Ldn immune study mccandless 2006 brief version by LDNIM

REPORT: LOW-DOSE NALTREXONE (LDN) EFFECT OF LOW-DOSE NALTREXONE (LDN) ON LABORATORY IMMUNE VALUES AND SELF-ASSESSMENT TRACK-SHEETS FOR PHYSIOLOGICAL AND MOOD EFFECTS PERIOD OF STUDY: 16 WEEKS MARCH – AUGUST 2006 PARTICIPANTS: (20) ASD CHILDREN AND (38) ADULTS (PRIMARILY PARENTS OR OLDER SIBLINGS OF ASD CHILDREN), MARCH – AUGUST 2006 PRIMARY INVESTIGATOR: JAQUELYN McCANDLESS, M.D., PRIVATE PRACTICE, CANOGA PARK, CA AND HONOKAA, HI CONSULTANT: ARISTO VOJDANI, PHD, DIRECTOR, IMMUNOSCIENCES LAB, BEVERLY HILLS, CA EARLY INFORMAL ADVISORS ON THE STUDY: JAAK PANKSEPP, PHD PAUL SHATTOCK, PHD STATISTICAL ANALYSIS PROVIDED BY DAVID GAYLOR, PhD, ASSISTED BY JACK ZIMMERMAN, PhD SOURCES OF SUPPORT FOR THIS STUDY LDN TRANSDERMAL CREAM FOR CHILDREN AND CAPSULES FOR ADULTS WERE DONATED BY COASTAL COMPOUNDING PHARMACY, SAVANNAH, GA PARTICIPANTS PAID FOR DISCOUNTED LAB SERVICES FOR THE IMMUNE TESTING DONATED BY IMMUNOSCIENCES LAB, BEVERLY HILLS, CA OTHER SOURCES OF SUPPORT: NONE

TABLE OF CONTENTS: COVER PAGE

TABLE OF CONTENTS

HISTORY OF NALTREXONE AND AUTISM

USE OF LOW-DOSE NALTREXONE (LDN) IN AUTISM

DESCRIPTION IMMUNE TESTS

4-6

ADDITIONAL NOTES RE TESTING: MOLLOY’S WORK

STATISTICAL APPROACH AND REGULATORY MARKERS AND REGULATORY MARKERS

16-WEEK IMMUNE TEST RESULTS CHILDREN

8-9

SUMMARY IMMUNE TESTS FOR CHILDREN

9-10

16-WEEK TEST RESULTS, IMMUNE TESTING ADULTS

SUMMARY IMMUNE TESTS FOR ADULTS

DISCUSSION OF LDN STUDY

REFERENCES

HISTORY OF NALTREXONE AND AUTISM: NALTREXONE IS AN FDA-APPROVED MEDICATION, A PURE OPIOID ANTAGONIST, AVAILABLE AS THE BRAND NAME RE VIA AS WELL AS IN GENERIC FORM AND USED PRIMARILY FOR TREATING OPIATE AND ALCOHOL ADDICTION SINCE THE 1970’S. SINCE THE 1970’S, STUDIES HAVE CONSISTENTLY SHOWN A VARIETY OF IMMUNE SYSTEM DISORDERS IN AUTISTIC CHILDREN. (1) INFECTIOUS AGENTS, TOXIC CHEMICALS AND DIETARY PEPTIDES HAVE BEEN SHOWN TO BE TRIGGERS FOR IMMUNE DYSREGULATION AND AUTOIMMUNITY. (2) THE HYPERSENSITIVITY REACTIONS TO THE LARGE PEPTIDES FOUND IN CASEIN AND GLUTEN PRODUCTS BY AUTISTIC CHILDREN INSPIRED RESEARCH INTO THE OPIOID ANTAGONIST NALTREXONE IN HOPES OF AVOIDING RESTRICTIVE DIETS TO PREVENT CASEO-OPIOID AND GLUTEO-OPIOID COMPOUNDS IN THE BRAIN FROM CREATING DELETERIOUS EFFECTS COGNITIVELY AND BEHAVIORALLY. (3a, 3b, 3c) AN ITALIAN STUDY DONE IN 1996 WITH VARYING NALTREXONE DOSING SHOWED SIGNIFICANT REDUCTION OF AUTISTIC BEHAVIORS IN 7 OUT OF 12 CHILDREN. (4) BEHAVIORAL IMPROVEMENT WAS ACCOMPANIED BY ALTERATIONS IN THE DISTRIBUTION OF THE MAJOR LYMPHOCYTE SUBSETS WITH INCREASE IN NORMALIZATION OF THE CD4/CD8 (T/1) RATIO. (5) NO SIMILAR IMMUNE STUDIES HAVE SINCE BEEN REPORTED ON THE USE OF NALTREXONE IN AUTISM, WHILE A LARGE BODY OF RESEARCH HAS POINTED REPEATEDLY TO ENDOGENOUS OPIOID SECRETIONS AS PLAYING THE CENTRAL ROLE IN THE BENEFICIAL ORCHESTRATION OF THE IMMUNE SYSTEM. (6) OPIOIDS ARE ENDORPHINS AND OPERATE AS CYTOKINES, THE PRINCIPAL COMMUNICATION SIGNALERS OF THE IMMUNE SYSTEM. (7) STUDIES AT UC DAVIS MIND INSTITUTE SHOW THAT CYTOKINE RESPONSES ELICITED BY THE T-CELLS, B-CELLS, AND MACROPHAGE CELL POPULATONS FOLLOWING THEIR ACTIVATION DIFFERS MARKEDLY IN CHILDREN WITH AUTISM COMPARED TO AGEMATCHED CHILDREN IN THE GENERAL POPULATION. (8) STUDIES WITH NALTREXONE FOR AUTISM HAVE BEEN EQUIVOCAL, AND SUBJECT TO NON-COMPLIANCE DUE TO THE BITTERNESS OF THE DRUG WHICH HAS POSED A PROBLEM FOR AUTISTIC CHILDREN, MOST OF WHOM COULD NOT SWALLOW CAPSULES. BERNARD BIHARI, MD, A NEW YORK PHYSICIAN STUDYING THE IMMUNE RESPONSES IN ADDICTED HIV AIDS PATIENTS IN 1985, DISCOVERED THAT AN ULTRA-LOW DOSE OF NALTREXONE BOOSTS THE IMMMUNE SYSTEM WITHOUT SIDE EFFECTS OR TOXICITY. HE FOUND THAT WHEN GIVEN BETWEEN 9PM AND 2 AM THE PITUITARY IS

ALERTED AND THE BODY ATTEMPTS TO OVERCOME THE OPIOID BLOCK WITH AN ENDORPHIN ELEVATION, STAYING ELEVATED THROUGHOUT THE NEXT 18 HOURS. (9) RESTORATION OF THE BODY’S NORMAL PRODUCTION OF ENDORPHINS IN THOSE WITH CANCER OR AUTOIMMUNE DISEASES OR ANY DISEASE CHARACTERIZED BY INADEQUATE OR DISORDERED IMMUNITY IS THE MAJOR THERAPEUTIC ACTION OF LDN, WHICH WAS SHOWN TO BE NEEDED ONLY ONCE DAILY AT BEDTIME FOR IMMUNE OPTIMIZATION. USE OF LOW-DOSE NALTREXONE (LDN) IN AUTISM I (DR JM) STARTED USING LDN IN A GROUP OF AUTISTIC CHILDREN PLUS A FEW ADULTS WITH OTHER AUTOIMMUNE CONDITIONS IN EARLY 2005 AFTER AN EFFECTIVE TRANSDERMAL CREAM WAS CREATED WHICH COULD BE APPLIED TO THE CHILDREN AT THEIR BEDTIME OR AFTER. DOSES WERE 3MG OF LDN CREAM FOR CHILDREN AND 4.5MG CAPSULES FOR ADULTS PER DR. BIHARI’S DISCOVERY OF THIS WINDOW OF DOSING FOR OPTIMUM IMMUNE BENEFITS. POSITIVE RESULTS IN THIS PIONEER GROUP AND THE LACK OF SIGNIFICANT SIDE EFFECTS LED ME TO DO A LARGER INFORMAL 16WEEK STUDY FROM MAR – AUG 2006 WITH 20 AUTISTIC CHILDREN AND 38 ADULTS (ALMOST ALL PARENTS OF ASD CHILDREN). A PLASMA IMMUNE PANEL WAS PERFORMED AT THE START AND FINISH OF THE STUDY INCLUDING TOTAL WBC’S, T-CELLS, B-CELLS, NATURAL KILLER CELL ACTIVITY, LYMPHOCYTE SUB-POPULATION, AND BRAIN AUTO-ANTIBODIES. WEEKLY PHYSIOLOGICAL AND MOOD CHILD ASSESSMENTS AND PARENTAL SELF-ASSESSMENTS WERE ALSO NOTED ON TRACKSHEETS THROUGHOUT THE 16-WEEK STUDY PERIOD WITH A NUMBERING SYSTEM FROM 1-10, NEGATIVE EFFECT TO POSITIVE EFFECT, 5 BEING NEUTRAL OR NOT/APPLICABLE.

DESCRIPTION: IMMUNE TESTS PERFORMED FOR 16-WEEK LDN STUDY, MAR – AUG 06 LAB VALUES LDN STUDY: TOTAL WHITE COUNT (WBC): THERE ARE 5 KINDS OF WHITE CELLS: NEUTROPHILS, LYMPHOCYTES, MONOCYTES, EOSINOPHILS, AND BASOPHILS. COUNT CAN BE AFFECTED BY GENERAL HEALTH,

INFECTIONS (BACTERIAL RAISE THE NEUTROPHIL COUNT, VIRUSES GENERALLY LOWER NEUTROPHILS AND CAN RAISE LYMPHOCYTE COUNT AND STIMULATE IT TO PUT OUT CYTOKINES) TOTAL T-CELLS: LYMPH CELLS THAT MATURE IN THE THYMUS, CONTROL INNATE CELL-MEDIATED IMMUNE RESPONSES AND ACTIVATE RESPONSES TO MOST ANTIGENS. T-CELLS DEAL WITH VIRUSES AND PATHOGENS THAT REPLICATE INSIDE CELLS WHERE THEY CANNOT BE REACHED BY ANTIBODIES. TOTAL T-HELPER (CD4+): RECOGNIZE ANTIGENS, PRODUCE CYTOKINES (CHEMICAL MESSENGERS) AND ACTIVATE OTHER T & B CELLS, CONTROL INTRACELLULAR BACTERIAL INFECTIONS BY ATTRACTING MACROPHAGES, LEAD ATTACKS AGAINST INFECTION, CONTROL OTHER CELLS, COUNT DETERMINES PRESENCE OF IMMUNE DEFICIENCIES AND AUTOIMMUNITY; COUNT HELPS GUIDE TREATMENT FOR HIV+AIDS. (STUDIES SHOW ANTI-AIDS DRUGS POTENTIATED BY NALTREXONE). (10) TOTAL SUPPRESSOR (CD8+) IMMUNITY “BRAKES”, STOP IMMUNE RESPONSE AFTER INVADING PATHOGENS ARE DESTROYED, “KILLER” CELLS SUPPRESSE IMMUNE RESPONSE, ANTIINFLAMMATORY, CYTOTOXIC (KILLS VIRUSES & CANCER CELLS) T-HELPER/T-SUPPRESSOR THE RATIO OF CD4+ AND CD8+ SHOULD BE 12.5. OVER 2.5 RELATES TO IMMUNE ACTIVATION OR AUTOIMUNITY, LESS THAN ONE RELATES TO IMMUNE SUPPRESSION OR DEFICIENCY. HEAVY METALS AND MYCOTOXINS CAN ALSO SUPPRESS IMMUNE ACTIVITY. TOTAL NATURAL KILLER LYMPHOCYTES IN INNATE IMMUNE SYSTEM, PROVIDE AGGRESSIVE DEFENSE AGAINST CELLS INFECTED BY MICROORGANISMS, ESP VIRUSES AND CANCER CELLS, TARGET INFECTIONS BOTH INSIDE AND OUTSIDE THE CELL INNATELY WITHOUT HAVING TO BE DIRECTED BY THE CD4+ “HELPER” CELLS; WHEN HIGH USUALLY MEANS VIRUSES. INCLUDES TOTAL IMMUNOCOMPETENT CELLS (+ CELLS) AND TOTAL NKHT3 (-CELLS). TOTAL CD3+ CD 26+: MEMORY LYMPHOCYTES, IMPORTANT CELLS, FIGHT TOXIC CHEMICALS, HEAVY METALS, MOLDS AND RELATED MYCOTOXINS, “POLICEMEN”. (THIS IS DPP-1V PEPDIDASE)

IMMUNOREGULATORY CELLS: MAINTAIN HOMEOSTASIS AND SELF-TOLERANCE TOTAL CD4+ CD25 (11) : CD4+CD25+ T-REGULATORY CELLS ARE IMPORTANT IN THE MAINTENANCE OF IMMUNOLOGICAL SELF TOLERANCE AND IN THE PREVENTION OF AUTOIMMUNE DISEASE, FIGHT AUTOIMMUNITY, NEED TGF-B & IL-10 CONTACT TO BECOME POTENT SUPPRESSOR CELLS IL-10 UNSTIMULATED (11): NEUTRALIZE INFLAMMATION, CREATE ANTI-INFLAMMATORY CYTOKINES IL-10 STIMULATED (11): ANTI-INFLAMMATORY IN RESPONSE TO ANTIGENIC STIMULATION TGF-BETA 1 UNSTIMULATED (12): SUPPRESSIVE CYTOKINES

IMMUNOSUPPRESSIVE , CREATE

TGF-BETA 1 STIMULATED (12): SUPPRESSIVE TO ANTIGEN

ADDITIONAL NOTES RE CYTOKINES AND DR. CYNTHIA MOLLOY’S RECENT WORK ON IMMUNE SYSTEMS OF AUTISTIC CHILDREN (PLEASE SEE REFERENCE #13, MARCH 06 JOURNAL OF NEUROIMMUNOLOGY) MOLLOY’S STUDY FOUND THAT IMMUNE CELLS OF CHILDREN WITH AUTISM PRODUCED HIGHER LEVELS OF BOTH THE TH1 AND TH2 CYTOKINES THAN IN CONTROLS, DEMONSTRATING POTENTIAL UNDERLYING HYPERSENSITIVITY TO EXPOSURES IN THE GENERAL ENVIRONMENT FOR ASD CHILDREN. THE MORE HIGHLY ACTIVATED T1 AND T2 CYTOKINES CONFIRM THE IMMUNE DYSREGULATION FOUND IN MANY OTHER STUDIES OF ASD CHILDREN, AND CONFIRM THAT THESE CHILDREN EXHIBIT HYPER SENSITIVITY IN BOTH INNATE AND ADAPTIVE SYSTEMS. THE EXCEPTION SHE FOUND WAS IN RELATIVELY LOWER LEVELS OF THE CRITICAL REGULATORY CYTOKINE, IL-10. DR. MOLLOY SAYS: “MANY PARADOXICAL FINDINGS THAT HAVE BEEN REPORTED ABOUT IMMUNE RESPONSES IN AUTISM COULD POSSIBLY BE EXPLAINED BY THE GENERAL DYSFUNCTION OF IL-10.” SHE IS PLANNING FURTHER STUDIES ON IL-10.

TEST RESULTS FOR 16 WEEK STUDY OF CHILDREN’S IMMUNE SYSTEMS STATISTICAL APPROACH: THE DATA FROM THE STUDY WAS SUBJECTED TO TWO DIFFERENT KINDS OF ANALYSES: QUANTITATIVE AND QUALITATIVE. THE FORMER INVOLVED TAKING THE RATIOS OF FINAL TO ORIGINAL VALUES USING THE CONVENTIONAL t-TESTS TO SEE IF THESE RATIOS WERE SIGNIFICANTLY GREATER THAN ONE.( WHEN THE DATA EXHIBITED WIDE VARIATIONS, THE LOGS OF THESE RATIOS WERE USED.) THESE QUANTITATIVE TESTS ASSUME THAT INCREASES IN THE MARKERS ARE ALWAYS “GOOD,” WHICH IS OBVIOUSLY NOT THE CASE IF THE MARKER VALUES EXCEEDED THE UPPER LIMIT OF THE NORMAL RANGE. FOR THIS REASON—AND ALSO BECAUSE THE SCATTER OF THE MARKER DATA WAS USUALLY GREAT FOR THE CHILDREN (MUCH LESS SO FOR THE ADULTS)—WE ALSO USED A SIMPLE BINARY TEST FOR EACH MARKER BASED ON WHETHER IT MOVED IN A “GOOD” OR “BAD” DIRECTION OR MOVED WITHIN OR OUTSIDE OF NORMAL RANGE.. BECAUSE OF THE SMALL SAMPLE SIZE FOR THE CHILDREN AND THE TENDENCY FOR THE DATA TO SCATTER DUE TO ONE OR MORE UNCONTROLLED VARIABLES (INFECTIONS DURING THE SIXTEEN WEEKS, DIETARY INCONSISTENCIES, ETC.), THESE LATTER TESTS ARE TO BE TAKEN AS SUGGESTIVE EVEN WHEN THE SIGNIFICANCE LEVEL WAS 95% OR GREATER. A LARGER SAMPLE SIZE (AT LEAST EQUAL OR GREATER THAN THAT OF THE PARENTS IN THE STUDY) AND GREATER CONTROL OF OTHER VARIABLES AFFECTING THE POPULATION OF CHILDREN ARE CLEARLY NEEDED IN FUTURE STUDIES. IN WHAT FOLLOWS WE SPEAK OF A MARKER GOING UP OR DOWN OR STAYING ESSENTIALLY THE SAME. THE CRITERION FOR THESE STATEMENTS IS BASED ON THE AMOUNT OF CHANGE RELATIVE TO THE STANDARD DEVIATION OF THE DATA. REGULATORY MARKERS: IL-10: IN OUR NON-PLACEBO (AND NON-CONTROLLED) STUDY - THIS MARKER VARIED WIDELY FROM CHILD TO CHILD AND SEVERAL INDIVIDUAL CHILDREN SHOWED WIDE CHANGES DURING THE STUDY, WITH AS MANY CHILDREN RAISING THEIR VALUES AS LOWERING THEM. THE AVERAGE VALUES WERE RATHER LOW RELATIVE TO THE NORMAL IL-10 RANGE, AGREEING WITH MOLLOY’S WORK. THUS IN 16 WEEKS LDN DID NOT SEEM TO HELP THIS IMPORTANT REGULATOR CYTOKINE TO INCREASE; WE AS YET HAVE NO UNDERSTANDING OF THIS. NONE OF THESE CHANGES WERE STATISTICALLY SIGNIFICANT. 7

TGF-B: THE TGF-B BEGINNING AVERAGE WAS QUITE A LOT HIGHER THAN THE MEDIAN OF THE NORMAL RANGE AND THE AVERAGE FINAL AVERAGE WAS LOWER THAN THE MEDIAN VALUE (ALTHOUGH, AGAIN, THIS CHANGE WAS NOT STATISTICALLY SIGNIFICANT). THIS MAY MEAN THE IMMUNOSUPPRESSIVE EFFECT OF TGF-B WAS LOWERED ON THE AVERAGE BY THE LDN, WHICH COULD BE AN INDICATION THAT AUTOIMMUNITY WAS IMPROVED IN SPITE OF THE IL-10 FINDING, PROBABLY POINTING TO INFLAMMATION STILL BEING SIGNIFICANT IN THIS GROUP. THEIR RELATIONSHIP IS AS YET UNCLEAR. THE SIGNIFICANCE OF THESE VALUES RELATIVE TO THE USE OF LDN IS DIFFICULT TO UNDERSTAND EXCEPT IN VERY GENERAL TERMS, OTHER THAN TO SAY THAT LDN IN THIS STUDY DID NOT SEEM TO STABILIZE THE LEVEL OF IL-10 REGULATORY BENEFIT FOR INFLAMMATORY ISSUES. 16 WEEKS MAY NOT BE LONG ENOUGH FOR THIS VALUE TO CHANGE – IT IS ALSO DIFFICULT IN ANY CIRCUMSTANCE TO DETERMINE AT WHAT PHASE THE INFLAMMATION REACTION IS IN – THIS WOULD HELP DETERMINE WHETHER IT WOULD BE PREFERABLE FOR INFLAMMATORY MARKERS TO BE INCREASING OR DECREASING. TOTAL WBC: IN REGARD TO THIS MARKER MOST CHILDREN WERE WITHIN NORMAL RANGE BOTH BEFORE AND AFTER THE TEST. THREE (3) STARTED BELOW RANGE AND FOUR (4) ENDED UP BELOW RANGE, THIS RESULT WAS CLEARLY NOT STATISTICALLY SIGNIFICANT. TOTAL LYMPHS: MOST CHILDREN WERE WITHIN THE NORMAL RANGE FOR THIS MARKER BOTH BEFORE AND AFTER THE TEST; ONE CHILD STARTED WITH AN ELEVATED VALUE AND ENDED UP WITHIN NORMAL LIMITS; ONE CHILD STARTED WITH NORMAL VALUE AND ENDED UP WITH ELEVATED VALUE. OBVIOUSLY THESE RESULTS WERE NOT STATISTICALLY SIGNIFICANT. TOTAL NATURAL KILLER CELL LEVEL: FIFTEEN (15) CHILDREN MOVED IN A POSITIVE DIRECTION, THREE (3) IN A NEGATIVE DIRECTION, AND ONE (1) STAYED THE SAME. ALTHOUGH THE t-TESTS FOR THIS MARKER WERE NOT SIGNIFICANT, A SIMPLE BINARY TEST SUGGESTS THAT THE CHILDREN AS A GROUP MOVED IN A POSITIVE DIRECTION WITH A SIGNIFICANCE OF 95%. THE AMOUNT OF MOVEMENT VARIED CONSIDERABLY. CD3+CD26: RELATIVE TO THE MEDIAN OF THE NORMAL RANGE, ELEVEN (11) CHILDREN MOVED TOWARD A BETTER SCORE, TWO (2) STAYED ESSENTIALLY THE SAME, AND SEVEN (7) MOVED TO A WORSE

SCORE. . ALTHOUGH THE QUALITATIVE TEST WAS NOT QUITE SIGNIFICANT FOR THIS MARKER, THE SCATTER OF THE DATA WAS SMALL ENOUGH AND THE IMPROVEMENTS OUTWEIGHED THE LESSENING SCORES QUANTITATIVELY FOR A t-TEST SIGNIFICANCE ON THE 95% LEVEL—THE ONLY SUCH QUANTITATIVE RESULT FOR THE GROUP OF CHILDREN. THIS MEANS WE CAN SAY THAT LDN SIGNIFICANTLY INCREASES THIS MARKER AT THE 95% CONFIDENCE LEVEL. THE AVERAGE INCREASE WAS 23%. TOTAL T3: ALL CHILDREN REMAINED WITHIN THE NORMAL RANGE BOTH BEFORE AND AFTER THE TEST, WITH ELEVEN (11) INCREASING AND NINE (9) DECREASING (CLEARLY NOT SIGNIFICANT). TOTAL SUPPRESSOR (CD8+): A MEDIAN OF 900 IS CONSIDERED GOOD; FOURTEEN (14) CHILDREN MOVED IN THE POSITIVE DIRECTION, TWO (2) REMAINED ESSENTIALLY THE SAME, AND FOUR (4 ) MOVED IN A NEGATIVE DIRECTION. ALTHOUGH THE t-TESTS SHOWED NO SIGNIFICANCE, USING A BINARY TEST, THE DIRECTION OF MOVEMENT OR “IMPROVEMENT” FOR THIS MARKER WAS STATISTICALLY SIGNIFICANT ON THE 95% LEVEL. TOTAL T HELPER (CD4+): THIS MARKER IS A KEY MEASURE OF IMMUNE SYSTEM HEALTH. DR. VOJDANI SAYS THAT IF VALUES STAY WITHIN NORMAL RANGE, THE HIGHER THE BETTER. WE FOUND THAT EIGHTEEN (18) OF THE TWENTY (20) MOVED HIGHER, ONE MOVED LOWER, AND ONE STAYED ESSENTIALLY THE SAME., WE CONCLUDED FROM THE BINARY TEST THAT LDN HAD A SIGNIFICANT POSITIVE EFFECT ON A 99% CONFIDENCE LEVEL. (NOTE: THIS BODES WELL FOR THE UPCOMING HIV+ AIDS STUDY ON LDN IN AFRICA). CD4/CD8 RATIO (T-HELPER/T-SUPPRESSOR): THE AVERAGE RATIO OF EIGHTEEN (18) POSITIVE CD4+ TO THE 14 POSITIVE CD8+ GAVE A VALUE OF 1.28 FOR THE CD4+CD8+ RATIO, WHICH SHOULD BE BETWEEN 1-2.5. FOUR (4) CD4+CD8+ RATIOS WERE ABNORMAL IN THE BEGINNING (TWO TOO HIGH, TWO TOO LOW); ALL OF THE CD4+/CD8+ RATIOS WERE WITHIN NORMAL LIMITS EXCEPT ONE (TOO HIGH) AT THE END OF THE STUDY; ALTHOUGH THIS RESULT IS ENCOURAGING IT IS NOT SIGNIFICANT ON THE 95% LEVEL. SUMMARY, CHILDREN: THOSE TESTS SHOWING THE BENEFIT OF LDN WITH STATISTICAL SIGNIFICANCE AT LEAST AT THE 95% LEVEL FOR THE POPULATION OF CHILDREN WERE: TOTAL NATURAL KILLER CELL LEVEL (QUALITATIVELY) TOTAL T-HELPER CELL (CD4+) LEVEL (QUALITATIVELY)

TOTAL SUPPRESSOR CELL (CD8+) LEVEL (QUALITATIVELY) CD3+CD26 LEVEL (QUANTITATIVELY) THOSE TESTS THAT DID NOT SHOW STATISTICAL SIGNIFICANCE: TOTAL WHITE COUNT TGF BETA 1 UNSTIM TOTAL LYMPHS TOTAL T3 POSITIVE TOTAL T-CELL NK ACTIVITY T-HELPER/T-SUPPRESSOR IL 10 TOTAL CD4/CD25 (NOTE FROM STATISTICIAN: THE RELATIVE CHANGES FOR TOTAL LYMPHS, TOTAL T-CELLS, TOTAL T-HELPER, AND TOTAL T3 POSITIVE WERE HIGHLY CORRELATED, INDICATING THAT THESE EFFECTS TEND TO VARY IN A SIMILAR MANNER FOR A PARTICULAR CHILD.) TEST RESULTS 16 WEEK STUDY, ADULT IMMUNE TESTING: STATISTICAL APPROACH: THE ADULT POPULATION WAS TWICE THAT OF THE CHILDREN’S AND THE DATA GENERALLY WERE FAR LESS SCATTERED, INDICATING THAT THE ADULT IMMUNE SYSTEMS WERE MUCH LESS LIKELY TO BE AFFECTED BY UNCONTROLLED VARIABLES DURING THE COURSE OF THE STUDY. BECAUSE OF THIS STABILITY IN THE POPULATION, WE LIMITED OUR ANALYSIS TO THE STANDARD (QUANTITATIVE) t-TESTS. TOTAL CD3+CD26: THERE WAS ON THE AVERAGE A 15% INCREASE IN THIS VARIABLE THAT WAS SIGNIFICANT ON THE 98% CONFIDENCE LEVEL. SIXTEEN (16) PARTICIPANTS MOVED UP, NINE (9) MOVED DOWN, AND THIRTEEN (13) SHOWED NO SIGNIFICANT CHANGE. CD4/CD25: THIS RATIO INCREASED SIGNIFICANTLY ON A 99.9% CONFIDENCE LEVEL. EIGHTEEN (18) ADULT VALUES INCREASED, 12 DECREASED, AND 8 STAYED ESSENTIALLY THE SAME, THERE WERE THREE (3) MORE ABNORMAL RESULTS AT END THAN AT BASELINE. (NOTE: TOTAL CD4/CD25 AND IL-10 UNSTIMULATED WERE HIGHLY CORRELATED) NATURAL KILLER CELL LEVEL: LEVEL INCREASED 14% (WITH A SIGNIFICANCE LEVEL OF 99%). EIGHTEEN (18) VALUES INCREASED, FIFTEEN (15) STAYED ROUGHLY THE SAME, AND FIVE (5) WENT DOWN, ALL VALUES WERE GENERALLY LOW BEFORE AND AFTER THE STUDY. THERE WAS ONE (1) ABNORMAL VALUE AT THE END AND NONE AT THE BEGINNING.

TOTAL T HELPER (CD4+): THIS INCREASED AN AVERAGE OF 13% (99.5% LEVEL OF SIGNIFICANCE). TWENTY-SIX (26) OF THE THIRTY-EIGHT (38) PARTICIPANTS INCREASED THEIR LEVEL WHILE STAYING WITHIN THE NORMAL LIMITS, TWELVE (12) WENT DOWN AND ONE (1) STAYED ABOUT THE SAME. THERE WERE NO ABNORMAL VALUES AT BEGINNING OR END OF STUDY. (DR. VOJDANI SAYS INCREASES ARE GOOD AS LONG AS THEY STAY WITHIN THE NORMAL RANGE). TOTAL SUPPRESSOR CD8+: VALUE INCREASED 13% (99.5% CONFIDENCE LEVEL). TWENTY-SIX OF THE THIRTY-EIGHT MOVED UPWARD. THERE WERE NO ABNORMAL VALUES AT BEGINNING OR END OF THE STUDY. ALTHOUGH BOTH CD4+ AND CD8+ INCREASED SIGNIFICANTLY, THE RATIO, CD4+/CD8+ DID NOT CHANGE SIGNIFICANTLY. THERE WERE NINE ABNORMAL VALUES AT THE BEGINNING AND TEN AT THE END OF THE STUDY. THE REASON THE RATIO REMAINED CLOSE TO ONE WAS THAT THE SUPPRESSOR AND HELPER CELLS BOTH INCREASED PROPORTIONALLY. TOTAL T3 POSITIVE: INCREASED 13% (SIGNIFICANT ON THE 99.5% LEVEL). TWENTY-SEVEN ADULTS MOVED UP AND ELEVEN MOVED DOWN. ALL SCORES BOTH BEFORE AND AFTER WERE WITHIN NORMAL REFERENCE RANGE. OF THE REMAINING SEVEN MARKERS, SIGNIFICANT INCREASES WERE NOTED IN TOTAL LYMPHOCYTES , TOTAL T-CELL , AND TGF BETA 1 UNSTIM . (NOTE: THE RELATIVE CHANGES FOR TOTAL LYMPHOCYTES, TOTAL T-CELL, TOTAL T-HELPER, TOTAL SUPPRESSOR AND TOTAL T-3 POSITIVE WERE HIGHLY CORRELATED.) SUMMARY, ADULTS: IN TERMS OF ABNORMAL VALUES FOR ADULTS: FIVE VALUES HAD HIGHER ABNORMALS AT THE END (CD4+/CD8+ RATIO, TOTAL NATURAL KILLER, TOTAL CD4/CD25, TGF-BETA UNSTIMULATED, AND NK CELL ACTIVITY); SIX VALUES STAYED THE SAME, AND FIVE VALUES IMPROVED FOR THE ADULT GROUP BY THE END OF THE STUDY.

DISCUSSION OF LDN STUDY: LABORATORY MEASUREMENTS OF THE CHILDREN’S IMMUNE SYSTEMS VARIED WIDELY. RANGES WERE BROAD IN ALMOST ALL CATEGORIES. CLEARLY, THERE WERE NOT ENOUGH PARTICIPANTS IN THE CHILDREN’S GROUP TO COME TO MORE THAN A FEW STATISTICALLY SIGNIFICANT CONCLUSIONS. HOWEVER, THERE WERE SEVERAL SUGGESTIVE CONCLUSIONS THAT INDICATE LDN DOES HAVE A POSITIVE EFFECT ON ASD CHILDREN’S IMMUNE SYSTEMS. SOME IMPORTANT MARKERS SUCH AS THE CD4+ CELL COUNT DID GO UP FOR THE MAJORITY OF PARTICIPANTS AND NONE ENDED UP OUT OF THE REFERENCE RANGE OF NORMAL. THE ADULT GROUP WAS LARGER AND SIGNIFICANTLY MORE CONSISTENT IN MAKING IMMUNE SYSTEM IMPROVEMENTS, ALTHOUGH THE AMOUNT OF IMPROVEMENT FOR THE CHILDREN WAS ACTUALLY GREATER THAN THAT FOR THE ADULTS, PROBABLY BECAUSE THEY HAD GREATER IMMUNE IMPAIRMENT GENERALLY. THIS MAY ALSO BE INFLUENCED BECAUSE THE CHILDREN ARE UNDERGOING INTENSIVE TREATMENT WITH SPECIAL DIETS, NUTRIENTS, ANTIPATHOGEN STRATEGIES AND OTHER THERAPIES TO IMPROVE THEIR IMMUNE SYSTEMS (METHYLATION, DETOXIFICATION, HBOT, ETC.) THE VARIABILITY EXHIBITED BY THE CHILDREN MAY BE BECAUSE OF THIS CONSTANT ARRAY OF NEW STRATEGIES, TREATMENT OF FREQUENT GUT INFECTIONS, ETC. GENETIC INFLUENCE WAS EVIDENCED IN THE PARENTAL GROUP BY HISTORY SINCE A SIGNIFICANT NUMBER OF THE ADULTS LISTED ALLERGIES, FATIGUE, AUTOIMMUNE AND GUT ISSUES AS CHIEF COMPLAINTS IN THEIR QUESTIONNAIRE. I DOUBT THAT AS A GROUP THEY ARE SPENDING AS MUCH TIME ON THEIR OWN HEALTH AS THEY SHOULD, EVEN THOUGH MANY WOULD LIKELY BENEFIT BY ATTENTION TO A BETTER DIET, TESTING FOR THE BEST NUTRIENTS FOR THEIR METABOLISM, ETC., ALL OF WHICH THEY DO FOR THEIR CHILDREN. PARENTS OF ASD CHILDREN ARE SUBJECTED TO A LOT OF DEMANDS WHICH STRESS EMOTIONALLY, PHYSICALLY, AND FINANCIALLY, NO DOUBT AFFECTING THEIR IMMUNE STATUS.

IMPORTANT MARKERS SUCH AS THE TOTAL T HELPER (CD4+), THE TOTAL SUPPRESSOR (CD8+) AND CD3+CD26 ALL SHOWED SOME LEVEL OF STATISTICAL SIGNIFICANCE FOR BOTH GROUPS OF ADULTS AND CHILDREN. CLEARLY, MORE STUDIES ARE NEEDED WITH A CONTROL GROUP AND DONE WITH PARTICIPANTS WILLING TO MAINTAIN THEIR CHILDREN’S (AND/OR THEIR OWN) NUTRITIONAL SUPPLEMENT REGIME, DIET AND OTHER STATUS QUO EXCEPT FOR URGENT SITUATIONS. THIS IS CHALLENGING TO DO IN OUR POPULATION WHERE MANY TREATMENTS ARE OFTEN DONE SIMULTANEOUSLY. NOTE ON SAFETY OF LDN: AT THE END OF THIS STUDY IN AUGUST 2006 BEFORE PRESENTING MY FINDINGS TO THE DAN! THINK TANK , I QUERIED THE TWO MOST POPULAR COMPOUNDING PHARMACIES WHICH OFFER THIS MEDICATION TO THE AUTISM COMMUNITY AS TO NUMBER OF PRESCRIPTIONS BEING FILLED. THE INVENTOR OF THE EMU-BASED TRANSDERMAL CREAM, MOLECULAR PHARMACOLOGIST DR. TYRUS SMITH AT COASTAL COMPOUNDING PHARMACY IN SAVANNAH, GA, REPORTED THAT HE HAD GIVEN THE SKIN FORMULA TO 50+ PHARMACIES ACROSS THE UNITED STATES, PLUS PHARMACIES ABROAD WHERE I HAVE TAUGHT (ISRAEL, SCOTLAND, AND HONG KONG). THE ESTIMATE AT THAT TIME FROM THESE QUERIES WAS THAT BETWEEN 4000-5000 CHILDREN WERE USING LDN AT THAT TIME WITH ESTIMATED 75-80% POSITIVE RESPONSE. THE PRIMARY ADVERSE EFFECT ON THESE CHILDREN WAS SOME HYPERACTIVITY OR INSOMINA WHEN THEY FIRST STARTED. THIS HAS ALWAYS SUBSIDED WITH CONTINUING PAST THE EARLY RESPONSES, LOWERING THE DOSE, OR STOPPING. FOR SOME PARENTS THESE SIDE EFFECTS WERE INTOLERABLE AND THEY PREFERRED TO STOP THE MEDICATION. THERE HAVE BEEN NO IRREVERSIBLE NEGATIVE REPORTS OTHER THAN TWO MOTHERS REPORTING THAT ALLERGIES WORSENED WHILE ON LDN, AND THEY BOTH STOPPED WITH GRADUAL RESUMPTION OF THEIR FORMER LEVEL OF ALLERGIC ACTIVITY. INTERESTINGLY, MANY PARENTS HAVE REPORTED ALLERGY LESSENING OR EVEN DISAPPEARANCE, BUT CLEARLY ALL THE CHILDREN ARE UNIQUE. AT THE PRESENT TIME, THE NUMBER OF FAMILIES ON MY “LDN IN AUTISM” E-LIST NUMBERS MORE THAN 1600. HERE PARENTS AND SOME PROFESSIONALS (INCLUDING ME WHEN I HAVE THE TIME) HELP NEW PARENTS GET USED TO THE MEDICATION AND HELP EACH OTHER WITH SIDE EFFECTS, DOSING QUESTIONS AND EXPECTATIONS.) THE MOST OBVIOUS AND IMMEDIATE GOOD EFFECTS OF THIS MEDICATION ARE ON SOCIALIZATION, COGNITION, AND LANGUAGE. IMMUNE BENEFITS ARE LATER, MORE SUBTLE, AND REPORTED BY PARENTS AS “BETTER HEALTH.” INTERESTINGLY, THE GREATEST BENEFIT REPORTED BY THE PARENTS WAS “BETTER QUALITY OF SLEEP.” FOR SOME PERSONS IN THE STUDY, BOTH ADULT

AND CHILDREN, CHANGING THE DIET TO FURTHER RESTRICT CASEIN AND GLUTEN HELPED ELIMINATE THE EARLY HYPERACTIVE RESPONSES.

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