specificity, potency and memory. So far, no pharmacological treatment for any diseases could possibly provide comparable level of safety, efficacy and lasting effect as the human immune response. In the treatment of cancer, after the primary therapies – i.e. surgery, radiotherapy and chemotherapy – immunotherapy is being explored as a fourth option, especially for advanced stage cancers. To date, several kinds of novel immunotherapeutics, including cancer peptide vaccines, dendritic cell vaccine, immune checkpoint blockade and adoptive cell therapy are increasingly being introduced in clinics. This article peeks into the brief history of immune-stimulation, state-of-the-art advances, as well as some limitations of these approaches.

Of Spontaneous Cancer Regression and Coley’s Toxins

Teaching Your Immune System To Fight Cancer

By Litt-Yee Hiew

In the unavailing search for a cancer cure, it has been interestingly noted that certain cancers could spontaneously regress.

Albeit rare and only anecdotally reported, kidney, brain (neuroblastoma), uterine and skin cancer are amongst the four frequent cancers associated with cancer regression, according to a review of 176 published cases from 1900 to 1960 [1].

The sudden disappearance of not only the primary tumour but also their metastatic foci has been hypothesised to be a result of unexpected activation of the immune system leading to the recognition of non-selfproteins and subsequently the destruction of these cancer cells [2].

Indeed, Dr. William Coley, an orthopaedic surgeon, was one of the first physicians to relate the concept of immune system and its interface with cancer by leveraging on the serendipitous discovery of the record of an immigrant patient with a recurring sarcoma that spontaneously regressed following an extended postoperative surgical wound infection with Streptococcus pyogenes [3].

It was documented that the tumour, despite only partially removed, shrank over several months and finally disappeared completely.

Following discharge, he remained cancer-free in the subsequent reviews. Penned in his paper dated back in 1893 [4], Coley attributed the patient’s unexpected cure to the infection which likely

led to stimulation of some type of immune responses.

In an attempt to rationalise his hypothesis, series of experiments to deliberately infect cancer patients with S. pyogenes were carried out and resulted, at times, in failed infections or even death. Eventually, a version containing a mixture of killed S. pyogenes and Serratia marcescens was developed into what became known as the ‘Coley’s toxins’. While remarkable recoveries were documented in several cases of advanced diseases [5], its use eventually faded for various reasons.

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Harnessing the Natural Capacity of the Immune System

Cancer Immunotherapy: From Experimental to Mainstream

However, it was not until the middle of 20th century when the term “immunesurveillance” was coined by immunologists Lewis Thomas and Macfarlane Burnet [6, 7].

Accordingly, the host immune system is deemed capable of constantly monitoring and blocking cancerous cell development by detecting mutated cells and eliminate them through various mechanisms.

Nonetheless, with emerging mutations, some may incidentally evade immunosurveillance and continue to expand. Such consequent development of resistant mutant selected by the continuous pressure from the host immune system are referred to as “immune-

Over the years, intense research effort to identify immunogenic targets recognisable by the T-cells has identified several human tumour-associated antigens (TAAs) such as cancer-testis (CT) antigens.

Like the predecessors of existing immunotherapies, cancer immunotherapy was in favour of the humdrum concept of reinforcing the host immune response to eliminate cancer cells and produce lasting immunity [11]. It, therefore, seems logically sound to evoke T-cell responses against these tumour antigens through vaccination or similar mechanisms. One option for developing vaccines for infectious diseases includes using the inactivated form of pathogen to stimulate immune response. However, when a similar approach was employed to make tumour vaccines, it proved ineffective. One prominent example is the whole-cell melanoma vaccine known as Canvaxin which, despite a seemingly promising phase II studies [12], revealed no benefit in the subsequent phase III testing [13], ultimately leading to its discontinuation [14]. When such powerful-but-blunt approach failed, it led to the realisation that targeting

editing” [8]. In other words, cancer that present as clinically detectable mass are likely to have progressed beyond the initial stage of carcinogenesis that render them capable of evading the imposed immune equilibrium to achieve invasion and metastasis.

At its core, the human immune system is composed of both innate and adaptive immunity. Phagocytes and natural killer cells are effectors of innate immunity which recognise target antigens in a non-specific manner.

Cytotoxic T-cells (CTLs) and antibodies, on the other hand, are effectors of cellular and humoral immunity,

“..with

respectively, and function in an antigen-specific manner.

Although various innate and adaptive immune cells contribute to antitumour immunity, current available evidences strongly suggest T-cell responses specific to tumour antigens can mediate spontaneous tumour clearance [9, 10]. To evoke T-cell activation, two signals are indispensable: one of which is the signal through T-cell receptor (TCR) induced by the complex of antigenic peptide and major histocompatibility complex while the other is through surface molecules termed stimulatory co-receptors, such as CD28, 4-1BB and OX-40.

emerging

mutations,

evade immunosurveillance and some may incidentally

continue to expand.”

tumour-specific antigens that are solely expressed on the cancer is crucial. Though this ideal is rarely achieved, certain TAA, e.g. CT antigens and mutated antigens were found to be expressed only in certain cancer cells and are therefore plausible target as well [15]. Given the propensity of viruses to efficiently induce CTL production, these antigens were delivered using selected recombinant viral vectors and adjuvants. One such example is PROSTVAC, a pox virus-based prostate cancer vaccine containing prostate-specific antigen [16] which demonstrated an improved overall and median survival in phase II testing in advanced stage patients [17] and is currently tested in an ongoing phase III trial [18].

Dendritic cell-based (DCbased) vaccine is another antigen-specific approach but with the advantage of bypassing the vectored delivery step. The DCs are presented to the antigen directly ex vivo and, following priming, readministered back into the patient.

Notably, the only therapeutic cancer vaccine that has been licensed for clinical use so far is the DC-based sipuleucel-T (Provenge®) for used in

the treatment of advanced prostate cancer [19].

Specifically, sipuleucel-T involves autologous cell transplantation of peripheral blood monocytes primed with a fusion protein consisting of recombinant prostate acid phosphatase (another TAA expressed in prostate tumour cells), as well as granulocyte-macrophage colony-stimulating factor.

On the other hand, mutated antigens, also known as neoantigens, refer to antigens derived from tumourspecific genomic DNA mutations. Because they possess epitopes that are specific to individual tumour [20], hence the induction of neoantigen-specific effector T-cells may be less affected by T-cell tolerance compared with non-mutated self-antigens.

Notably, a recent publication has revealed an exciting finding that complex tumours with multiple mutations was found to have an increased chance of being spotted by the immune system [21].

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Overcoming the Inherent Immune Resistance in Established Cancers Vaccines aside, alternative approaches include genetically engineering patient’s own immune cells via tumourinfiltrating lymphocytes therapy or chimeric antigen receptor T-cells therapy to directly target cancer cells [22]. While some cancers demonstrated satisfactory clinical response [2327], they are relatively uncommon [28]. In other words, it appears that these potent immune cells are switched off by some tumour defence mechanisms. Indeed, the immunosuppressive condition in the tumour microenvironment is amongst the most crucial factors that account for inef fectiveness even when the therapeutic

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balance has a significant stimulation-dominant side, simply because providing adequate stimulatory cosignals to exceed the heavy inhibitory conditions in the tumour microenvironment is particularly challenging without int r o d u c ing significant adverse effects to the patient.

that facilitate immune escape. In order for tumours to grow, the immune system is often prevented from mounting an effective antitumour response. In the tumour microenvironment, this cancer-specific milieus are formed by several cellular populations, including tumour cells, stromal cells and infiltrating immune cells.

The current understanding Accordingly, this response of tumour immunology can be broadly categorised proposes that tumours into a few phenotypes [29]. could generate an Notably, the T-cell infiltrated immunologically-restrained phenot yp e has b e en milieu, typically by interfering demonstrated to confer solid with any one of the following tumour a positive prognostic steps, including the priming, value in colon, breast, skin recruitment, trafficking, (melanoma) and ovarian entry and accumulation of cancer [30-33]. Nonetheless, activated T-cells through subsequent studies looking various signalling pathways at advanced melanoma

Illustration by Mohd Arshad

“...tumours

could generate an immunologically-restrained milieu by interfering with activated T-cells through various signalling pathways that facilitate immune escape.” revealed that, notwithstanding their presence, the T-cell response is in actuality blunted albeit being largely reversible [34]. Further research eventually led to the discovery of inhibitory receptors such as immune checkpoint molecules, which were found to be highly expressed in tumour tissues and contribute to this immunosuppressive conditions [35, 36]. As a transducer of co-inhibitory signals, these immune checkpoint molecules inherently exist to maintain immunological homeostasis to limit over-activation of the host immune systems. Hence, the concept of immune checkpoint blockade is to induce therapeutic benefit by counteracting the immunosuppression in the tumour microenvironment.

antibodies against those molecules have been developed such as ipilimumab (antiCTLA-4 antibody), nivolumab and pembrolizumab (antiPD-1 antibody) which have been approved for use in a number of countries. Antibodies against programmed cell death ligand-1 (PD-L1) are also under development [39]. So far, treatments with these antibodies have shown promising results.

Durable clinical response were noted in 15–20% of the patients treated with CTLA-4 blockade while anti-PD-1 notably reported a 3-year survival rate of up to 40% in advance stage cancers [40-43].

While not all agents necessarily confer an overall survival (OS) advantage [44, 45], the long-term survival benefit is unobserved in conventional therapies for which Two most representative the clinical responses, though immune checkpoint molecules, immediate, are commonly at present, are the cytotoxic transient. Despite longer OS T-lymphocyte-associated protein-4 has been reported in skin, lung, (CTLA-4) and programmed cell kidney and bladder cancer death-1 (PD-1) [37, 38]. Accordingly, [46-48], other types of cancers

unfortunately do not respond well to these therapies, probably due to insufficient numbers or repertoires of neo-antigens to evoke host immunity. Apart from CTLA-4, PD-1 and PD-L1, other immune checkpoint molecules with potential as clinical targets include lymphocyte-activation gene-3, T-cell immunoglobulin mucin-3, and B- and T-lymphocyte attenuator [49, 50]. Research and development of those molecules are actively carried out at present.

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Conclusions and Future Directions The progression from Coley’s toxins to the myriad of immunotherapeutic approaches under development today speaks profoundly of the contribution of immunology in the evolution of cancer therapeutics.

Ultimately, whether T-cells are activated or inactivated upon TCR ligation will depend on the delicate balance between the stimulatory and inhibitory cosignals. On the other hand, multiple lines of evidence indicate that cancer stemlike cells or cancer-initiating cells (CSC⁄CIC) – which are notoriously resistant to the current standard therapies, including molecular-targeting therapy – also express several TAAs that are recognisable by CTLs both in vitro and in vivo [51]. Given their susceptibility to CTL and high level of expression of TAAs, it is likely that CSC⁄CIC would be the next target for future cancer immunotherapy.

Indeed, several trials of both pre-

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clinical and clinical settings have been reported, with DNAJB8-derived antigenic peptide being one example of a promising candidate for both colon and kidney CSC⁄CICtargeting immunotherapy [52].

Indeed, the research on cancer therapy has come a long way in the past few decades and each step of this journey has been marked by milestones that shaped the current clinical approach [53].

Still, the outstanding value of Coley’s immunotherapy regimen, perhaps more than any existing therapeutics, stemmed from the observed clinical recoveries even in advanced diseases, with such patients in remission for life.

While cases of spontaneous regression have been controversial, disregarding these exceptional examples may risk losing valuable opportunities to learn about the intricacies of our immune system.

Alongside the rapid advances in sequencing technologies, the knowledge of tumour immunology has been greatly expanded at the molecular level. More than just providing an incentive for the development of novel immunotherapies, a better mechanistic understanding will allow clinician to stratify individual patients accordingly and exploit their inherent immune capacity to complement the standard-ofcare treatment. The synergy of modulating various arms of immunity for potential incorporation into the existing surgery, chemotherapy, targeted therapy and radiotherapy may hold great promise as the curative combination for advanced stage malignancies and will probably form the foundation for future personalised treatment.

About The Author Litt-Yee Hiew is currently studying towards an MSc in Molecular

Medicine at the International Medical University. She finds great fulfilment from unravelling the wonders of science as well as in creative writing that could foster meaningful dialogue and bridge the gap between the scientific community and society. Find out more about Litt-Yee by visiting her profile at: http://www.scientificmalaysian.com/members/lyhiew/

References 1. Everson, T.C. (1967). Spontaneous regression of cancer. Prog Clin Cancer., 3: 79-95.

2. Papac, R.J. (1998). Spontaneous regression of cancer: Possible mechanisms. In Vivo., 12: 571-578.

3. Hoption Cann, S. A., van Netten, J.P., van Netten, C. (2003) Dr William Coley and tumour regression: A place in history or in the future? Postgraduate Med J., 79: 672-80.

4. Coley, W.B. (1893). The treatment of malignant tumors by repeated inoculations of erysipelas: with a report of ten original cases. Am J Med Sci., 105: 487-511.

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SUMMARY

C u r r e n t p r o g r e s s

on malaria vaccines

By Adaikalavan Ramasamy

N

on-vaccine approaches for tackling malaria have made an impact in reducing the number of malaria cases and deaths but a vaccine would help tremendously towards malaria elimination. New sources of funding such as those from the Bill and Melinda Gates Foundation, pharmaceutical companies and oil companies in the last decade has enabled rapid advances in malaria vaccine development. We are likely to see the RTS,S/AS01 vaccine to be licensed for use in the next few years despite its low

such a highly effective vaccine, we should consider RTS,S/AS01 as complementary to existing malaria eradication programs.

Resistance towards antimalarial drugs should be taken seriously as there are currently no new effective drugs in the pipeline. A genetic surveillance of sampling mosquitoes and pathogens such as those carried out by the Malaria Genomic Epidemiology Network (MalariaGEN) is vital to map out any emerging resistance and to monitor outbreaks. Additionally, we should actively monitor for counterfeit and substandard drugs.

A more effective vaccine would be one that targets the parasite at multiple stages Finally, we need to invest more funding (i.e. the sporozoite stage, liver stage, and research into other malaria species, blood stage and transmission blocking). especially P. knowlesi and P. vivax species that are more prevalent than P. falciparum to develop and implement. Until we have in Malaysia.

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About Malaria M

alaria is a mosquito-borne disease that Malaria could be prevented either by represents a major public health risk, eliminating the mosquitoes that act as the affecting nearly half the world population and vector for the parasites, limiting the spread of the disease by rapid diagnosis and treatment World Health Organization (WHO) estimates of suspected cases or by using vaccines to approximately 214 million new malaria cases enhance the human immune system to kill and 438,000 malaria deaths worldwide in the pathogen rapidly. In reality, a combination 2015 alone [1]. Children under the age of 5 of all these approaches will be required to living in sub-Saharan Africa accounted for eliminate malaria.

70% of these deaths. Other high-risk groups include pregnant women, immunosuppressed individuals (e.g. HIV infected patients), travellers and the elderly.

MALARIA LIFE CYCLE M

alaria is caused by the Plasmodium parasites transmitted by infected female Anopheles mosquitoes. A female mosquito requires blood to produce eggs and through the process of biting the parasite enters the host’s blood vessels, where it travels rapidly to the liver to mature and multiply. Then it

breaks out of the liver to infect red blood cells to multiply again, causing red blood cells to burst, thus releasing the parasites back into the bloodstream. If another female mosquito bites the infected human at this stage, the mosquito also becomes infected, thus completing the life cycle of malaria transmission (Figure 1).

Figure 1: The life cycle of malaria plasmodium (Image credit: OpenLearn Works)

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Figure 2A: Endemicity map of the Plasmodium falciparum in 2010 (Image credit: The Malaria Atlas Project)

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Figure 2B: Endemicity map of the Plasmodium vivax (bottom) in 2010 (Image credit: The Malaria Atlas Project)

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Figure 3: Potential geographical range of the Plasmodium knowlesi parasite reservoir map in 2013 in Asia region (Image credit: The Malaria Atlas Project)

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Different types of malaria S ix different species of Plasmodium are currently known to infect humans.

Each species has a different geographical spread, life cycles, incubation periods, disease severity and treatment approaches.

P. falciparum is the most common species worldwide (~75%) and the main cause of severe and complicated malaria (Figure 2A). The second most common species worldwide is P. vivax

(~20%) which is predominantly found in South America and Asia (Figure 2B).

However, the most common species in Malaysia [2] are P. knowlesi (38%), P. vivax (31%) and P. falciparum (19%) [2]. Note that P. knowlesi traditionally infected monkeys only but it has recently acquired the ability to infect humans as well and this is an emerging problem particularly in Malaysia (Figure 3).

Malaria symptoms, diagnosis and treatment S ymptoms usually begin between 10 - 30 days after being bitten. The symptoms include fever, chills, headache, fatigue and vomiting. Classic hallmarks of malaria is a repeated cycle of cold stage (intense cold and shivering lasting < 1 hour) followed by hot stage (intense heat, headache, dry

skin lasting 2 - 6 hours) and sweating stage lasting 2 - 4 hours.

Early and accurate diagnosis is key to selecting the correct treatment, shortening illness duration and to prevent lifethreatening complications (e.g. cerebral

malaria, severe anaemia, respiratory These drugs are considered to be the most distress). However, early diagnosis can be effective antimalarial currently available and contribute hugely to malaria reduction.

However, counterfeit or substandard drugs threaten to undermine this success. prick under a microscope (Figure 4) or Further, parasites that are resistant to these commercially available rapid diagnostic test antimalarial drugs have been reported in kits.

Cambodia, Laos, Myanmar, Thailand and Vietnam. This is worrying given that no new antimalarial drugs are anticipated in the Artemisinin-based combination therapies near future. for uncomplicated P. falciparum malaria.

Figure 4: P. falciparum culture which shows several red blood cells with the pathogen inside them (right).

(Image credit: Wikipedia)

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Malaria control and elimination efforts in Malaysia M alaria elimination programs focusing on mosquito control in Malaysia have been successful in reducing the number of cases from ~50,000 in the early 1990s to ~5,000 in 2012 with a corresponding reduction in deaths due to malaria per year from 43 to 16 [2].

However, these efforts require constant monitoring, funding and considerable foresight. Furthermore, majority of malaria cases arise from remote areas in Sabah and Sarawak where access

of malaria cases (~30%) are â&#x20AC;&#x153;imported casesâ&#x20AC;? â&#x20AC;&#x201C; where migrant workers from Indonesia and Philippines who acquire Elimination strategies include indoor malaria during home visit before residual spraying, using insecticide returning to Malaysia where their access treated bed nets and good and fast to medical doctors is more limited [2]. management of malarial outbreaks.

RTS,S/AS01 vaccine (tradename Mosquirix) R

TS,S/AS01 is a recombinant vaccine which fuses the P. falciparum circumsporozoite protein with surface antigen from Hepatitis B and adjuvanted with AS01 (to increase immune response). The vaccine induces high levels of anti-circumsporozoite antibodies which can attack the parasite before it can invade the liver cells. It also provokes a strong CD4 T-cell response which can kill the parasite in the liver before it can break out of the liver.

malaria, severe anaemia, malaria hospitalization, was dropped to 27% in infants who received four doses (same schedule but starting at 6 weeks of life). Infants and young children who did not receive the fourth dose had an even lower overall within 7 days of vaccination in the children in older groups compared to the control group.

The European Medical Agency evaluated the

Vaccine development for malaria V accines have been generally considered to be the cheapest and most effective public health measure for many infectious diseases and malaria is no exception. They could be integrated along with the routine vaccination schedule and could offer long lasting protection as well as herd immunity

vaccines will considerably enhance and complement the malaria elimination efforts. Numerous vaccines are in development but they mainly focus on P. falciparum and target the parasite before they burst out of the liver. In the following sections, we discuss the three most advanced and promising vaccines: RTS,S/AS01, ChAd-MVA population is vaccinated. Therefore, with MeTRAP and PfSPZ. the availability of affordable malaria

This vaccine was developed by GlaxoSmithKline (GSK) over three decades with funding support from the PATH Malaria Vaccine Initiative (MVI) is favourable from a regulatory perspective. In and Bill & Melinda Gates Foundation. This is October 2015, the WHO reviewed the evidence and recommended large-scale implementation critical Phase 3 clinical trial enrolling over 15,000 pilots to replicate the protection reported in the volunteers from 11 trial sites in seven African children aged 5 - 17 months who received the countries. Two age groups were included: infants 4-dose schedule [4]. GSK is now in the process aged 6 - 12 weeks and children aged 5 - 17 of planning Phase 4 study to further characterise months with a median participant follow-up of the safety and effectiveness of RTS,S/AS01 48 months [3]. vaccine. These studies are expected to eventually recruit 800,000 children aged 5 - 9 months in 3 The best protection was seen in the children - 5 sites located in sub-Saharan with moderate recruited at ages 5 - 17 months who received high malaria transmission settings. four doses (vaccination at 0, 1, 2, and 20 months

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ChAd63/MVA ME-TRAP vaccine V

iral vector vaccines use harmless and replication-defective viruses to carry and deliver pathogen sequences to train the human immune systems. The viruses are chosen from non-human species so they are not neutralised too quickly and designed to be harmless and replication-

(MVA), which is a type of smallpox vaccine, expressing ME-TRAP. This heterologous prime-boost strategy provokes strong immune response as the backbone for many candidate

and more recently with Ebola. The control group consisted of 60 adult been shown to induce potent T-cell males who received rabies vaccine response [5] which is required to as placebo. All volunteers were given destroy liver cells that have been antimalarial drugs after vaccinations infected with the parasites. to clear parasites and then monitored for 8 weeks post MVA vaccination for Researchers from Jenner Institute, infections.

University of Oxford and the Malaria Viral Vectored Consortium reported They found that vaccinations reduced the risk of infection by 67%. While They used a chimpanzee adenovirus these results are very promising, the (ChAd63), which is similar to human common cold virus, synthetically in a much larger trial with longer constructed to express the highly follow-up and participants from conserved regions of malaria multiple sites with different malaria antigens called ME-TRAP to prime transmission rates. Furthermore, the 61 healthy adult males in Kenya. results need to be replicated in infants Eight weeks later, the volunteers and young children who are at higher were boosted with an attenuated risk of malaria infection.

PfSPZ vaccine P

fSPZ vaccine was developed by Sanaria and made up of non-replicating irradiated whole sporozoites. Sporozoite is the plasmodium form that leaves the mosquito during the feeding process and infects the liver cells. Sanaria currently collect sporozoites manually from dissecting salivary glands of mosquitoes and then irradiate and freeze them for vaccination. In an earlier study

protection in 6 out of 9 volunteers who received four doses and 6 out of 6 volunteers who received the PfSPZ Vaccine Clinical consortium has set-up seven different clinical trials in USA, Africa and Germany which will recruit at least 450 volunteers.

very high, there are several criticisms. Collection intravenously in non-human primates and mice of large amounts of sporozoite manually will be was far more immunogenic than subcutaneous or challenging for large-scale implementation and intradermal vaccinations.

Sanaria is working on a robot to automate this Next, they recruited and vaccinated 40 adults with process. The second criticism is that the vaccine different dosages and number of vaccines and requires super-cold liquid nitrogen which would then deliberately challenged with the pathogen to be logistically challenging in Africa. Finally, the the highest dose of the vaccine, they observed young children needs to be considered further.

About The Author Adaikalavan Ramasamy is currently the Senior Leadership Fellow in Bioinformatics and heads the Transcriptomics Core Facility at the Jenner Institute, University of Oxford. His team uses gene expression as a tool to understand how the immune system responds to vaccination and why this response differs among individuals. Understanding the mechanisms of protection can help inform vaccine development for infectious diseases and cancer immunology. Adai and his team work on a broad range of novel and licensed vaccines in adults, children and

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References [1] Fact Sheet: World Malaria Report 2015 (updated 9th December 2015) http:// www.who.int/malaria/media/world-malaria-report-2015/en/ [2] Management guidelines of malaria in Malaysia, Ministry of Health Malaysia

sults of a phase 3, individually randomised, controlled trial. Lancet. 2015 Apr 23. pii: S0140-6736(15)60721-8 [4] Malaria vaccine: WHO position paper – January 2016.

[5] Ewer, K. J. et al. Protective CD8+ T-cell immunity to human malaria induced by chimpanzee adenovirus-MVA immunisation. Nature Communication. 4, 2836 (2013).

[6] Ogwang, C. et al. Prime-boost vaccination with chimpanzee adenovirus and Plasmodium falciparum infection in Kenyan adults. Science Translation Medicine 7, 286re5 (2015).

[7] Epstein, J. E. et al. Live attenuated malaria vaccine designed to protect through hepatic CD8+ T cell immunity. Science 334, 475–80 (2011).

The Dengue Vaccine Dilemma: Route to Prevention Are We There Yet?

[8] Seder, R. A. et al. Protection against malaria by intravenous immunization with a nonreplicating sporozoite vaccine. Science 341, 1359–1365 (2013).

By Nor Ilham Ainaa Muhsin

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Summary Although the Phase III clinical studies showed that DengvaxiaÂŽ was efficacious with satisfactory safety profile, we still have to continue monitoring the vaccine for long-term adverse effect by developing suitable models and active collaborations to decipher the immunological mechanisms that might be triggered by the vaccine.

We also require a thorough cost-benefit analysis to determine the best option for dengue vaccination program in Malaysia. Besides, we should not neglect other approaches such as vector control and dengue hotspot identification that can benefit other mosquito-borne diseases (e.g. malaria).

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Global Burden of Dengue Dengue represents a global health issue as it is endemic in over 100 countries, many with tropical and sub-tropical climate, and affects nearly 40% of the world population [1] (see Figure 1). The World Health Organization estimates 50 - 100 million infections and 22,000 deaths occur annually, commonly in children [2]. The economic burden due to diagnosis, hospital treatment,

time off work, impact on tourism is difficult to quantify but likely to be very high and comparable to malaria or TB. The epidemiologic and ecologic factors affecting the spread of dengue include urbanisation trends, global warming and increased global travel, which have led to large epidemics with high mortality in Southeast Asia.

Figure 1: Distribution of global dengue risk (Source: http://www.eliminatedengue.com/our-research/dengue-fever)

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Clinical Aspects of Dengue Symptoms appear 3 to 14 days after infection and can range from mild to high fever with severe headache, muscle and joint pain. Dengue Hemorrhagic Fever (DHF) is a severe form of dengue and can result in persistent vomiting, abdominal pain, bleeding and breathing difficulty.

There is currently no treatment for dengue fever but the symptoms can be managed and reduced by giving intravenous fluid to counteract the fluid leak from blood vessels in the critical defervescent phase.

Dengue Virus and Transmission In 1903, it was demonstrated that dengue fever was caused by a viral infection transmitted by mosquitoes during feeding [3]. The primary mosquito species is Aedes aegypti, which can also transmit the chikungunya, yellow fever and Zika virus.

The dengue virus is a single positive stranded RNA virus belonging to the Flavivirus genus of the Flaviviridae family. Albert Sabin (developer of the oral polio vaccine) identified two serologically different viruses

that cause dengue in 1944 and to date, four serotypes (DENV1 â&#x20AC;&#x201C; 4) have been identified. These serotypes are phylogenetically and antigenically distinct and therefore can be considered as separate viruses.

Infection with one dengue serotype can provide lifelong immunity against reinfection with that particular serotype, but not against the other serotypes. Secondary infection by a different serotype contributed to the majority of DHF cases [4].

Dengue in Malaysia Malaysia ranks in the top ten countries (see Figure 2A) with the highest dengue infection and death in the world. The dengue hotspots are predominantly found in Klang Valley (60%) followed by Johor (15%). Since 2013, Malaysia has been experiencing unprecedented

outbreaks (Figure 2B and 2C). In 2015, there were 120,836 cases with 336 deaths (almost 330 cases and 1 death per day) due to dengue. This outbreak was associated with a switch in the predominant circulating serotype from DENV3 and DENV4 to DENV2 in early 2013 [5].

Figure 2: A) The average annual number of dengue cases reported in the 30 most endemic countries to the WHO between 2004 to 2010. (Source: www. eliminatedengue.com), B) and C) Number of dengue cases and deaths in Malaysia from 1995 â&#x20AC;&#x201C; 2015. (Source: www.idengue.remotesensing.gov.my)

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Overview of Dengue Vaccine Development Scientists have been trying to develop a vaccine for dengue since the early 1930s. There are at least three main challenges for developing an effective dengue vaccine. First, one needs to achieve immunity toward all four serotypes (i.e. a tetravalent vaccine) as secondary infection with a different serotype leads to an increased risk of DHF. Secondly, it is difficult to achieve a balanced efficacy towards all four serotypes in a tetravalent vaccine formulation due to serotype interference. Finally, the lack of an animal disease model prevents the rapid testing of candidate vaccines.

Several approaches have been tried including live attenuated (where the pathogen is made safe by mutation), inactivated (using chemical, heat or irradiation), DNA vaccines and subunit vaccines. The most advanced vaccine to date is Dengvaxia®. We will now briefly review the vaccine construction and the subsequent clinical trials before discussing the merits of implementation this vaccine in Malaysia.

Construction of Dengvaxia® Vaccine The E and prM gene in the dengue virus have been identified as essential for dengue vaccine construction [6]. The E gene encodes the envelope glycoprotein, which stimulates the production of neutralising antibody in humans. The pre-membrane protein (encoded by the prM gene) is required to process and fold the E protein into the correct 3-dimensional structure.

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Phase III Clinical Trial Evaluation Dengvaxia® successfully completed two phase III clinical studies in 2014 with over 30,000 participants. Participants in both trials received three doses of the vaccine over the course of a year.

These studies aim to examine the efficacy of Dengvaxia® in reducing virologically-confirmed dengue and DHF. The first trial included 10,275 children aged between 2 to 14 years recruited from 11 sites in five Asian countries. The second trial included 20,869 children aged between 9 to 16 years from 22 sites in five Central and South America countries. The volunteers were randomised to vaccine and placebo groups in a 2:1 ratio. The interim result based on 1-year-follow-up from final dose [7]

is summarised in Table 1. The data for the Asian countries was stratified into the younger cohort (< 9 years) and older cohort (> 9 years).

This is the first dengue vaccine to reach clinical Phase 3. The overall vaccine efficacy in the older cohort was encouraging (67.8% in Asian countries and 64.7% in Latin American countries). The most striking finding from this study is that the vaccine dramatically reduced the incidence of DHF by more than 90%.

However, there are several negative findings from the trial. First, the vaccine efficacy was low in the younger cohort (44.6%) which is the group at the highest risk. The

In early 2000, scientists from Acambis (now acquired by SanofiPasteur, a French pharmaceutical company) adopted the strategy to incorporate these two genes into a yellow fever vaccine 17D strain genomic backbone (see Figure 3) to produce a recombinant, liveattenuated, tetravalent dengue vaccine (CYD-TDV), commercially known as Dengvaxia®.

Figure 3: Construction of Dengvaxia® vaccine. The E and prM genes from all serotypes were incorporated into the yellow fever vaccine backbone. (image credit: Guy et al (2015) [8])

Table 1: Main findings of the Phase III clinical studies with CYD-TDV (Dengvaxia®)

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investigators hypothesised that the low efficacy in younger cohort could be due to the ongoing development of some vascular physiology such as the capillary system in young children. Secondly, the vaccine efficacy for DENV2, the predominant circulating serotype in Malaysia, was disappointingly low (36.8% in Asian countries and 50.2% in Latin American countries) even in the older cohort. Finally, the investigators also found much higher vaccine efficacy in individuals who were seropositive at baseline, indicating previous exposure to dengue virus.

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Based on the interim results, the WHO recommended introducing the vaccine in highly endemic dengue countries. As of April 2016, the vaccine has been licensed in Mexico, Brazil and the Philippines.

These successful authorisations will allow collaboration with the national authorities to conduct further Phase IV studies to monitor any long-term adverse effects of the vaccine in reality and the feasibility of the vaccination programme.

About The Author

Dengvaxia® Dilemma in Malaysia Upon completion of Dengvaxia®’s Phase III clinical trials, the current Deputy Health Minister, Datuk Seri Hilmi Yahya announced that the vaccine will be made free for the public by mid-2015 but the decision was reversed several months later.

This is due to the fact that the overall vaccine efficacy at 60% was not convincing for larger scale usage.

Besides, Dengvaxia® was shown to be not as effective against the most current prevalent serotype in Malaysia, DENV2.

Professor Emeritus Dato’ Dr. Lam Sai Kit, an eminent virologist from

the University of Malaya and the Immediate Past Chairman of the Asia-Pacific Dengue Vaccine to Vaccination Steering Committee rebutted the statement. His argument is that even if ineffective, vaccines recipients will still benefit from vaccination as Dengvaxia® was shown to reduce hospitalisation and severe dengue by 80-90%. This can save our public health sector tremendous amount of money each year, as the economic burden of dengue in Malaysia is approximately RM 360 million per year.

Nor Ilham Ainaa Muhsin is currently in her final year of studying DPhil in Molecular Genetics at University of Oxford. She is a newbie in the scientific research field but eager to put her molecular science knowledge into translational studies, which hopefully will benefit the country.

An adventurous person, she enjoys nature the most and has plans to visit Malaysia’s wonderful islands and rainforests once she submitted her thesis. She is also an avid runner.

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References [1] http://www.who.int/mediacentre/ factsheets/fs117/en/ [2] http://www.cdc.gov/dengue/ epidemiology/ [3] Encyclopedia of Entomology, Volume 4 by John L. Capinera [4] Halstead, S.Á., Nimmannitya, S. and Cohen, S.N., 1970. Observations related to pathogenesis of dengue hemorrhagic fever. IV. Relation of disease severity to antibody response and virus recovered.

The Yale journal of biology and medicine, 42(5), p.311.

[5] Ng, L.C., Koo, C., Mudin, R.N.B., Amin, F.M., Lee, K.S. and Kheong, C.C., 2015. 2013 dengue outbreaks in Singapore and Malaysia caused by different viral strains.

The American journal of tropical medicine and hygiene, 92(6), pp.1150-1155.

[6] Mellado-Sánchez, G., GarcíaMachorro, J., Sandoval-Montes, C., Gutiérrez-Castañeda, B., RojoDomínguez, A., García-Cordero, J., Santos-Argumedo, L. and CedilloBarrón, L., 2010. A plasmid encoding parts of the dengue virus E and NS1 proteins induces an immune response in a mouse model.

Archives of virology, 155(6), pp.847856.

[7] Hadinegoro, S.R., ArredondoGarcía, J.L., Capeding, M.R., Deseda, C., Chotpitayasunondh,

T., Dietze, R., Hj Muhammad Ismail, H.I., Reynales, H., Limkittikul, K., Rivera-Medina, D.M. and Tran, H.N., 2015. Efficacy and long-term safety of a dengue vaccine in regions of endemic disease. New England Journal of Medicine, 373(13), pp.1195-1206.

[8] Guy, B., Briand, O., Lang, J., Saville, M. and Jackson, N., 2015.

Development of the Sanofi Pasteur tetravalent dengue vaccine: One more step forward. Vaccine, 33(50), pp.7100-7111.

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SciMy Ask Me Anything:

Question 1:

Have you always planned to transition from academic research to joining a biotech company, and then to founding your own startup? What were the factors that influenced you to take this path?

Not at all. It was always my interest and intention to be in academia. After my post-doctoral fellowship at Harvard, I was an Assistant Professor at Johns Hopkins in Baltimore, which was about a 45 minutes commute one-way to work each time. After six years and when our youngest was born, and with my husband Steve traveling a lot, the distance and time of getting to work became a consideration.

So when I was recruited for the position of Director of Molecular Biology at a new start-up biotech company, I resisted until it became clear that the labs were two minutes away from my home!

Question 2:

As a Malaysian scientist, have you been approached to contribute to, or help the local research community?

Dr. Betty Kim Lee Sim

I just made it as a visiting Professor at the University of Malaya, my alma mater, in December 2015.

During my last visit, I had the opportunity to lecture and meet and interact with the fabulous staff and students at the Faculty of Medicine. I am working on a nice collaboration with them now, linking with other institutions in the US and co-applying for grants with Malaysians in Malaysia. This is great and exciting!

Founder of Protein Potential LLC & Executive Vice-President at Sanaria Inc.

From the 1st to 13th of March 2015, Scientific Malaysian organised an Ask Me Anything (AMA) session on their online discussion platform with Dr. Betty Kim Lee Sim (@bkimleesim), Founder of Protein Potential LLC and Executive Vice President of Process Development and Manufacturing at Sanaria Incorporated. Dr. Sim’s team at Sanaria Incorporated has developed the world’s first attenuated sporozoite malaria vaccine, the PfSPZ Vaccine, which is now in clinical trials in the United States, Europe, and Africa. In particular, the PfSPZ Vaccine has also been shown to be 100% protective in two clinical trials.

Throughout this AMA session, registered members of Scientific Malaysian were given opportunities to post questions directly to Dr. Sim. Here, we summarise the discussion that took place during the session.

Question 3: Last year in the United States, pharmaceutical companies inflated the price of medication and vaccines way above the cost of making them. Considering that the PfSPZ vaccine is targeted at developing countries where malaria is most prevalent, what precautionary steps would you be expected to take after it is approved for use?

It is now estimated that it costs more than US$2.6 billion to bring a new drug or vaccine to market in the United States1. In order to keep new drugs and vaccines coming, these costs must be recouped.

Having said that, Sanaria’s goal is to bring a PfSPZ vaccine to market as soon as possible and to use it to halt transmission of and eliminate malaria through mass vaccine administration campaigns. This will only be possible if the vaccine is available at the lowest possible cost. We have already begun making agreements with African governments to assure this. That also means that the vaccine must be sold at a significant profit in the developed world for travelers to help fund the lowest possible cost in the developing world.

http://csdd.tufts.edu/news/complete_story/pr_tufts_csdd_2014_cost_study

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Question 4:

There appears to be little vaccine research and development in Malaysia. How could Malaysia build capacity in terms of research into vaccine research and development? What vaccines or diseases would you suggest to prioritise?

These are critically important questions, but not so easy to address, as this will represent a tremendous shift in perspective and national commitment backed up by investment.

Singapore is probably a good model. Here are several suggestions, (i) the government needs to set up a funding mechanism with significant amount of funds behind it to support this type of research (the Small Business Innovative Research [SBIR] program in the United States is a good example),

Question 6:

What do you think is the best way we can shape this pharmaceutical industry to enable less profitable cures and vaccines to be researched and funded?

The only way to shape anything is to move forward to achieve your goals.

Crowdfunding is a good way to bring attention to these diseases, and to get some seed money. However, the amount of money required to bring a new drug or vaccine to market (currently estimated to be $2.3 billion) cannot be raised through crowdfunding.

(ii) the government needs to work with excellent Malaysian physician scientists to establish a first rate Phase I and then Phase II clinical trials center like he Jenner Institute in Oxford, United Kingdom (we work at such a center in Tanzania), (iii) the team at this clinical trials center needs to be fully trained in GxP (GMP, etc.) and have the personnel (quality, regulatory, clinical, laboratory) and infrastructure (IT, laboratory, etc.) to run clinical trials at a level acceptable to the entire world, and (iv) a few diseases such as dengue, which is now devastating in Malaysia, need to be focused on to get the enterprise off the ground. With such a commitment and resource, I think the Malaysian investment community, which has a large international presence in biotech, could be convinced to invest in Malaysian enterprises.

Question 5:

Most clinical trials involving vaccines or drugs are very much based on western population. Studies have shown that several reasons (genetic predisposition, environmental factors, etc.) could contribute to the poor vaccines/drugs efficacy in non-western countries (i.e., developing countries). How can we address this issue to improve human health and to tackle diseases in the non-western countries?

We are quite mindful of these concerns, and in the context of the PfSPZ vaccine, we will soon be reporting on dramatic differences in immunological responses to immunisation with the exact same regimen of PfSPZ Vaccine in non-immune U.S. and semi-immune Malian (West African) subjects. For these reasons, and because of the importance of malaria worldwide, we are testing our PfSPZ vaccines simultaneously in multiple ethnic groups at multiple sites in the U.S., Germany, and six countries in Africa, and are working on getting studies going in Southeast Asia and South America. The answer is to assess products where they will be used.

Dr Betty Sim Kim Lee pictured here with local children while working on clinical trials in Doneguebougou, Mali (Source: The Malay Mail Online, picture courtesy of Dr Sim)

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Question 7:

Have you ever felt that your gender played a role in your career progression, especially as you started during an earlier generation when there might have been more barriers for women? In relation to that, how have you coped with juggling a career and a family?

No. I have never felt that my gender inhibited my work or success in anyway. Perhaps it is an attitude. Yes, I did juggle my career and family, but in a happy way. I never distinguished my work from family life. Scientific discussions and discourse were often at the dinner table. My sons met and got to know our collaborators and friends in science and medicine from all over the world. I brought my kids into the lab even when they were young when I had to tend to experiments! Gone are those days when security and lab safety issues were non-existent.

Question 8: Any words for Malaysians that chose to have a career in scientific research?

Biography: Dr. Betty Kim Lee Sim hails from Kota Bharu, Kelantan. Dr. Sim did her undergraduate (B.Sc., Honors, First Class) and graduate studies (M.Sc., Ph.D.) at the University of Malaya, Kuala Lumpur. After a postdoctoral fellowship in molecular biology at the Harvard School of Public Health, she became a research assistant professor at the Johns Hopkins School of Public Health and worked with the Walter Reed Army Institute of Research in their malaria program.

In 2003, Dr. Sim founded Protein Potential LLC, a company focused on discovering, producing, and developing subunit recombinant vaccines, therapeutics, and diagnostics.

Protein Potentialâ&#x20AC;&#x2122;s core capabilities include the capacity to rapidly and efficiently take newly discovered molecules through all steps required to initiate and conduct clinical trials. Protein Potentialâ&#x20AC;&#x2122;s platform technology includes recombineering foreign genes into Ty21a the attenuated Salmonella Typhi typhoid fever.

Dr. Sim is also Executive Vice President of Process Development and Manufacturing at Sanaria Inc., where she built the team and led the manufacturing effort for the PfSPZ Vaccine. Dr. Sim currently lives in Maryland, United States with her husband Dr. Stephen L. Hoffman, the founder, CEO and Chief Scientific Officer of Sanaria. Their three sons are pursuing careers in law, medicine, and science.

To find out more about Dr Sim, visit her Scientific Malaysian profile at http://www.scientificmalaysian.com/members/bkimleesim/

Research is a challenging career. But it selects for those who like the wild ride. Be bold and enjoy the ride!

Question 9: Roti canai or nasi lemak?

Roti canai :)

This interview has been edited for brevity and clarity. The original version of this interview can be accessed at: http://www.scientificmalaysian.com/groups/general-research/forum/topic/ask-meanything-ama-dr-betty-kim-lee-sim/

Dr Betty Sim Kim Lee at work in a laboratory in Equatorial Guinea (Source: The Malay Mail Online, with picture courtesy of Dr Sim)

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GEms From Our Web A rt ic l e s

http://magazine.scientificmalaysian.com/life-as-ascientist/interview-hafizah-noor-isa-gravitationalwaves/

Scientific Malaysian team? http://magazine.scientificmalaysian.com/life-as-ascientist/scimy-interview-professor-dato-dr-mokhtarsaidin/

Web Developers http://magazine.scientificmalaysian.com/life-asa-scientist/interview-prof-philip-crosier-universityauckland/

Magazine Illustrators or Designers

News Editors http://magazine.scientificmalaysian.com/life-as-ascientist/scimy-interview-professor-mark-stonekingpart-ii/

Publicity Officers http://magazine.scientificmalaysian.com/lorealunesco-malaysia-for-women-in-science-2015fellows-interview

University Ambassadors

http://www.scientificmalaysian.com/2016/01/12/ event-report-icyc-2015/

we are always looking forward to new ideas! http://www.scientificmalaysian.com/2016/02/01/ event-report-effective-science-communicationworkshop/

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