Treating Chronic Fatigue Syndrome (CFS) & Fibromyalgia (FM) by Targeting the Methylation Cycle

1. Treating Chronic Fatigue Syndrome (CFS) & Fibromyalgia (FM)byTargeting the Methylation Cycle Derek Enlander, MD 860 Fifth Avenue New York, NY 10065 USA1-212-794-2000 denlander@aol.com

2. Basic cell process

3. Pathogenisis

4. Virus as Causative Agent • CFS often starts with a flu-like episode • CFS symptoms wax and wane • Antiviral pathways are activated • CFS symptoms are similar to many viral conditions including: • EBV mononucleosis • Ross River virus • Geographic outbreaks have been reported • Gene expression profiling found genetic variants that impact antiviral defenses • Antiviral treatments have been effective in in small studies • Ampligen • isoprinosine • beta interferon • valganiclovir

5. Viruses Implicated In CFS • Human herpesviruses • EBV (HHV-4) • CMV (HHV-5) • HHV-6 • HHV-7 • Stealth Virus (Simian) • Enteroviruses • Polio • Coxsackie A & B • Echovirus • Foamy virus [spuma virus] • Parvo virus • B-19 • Hepatitis-C virus • JHKV • Rubella virus • Ross River virus (RRV) • Inoue-Melnick virus • Borna virus • HERV Human Endogenous Retrovirus

6. HHV6 virus • Discovered in 1986 by Ablashi and Salahuddin at NCI in patients with lymphoproliferative disease( slide by kind permission of Dharam Ablashi) • HHV-6 is an enveloped, double-stranded DNA virus with an icosahedral capsid and virion size of 160-200 nm. The genome contains 70 proteins. Some of these proteins (early antigen and immediate early antigen) can be used to detect active infections. • Two very distinct variants with 90% nucleotide sequence homology. The genome of variant A or B ranges from 159 – 170 kb. • Beta herpesvirus and genus roseola virus • Over 90% of adults seropositive • Predominantly CD4 lymphotropic

7. HHV6 • HHV-6 has two variants, A and B.  Each variant is associated with a subset of illnesses.  • HHV-6A infection comes later in life and has been linked to the pathogenesis of CFS, rhomboencephalitis, and MS. It also plays a role in HIV, causing more immunosuppression in AIDS patients.  • HHV-6B is the causative agent of Exanthem subitum, is strongly linked to infections in transplant patients and is also associated with epilepsy and post-transplant acute limbic encephalitis. • HHV-6A is more lytic, and readily infects a variety of neural and other cells such as astrocytes , and is also more neurotropic, leading to cognitive disorders in CFS and MS patients. • Both A & B strains cause encephalitis, amnesia, facial paralysis, chronic myelitis and transverse myelitis. • HHV-6A can lead to enhanced production of EBV, HSV and HHV-8. Both variants also induce expression of human endogenous retrovirus K-18 encoded super antigen.

8. HHV6 (cont.) • Both strains establish latency in monocytes, macrophages and target CD4 cells for infection. • Since HHV-6, like other human herpesviruses, is ubiquitous, its role in the pathogenesis is established when the virus is in the active state of infection. • Anti-viral agents foscarnet, cidofovir and ganciclovir inhibit CMV infection do not block HHV-6 infection, especially of the A variant as efficiently. • HHV-6 infection contributes to immune suppression by: • Disturbing key immune activation pathways and cytokine networks. • Depleting CD4 T lymphocytes via direct infection and induction of apoptosis (Lusso) • Upregulating TNF alpha, TNF- gamma, IL-1beta and IL-10.(Flamand, Dockerell, Li)and IL-21. • Downregulating complement activity through the CD46 receptorsuppressing macrophages to produce IL-12 stimulated with gamma interferon.(Lusso 2004)

9. Localization of HHV-6 variants in myocardial tissue by IHC Myocardial hhv6 HHV-6 B HHV-6 A Detection of interstitial cells IHC Labeling of cardiomyocyte www.ikdt.com Kühl et al., 2008, submitted

10. Astrocytes obtained from lateral temporal lobe infected with HHV-6 Astrocyte hhv6 Infected astrocyte

11. Antiviral / immuno used for CFS Antiviral Study effective? ampligen Strayer 1994 yes ??? isoprinosine (Immunovir) Pinching 2002 yes alpha 2a interferon See, 1996 yes ? acyclovir (Zovirax) Straus 1990 (5 weeks) no valacyclvir (Valtrex) Lerner, 2001 (6 mos) yes ?? valganciclovir (Valcyte) Lerner, 2006 (6 mos) yes valganciclovir (Valcyte) Montoya, 2006 (6 mos) yes Hepapressin, Immunoprop Enlander 2007 (12 mos) yes

12. Methylation cycle

13. Reasons to Target Methylation Cycle • Paul Cheney, M.D. reported “almost universal” glutathione depletion in CFS in 1999 • Rich Van Konynenburg, Ph.D. reported that CFS symptoms can be explained by glutathione depletion [AACFS 7th Intl. Conf.] • Personal experience with glutathione supplementation and methylation targeting

14. Current Treatment part 1 • Hepapressin Complex • Weekly Injection • Hepapressin • Magnesium Sulphate • Folic Acid • B12 • Calphosan • Glutathione • Trace elements • Clinical trial 1994 • Kutapressin Derivative • History of treatment

15. Current Treatment part 2 • BetaMax • Methyl B12 Spray • Methyl Cobalamine • Vitamin B6 • Lectrolyte • Sodium • Potassium • Calcium • Magnesium • Trace Zinc • Catapult • Picamilon • Cat’s Claw • Glutathione • L-cystine • Trace Selenium • Provides energy • Relieves muscle pain • Clears Brain Fog

16. Current Treatment part 3 • Immunoprop • Glutathione • L-cystine • Picamilon/Ascorbic Acid? • Trace Selenium • Immunoplus • Folinic Acid • Folic Acid • Vitamin B complex • Glutathione • L-cystine • Phosphadylserine • Trace Selenium • Trace Magnesium • Trace Zinc

17. Major Pathways - Methylation Cycle Diet Choline DHF THF Betaine TMG Methyl Synthase (MS) DNA, RNA Zn Protein synthesis, Carnitine SAM Methionine MAT (methionine adenosyltransferase) SAH BHMT Homocysteine B12 Zn, B6 Cystathionine Glutathione

18. Main Role of Methylation Cycle • Methyl Group Source for the body • Coordinate sulfur metabolism • Coordinate production of DNA

19. Methionine-Homocysteine Pathway Methionine S-Adenosylmethionine S-Adenosylhomocysteine Homocysteine B12, B6, Mg Cysteine Cystathionine Toxic Sulphites Glutathione Sulphates

20. Role of Hepapressin Injection Methionine S-Adenosylmethionine S-Adenosylhomocysteine • Folic Acid Homocysteine B12, B6, Mg Cysteine Cystathionine Toxic Sulphites Glutathione Sulphates

21. Antiviral treatment • Hepapressin / Kutapressin / Nexavir • Bovine/Porcine Liver Extract once weekly • Phase I clinical trial 1994 • Invivo antiviral activity • Valganciclovir ..Valcyte 450mg twice daily • Montoya, 2006 • Case Study, Retinal Uveitis + CFS • Modest Effect • Important to identify appropriate CFS patient titer 1:640

22. Gene expression 2005

23. Kerr collaborators

24. CFS Gene microarray

25. Gene Microarray

26. CFS Associated genes

27. Diagnostic slide test

28. GcMaf • GcMaf protein.. Gc Macrophage Activation Factor original work Yamamoto • Reduced by action of Nagalase • Nagalase if increased GcMaf is decreased • Research into ME/CFS serum nagalase levels • Administration of GcMaf injection in research protocol • Cheney researches MAF 314 probiotic yogurt • ME/CFS center researches MAF 878 probiotic

29. Ampligen • Research treatment protocol • Thought to act on the immune system • 5 centers chosen in US • California, New York, Utah, Carolina, Florida • Hemispherx part funding • Patient cost for 6 months $12,000 • Not yet FDA approved

30. Retro Virus XMRV • New retrovirus XMRV testing in Nevada…. Judy Mikovits • Exciting press releases Oct 2009 • Helps patients and doctors stress the physiological aspect of CFS / ME • Harvey Alter, NIH confirms MLV results 2010 • Small number of patients tested, not adequately replicated as yet • Moderation in determining this as causative agent • Researcher battle ensues • Science requests withdrawal of original paper May 2011 • WPI dismisses Dr Mikovits , sues her for stealing her own notebooks , jails her for 4 days

31. XMRV retro virus • Whitmore Pederson Institute, Reno, Nevada • XMRV retro virus study • Science journal publishes article 8th Oct 2009 • Battle ensues by other researchers some confirm, other deny • May 2011 Science asks Judy Miklovits to withdraw article

32. Dr Judy Mikovits

33. Proc. Natl. Acad. Sci. USA Vol. 88, pp. 2922-2926, April 1991 Medical Sciences Retroviral sequences related to human T-lymphotropic virus type II in patients with chronic fatigue immune dysfunction syndrome (Epstein-Barr virus syndrome/infectious mononucleosis/myalgic encephalomyelitis/polymerase chain reaction/in situ hybridization) ELAINE DEFREITAS*, BRENDAN HILLIARD, PAUL R. CHENEY, DAVID S. BELLS, EDWARD KIGGUNDU, DIANE SANKEY, ZOFIA WROBLEWSKA, MARIA PALLADINO, JOHN P. WOODWARD§, AND HILARY KOPROWSKI The Wistar Institute, 3601 Spruce Street, Philadelphia, PA 19104 Contributed by Hilary Koprowski, Nov13, 1990 ABSTRACT Chronic fatigue immune dysfunction syndrome (CFIDS) is a recently recognized illness characterized by debilitating fatigue as well as immunological and neurological abnormalities [Straus, S. E. (1988) J. Inf. Dis. 157, 405412]. Once thought to be caused by Epstein-Barr virus, it is now thought to have a different but unknown etiology. We evaluated 30 adult and pediatric CFIDS patients from six eastern states for the presence of human T-lymphotropic virus (HTLV) types I and II by Western immunoblotting, polymerase chain reaction, and in situ hybridization of blood samples. The majority of patients were positive for HTLV antibodies by Western blotting and for HTLV-II gag sequences by polymerase chain reaction and in situ hybridization. Twenty nonexposure healthy controls were negative in all assays. These data support an association between an HTLV-ll-like virus and CFIDS.

34. Cohort population Initiation 3 months 6 months 9 months Control Placebo 215 48 (2.4) 49 (2.6) 53 (2.4) 50 (2.3) Kutapressin /Hepapressin alone 209 46 (2.6) 57 (2.4) 55 (1.8) 62 (1.5) Kutapressin /Hepapressin complexed 210 47 (2.1) 47 (1.9) 64 (1.3) 74 (0.8) Kutapressin /Hepapressin complexed Immunoprop Immunoplus 216 45 (2.5) 52 (1.7) 73 (0.9) 81 (0.5) Hepapressin complex treatment Karnofsky & Fatigue scores

35. Ongoing research • GcMaf / Nagalase…. Collaboration with Dr. Kenny deMeirleir • GcMaf Study in treatment of CFIDS/ME patients • Ampligen treatment study • Part Sponsored by Hemispherx Inc • B12 changes with BetaMax (sublingual methyl B12) • Average increase 56% after avg. 3 months of use • Retrospective analysis of Carnitine in CFS • 120 patients, randomly selected serum Carnitine Low • Analysis of urinary H2S with Lead Arsenate • 70 patients, randomly selected 35% positive Lead arsenate analysis

36. Proposed Research • Retuximab • CMX 1000 • Enbrel • Pre and post Exertional Malaise study

37. Retuximab Study • Norwegian Study Oct 2011 • ME/CFS Patients with lymphoma • ME/CFS seemed to improve after chemo • Initial study 5 patients • Followup with 15 patients 15 controls

38. Retuximab Study • Patients have to be hospitalised overnight • Retuximab is a chemotherapeutic infusion • Administered monthly for 6 months • Not FDA / GMC approved • Past side effects including fatal outcome • Relapse 6 months after infusion completion • Reversed after second infusion

39. The UK and Ireland scene • Problems in finding the diagnosis • If diagnosis is made here or elsewhere problem in finding suitable care and treatment • Doctors are only given psychiatric methods of treatment CBT & GET • Research into physical aspect of ME/CFS is neglected • Disability is undercompensated • National cost in GNP is estimated at 10,000 million pound annually

40. The UK and Ireland scene • Provide research funds to physical diagnosis and treatment • Provide training to young doctors • Provide information to old doctors • Set up Central and regional ME/CFS centres to diagnose and treat this physical disease • Reverse old ideas of psychiatric treatment

41. Immediate Provisional solution • Initiation and funding of two fellowships in the diagnosis and treatment of ME/CFS by the Government and/or Private sources • We have set up the fellowship training for 12 months for two post Doc physicians • The training will be provided at the ME/CFS Centre in New York • Dr Eric Schadt will supervise the research part of the fellowship • Dr Derek Enlander will supervise the clinical part of the fellowship

42. ME/CFS Center Mount Sinai Medical Center, New York set up in Nov 2011 with a donation of one million dollars by a patient of Dr Enlander First ME/CFS centre in a major School of medicine • Research in Genomics Eric Schadt • Immunology Miriam Merad • Virology Ila Singh • Pulmonology Christian Becker • Clinical diagnosis and treatment Derek Enlander and David Bell