The Secret Correlation between Cancer and Genetics

By Ryan Gomes

Image by Ruslan Kalendar. [CC-BY-SA 4.0]

DNA – the foundational component of sequences in our bodies that helps us live and be unique from one another. This important structure in every single living organism holds many secrets that tell us about our past ancestral generations and our futures as well. Scientists are constantly trying to uncover these secrets and discover new information that will help improve the quality of human life. Dr. Federico Innocenti is an associate professor in the Division of Pharmacotherapy and Experimental Therapeutics of the UNC Eshelman’s School of Pharmacy. He uncovers secrets of our DNA and genetics that will be able to help treat future patients suffering with cancer, particularly cancers in the gastrointestinal regions of the body. Throughout his research, Dr. Innocenti observed how certain sequences of DNA might influence survival rates from gastrointestinal tumors and how those sequences might affect how our bodies respond to chemotherapies. The particular chemotherapy in Dr. Innocenti’s lab is a widely known drug called irinotecan, or CPT-11. Irinotecan inhibits the topoisomerase, a crucial enzyme that is needed for DNA replication. Cancer cells progress and spread through the body via DNA replication. When inhibiting topoisomerase, DNA replication of cancer cells cannot proceed, therefore preventing further progress of the tumors. Irinotecan is used to treat many types of cancers within patients, including metastatic colorectal cancer, the second most lethal form. Although irinotecan has a reputation to be very efficient and beneficial, studies show that roughly more than 30% of cancer patients who have taken irinotecan to treat their cancer, experienced a worse state of health.⁴ This is largely due to the dosage of irinotecan that doctors prescribe for patients. A higher dosage than necessary for the patient can result in CPT-11-induced toxicity that can cause severe neutropenia. Neutropenia is a reduction in blood counts of neutrophils, a type of white blood cell that helps the body defend against pathogens. Initially, scientists theorized that the link between the amount of dosage and patient health was related to certain factors such as the demographics of the patient like their gender and age, but research has shown that data does not reflect this.⁴ In order to understand and gain more informa24

tion about the correlation between the dosage of irinotecan and the patient’s health, Dr. Innocenti’s lab has looked into the genetic makeup of cancer patients. In order to look into this, the Innocenti lab investigated a certain gene inside the human body called the UGT1A1

Figure 1. This figure above shows how much Irinotecan is necessary in each genotype (the top) in order to get similar SN-38 results (the bottom). It is seen that a patient with *1/*1 genotype needs much more irinotecan dosage in order to maintain similar SN-38 levels compared to the other 2 genotypes. Reprinted with permission. © (2014) American Society of Clinical Oncology. All rights reserved.

“Learning more about the genetic makeup in humans would not only allow new treatments to be developed to help with different forms of cancer.”

gene and measured its common variation, called UGT1A1*28. The UGT1A1*28 gene helps in maintaining adequate levels of SN-38, the by-product formed when irinotecan is metabolized and processed in the body. SN-38 is the main component of the chemotherapy drug that helps in preventing the functionality of the topoisomerase in cancer cells, which helps in blocking further progress of the tumor. When looking into the UGT1A1*28 gene, the researchers found that there were three different forms of the gene and every person has one of those versions of the gene in their DNA.2 The three different forms are: *1/*1, *1/*28, and *28/*28.2 *1 and *28, are examples of alleles, which are variations of the gene that can be combined together in various ways to create what is called the genotype. Since this gene is hereditary and passed down from our biological parents, we all have two alleles or variations of the gene, one from each parent (either alleles *1 or *28), that come together to create one of the three combinations. Depending on the combination, a certain individual with cancer will react differently to a particular dose of irinotecan. In order to test this, Dr. Innocenti designed a clinical trial with a handful of cancer patients that had either the *1/*1 genotype, the *1/*28 genotype, or the *28/*28 genotype. Based on their genotype, cancer patients were given a specific dosage of irinotecan every three weeks. While monitoring the patients’ health conditions during the three-week period, Dr. Innocenti observed whether or not the cancer patients had a negative reaction to the dosage and were at risk of severe neutropenia. The patients with *1/*1 and *1/*28 genotypes started off with intaking 700 mg of irinotecan every three weeks while the patients with the *28/*28 genotype began with 500 mg of irinotecan.2 Patients with *28/*28 genotype started with a smaller dosage amount because “*28/*28 would likely tolerate less irinotecan than the other genotypes and it would not have been ethical to treat all patients at the same dose of irinotecan.”3 As the patients were monitored, the dosage of irinotecan was slowly adjusted to meet the amount that they would be able to tolerate in their bodies. After making proper adjustments to the dosage of irinotecan for the cancer patients, Dr. Innocenti observed that every genotype had a preferred amount of irinotecan that the patient could tolerate in their bodies. Patients with the *1/*1 genotype could intake 850 milligrams but not more than 1,000 milligrams. Those who had the *1/*28 genotype could have 700 milligrams of irinotecan but not any more than 850 milligrams. Lastly, cancer patients with the *28/*28 genotype had by far the lowest level of tolerance to the dosage of irinotecan, being able to take 400 milligrams of irinotecan but no more than 500 milligrams. The results of this experiment can be seen in figure 1 above. This helps to shows that across the three different genotypes, *1/*1, *1/*28, and *28/*28, patients are equally exposed to SN-38, but patients with the *28/*28 genotype need less amount of irinotecan than the patients with the other two genotypes. Although this seems beneficial, further studies have indicated that patients with the *28/*28 genotype are more susceptible to neutropenia since their tolerance levels to irinotecan are so low.1 With further observations, the Innocenti lab were able to conclude that the UGT1A1*28 variation has a significant effect on the amount of irinotecan dosage that cancer patients can intake, providing lower chances of side effects while preserving antitumor potential.2 Dr. Innocenti’s research has helped to demonstrate that although there might be a particular treatment that is found for cancers in the body, the specific amount of treatment that is given to the patient is a very crucial aspect that needs to be considered. Not everyone will be able to take the same amount of dosage and Dr. Innocenti’s research has provided more information about how the genetic makeup of an individual can influence the amount of chemotherapy 25

required for it to have an effect towards treating cancer without risk of side effects. Even though this research is very important, Dr. Innocenti realizes that there is still much to learn about the secrets hidden in our DNA that can help in developing the treatments for different forms of cancer: “the direction I foresee going in towards [the future] is the understanding of how all of these genetic mutations behave with tumors and whether these mutations can open the door to new drugs that can be given to patients.”3 Learning more about the genetic makeup in humans would not only allow new treatments to be developed to help with different forms of cancer, it would also allow doctors to prescribe certain amounts of treatment needed for maximum efficacy and minimum risk of side effects.

Figure 2. This figure shows that irinotecan metabolizes in the body into SN-38

References

1. Dean L. Irinotecan Therapy and UGT1A1 Genotype. 2015 May 27 [Updated 2018 Apr 4]. In: Pratt VM, McLeod HL, Rubinstein WS, et al., editors. Medical Genetics Summaries [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2012-. Available from: https:// www.ncbi.nlm.nih.gov/books/ NBK294473/ 2. Innocenti F, Schilsky RL, Ramírez J, et al. Dose-finding and pharmacokinetic study to optimize the dosing of irinotecan according to the UGT1A1 genotype of patients with cancer. J Clin Oncol. 2014;32(22):23282334. doi:10.1200/JCO.2014.55.2307 3. Interview with Federico Innocenti, M. D., Ph. D. 9/11/2020 4. Karas S, Etheridge AS, Tsakalozou E, et al. Optimal Sampling Strategies for Irinotecan (CPT-11) and its Active Metabolite (SN-38) in Cancer Patients. AAPS J. 2020;22(3):59. Published 2020 Mar 17. doi:10.1208/ s12248-020-0429-4

Re-engineering the Immune System

By. Reva Kodre

Figure 1. : Image of cross-section of a human breast cancer with cancerous regions. Image Courtesy of Nazanin Rohani.

Our immune system is an army of soldiers, armed body to grow rapidly and possibly metastasize, in which with proteins that tag and alert killer cells to attack the abnormal cells spread to a different region of the body. a foreign body or antigen. The B lymphocytes (B- As the director of the Cancer Cellular Immunotherapy procells) in our immune system are the military intelligence gram at UNC, Dr. Dotti is focused on Chimeric antigen resystem of our body—trigger the production of Y-shaped ceptor (CAR) T-cell therapy, which uses gene modified Tproteins called antibody that lock onto specific antigens cells to create a targeted attack on cancer cells. CARs have such as bacteria. After been more commonly used to attack leukemia, or blood latching onto a certain an- cancers, which typically begins with early forms of white tigen, the antibodies signal blood cells, but it can originate from other types of blood the T lymphocytes (T-cells) cells as well (Figure 2).1 There are many different types of to destroy the tagged an- leukemia, but it is Acute Lymphoblastic Leukemia (ALL) tigens or infected cells.1 Antibodies seems like a foolproof plan of defending our bodies from getting sick, but an issue arises when the immune system is unable to proDr. Gianpietro Dotti, M.D. duce enough antibodies to defend itself. The issue becomes even worse if the B-cells have transformed into malignant cells, unable to fulfill its task of protecting the body. The progression would result in something that is more daunting: cancer. To help with the loss of defenses, UNC-Chapel Hill Lineberger Comprehensive Center’s Dr. Gianpietro Dotti, M.D. has been actively involved in novel types of treatment in cancer immunotherapy clinical trials, specifically the re-engineering of immune cells. Cancer generally develops when there is an abnor- Figure 2. Anatomy of the bone. Image Courtesy of Terese mality in a specific gene that causes a type of cell in the Winslow [CDR755927].

that targets lymphocytes specifically. ALL is common in young patients and can be fatal within a few months if not treated properly.1 Normally, the bone marrow would produce blood stem cells that eventually mature to become either myeloid or lymphoid stem cells and the lymphoid stems cells would then develop into infection-fighting cells.1 However, it is difficult for the immune system to defend itself when the cancer originates in the infectionfighting cells themselves. CAR T-cell therapy involves taking T-cells from a patient diagnosed with cancer and are genetically engineering them in the laboratory to include special receptors on their surface, at which point the cells are classified as CAR T-cells.2 By changing the mechanism of the T-cells and converting them into CAR T-cells, they are now more capable of binding to and attacking the cancer cells (Figure 3). Because these T-cells are taken directly from the patients, this type of treatment is personalized to the specific type of cancer present in their body, making the attack direct

Figure 3. Diagram of CAR T-cell Therapy Treatment. Image Courtesy of Terese Winslow CDR774647]. and specific. One of the most common issues with cancer treatments is that the drug itself might negatively affect normal cells and have a toxic effect on the body. However, CARs are able to attack specific types of antigens that are present in higher levels on cancerous cells compared to normal cells. For example, with B-cell malignancies, it is becoming increasingly more common to target the CD19 antigen that is prevalent on surface of some blood cancer cells.2 In the case of childhood ALL, CD19 CAR T-cells can be used to treat relapsed ALL, along with chemotherapy.2 After observing remarkable regression of blood cancers after using CAR T-cell therapy, scientists like Dr. Gianpietro Dotti hope to apply this mode of therapy to solid, tissue-like tumors as well. The primary challenge with attacking solid tumors is the difficulty for CAR T-cells to reach the small, focused sites of the cancer while also avoiding the healthy cells that surround it (Figure 1). The challenge for solid tumors is different from those of liquid cancers like leukemia, where the infused CAR T-cells are more likely to bump into the cancerous cells that are able to travel freely in the blood and target them. The concern is similar with chemotherapy, which Dr. Dotti explains, “when we do chemotherapy, we don’t kill all the tumor, we kill also a lot of normal cells—you lose your hair.”3 However, Dr. Dotti and his colleagues at UNC Lineberger have developed strategies to allow CAR T-cells to regulate their activity to effectively kill the lymphoma tumor cells without triggering harsh side effects. Dr. Dotti’s team has found that a molecule called LCK is able to increase the activity of a CAR T-cell that responds to the 4-1BB protein signal.4 The LCK molecule enhances the activity of the modified T-cells and thus results in a better tumor-targeted attack. In order to decrease the activity of the CAR T-cells, they have found a new “safety switch” called SHP1 that binds to the CAR T-cells and reduces their activity without killing them.4 SHP1 allows the genetically engineered T-cells to slow down their attack on tumor cells and continue to grow and expand, while avoiding any side effects that may occur.4 There are still many obstacles with CAR T-cell treatments for solid tumors such as pancreatic cancer and ovarian cancer, but Dr. Dotti remains optimistic for future breakthroughs. As these newer and more innovative models of treatment are discovered, researchers at UNC Lineberger hope to see progress towards creating safe treatments to alleviate the toxic effects of cancer. Although this process may take years of rotating through lab experiments and clinical trials, it is necessary to promote the patient’s safety first. Dr. Dotti expresses that CAR T-cell therapy may never be the one true cure for cancer, but when combined with chemotherapy and other forms of immunotherapy, it can dramatically reduce the size of the tumor, which is a huge step closer to the ultimate goal of healing the patient. As Dr. Dotti states, “with every single patient that will come in, you have step one, step two, step three, and hopefully after step three, we’re done.”3

References

1. PDQ® Pediatric Treatment Editorial Board. PDQ Childhood Acute Lymphoblastic Leukemia Treatment. Bethesda, MD: National Cancer Institute. Updated July 13, 2019. Available at: https://www.cancer.gov/types/leukemia/patient/child-all-treatment-pdq. Accessed Sept. 13, 2020. [PMID: 26389385] 2. Dotti, G., Gottschalk, S., Savoldo, B., & Brenner, M. K. (2013). Design and development of therapies using chimeric antigen receptor-expressing T cells. Immunological Reviews, 257(1), 107126. doi:10.1111/imr.12131 3. Interview with Gianpietro Dotti, M.D. 09/11/2020. 4. Sun, C., Shou, P., Du, H., Hirabayashi, K., Chen, Y., Herring, L. E., . . . Dotti, G. (2020). THEMIS-SHP1 Recruitment by 4-1BB Tunes LCK-Mediated Priming of Chimeric Antigen Receptor-Redirected T Cells. Cancer Cell, 37(2). doi:10.1016/j.ccell.2019.12.014