Monday, September 13, 2010

XMRV Conference Recap


One thing that resonates from the two-day XMRV conference at the National Institutes of Health is the, at times, testy battle between two camps of scientists. The cohort that believes that mouse retrovirus contamination accounts for the positive XMRV and related murine leukemia virus studies was pitted against the cohort that believes contamination has been ruled out and that the positive studies show a clear link between this family of retroviruses and Chronic Fatigue Syndrome and prostate cancer.  Not surprisingly, the discussions around CFS grew more heated than those around prostate cancer.  Dr. Mary Kearney of the National Cancer Institute may be the one to resolve this issue, as she reported on a test that discriminates between mouse retroviruses and XMRV with, she said,100 percent accuracy.

Despite several positive retroviral findings, by the time the Q and A began at the end of the second day, the upbeat mood many felt due to the recently published FDA/NIH/Harvard murine leukemia virus study—which found XMRV-related MLV viruses in 86.5 percent of CFS patients—had morphed into uncertainty.  Dr. John Coffin of Tufts University, however, reassured the audience that it takes time to grind the sausage—his oft-used phrase—and that the XMRV findings were, in fact, moving along at a brisk clip.

Lyme expert Dr. Joseph Burrascano asked one of the most salient questions of the conference that so far has no definitive answer:  Why are some scientists finding XMRV and related MLVs in most of their samples while others aren’t finding it at all?  What is responsible? Is it the reagents, the collection, the processing, the methods, the patient cohorts or a combination of the above?  The representative of the Blood Working Group suggested that blood collection and processing may hold the key. Dr. Judy Mikovits echoed that sentiment.

Here are some highlights of the conference:

--Dr. Sam Chow of UCLA discussed data from a Chinese study of men with prostate cancer and healthy controls. Of the 117 controls, 17 or 14.5 percent were positive for one of several types of MLVs, including XMRV, polytropic MLVs and modified polytropic MLVs.  (Polytropic viruses infect the original host—in this case mice—as well as other species, whereas xenotropic viruses like XMRV infect species other than the original host.)  Of the 34 men with prostate cancer, five or 14.5 percent were also positive.  One reason for the hefty numbers in controls may be that Chinese keep mice as pets. Chow said that older men were more likely to have an MLV than younger men.  In addition, Chow reported that XMRV has entered the blood supply in China: 3.4 percent of blood donors were positive for XMRV. There was no association of the RNASE-L genetic defect in the Chinese study.


In contrast, Dr. Robert Silverman of the Cleveland Clinic and co-author of the first study linking prostate cancer to XMRV,  talked about the RNASE-L mutation in his cohort.  In one family of five sons, for instance, four had prostate cancer, along with the RNASE-L mutation.  He also discussed the study he coauthored on macaques infected with XMRV.  In these animals, he explained, the prostate epithelium is infected early on, followed by the seminal vesicles and epididymis, and finally the stromal fibroblasts. Prostate cancer is often preceded by inflammation and XMRV stimulates proinflammatory genes, so it’s conceivable, he said, that XMRV infection could cause inflammation and eventually prostate cancer. The enzyme APOBEC3G, a potent inhibitor of XMRV, mutated XMRV DNA in blood cells of the macaques. Silverman also said that androgen stimulates XMRV, and anti-androgens inhibit the retrovirus.

--Dr. Eric Klein, chair of the Glickman Urological and Kidney Institute at the Cleveland Clinic and co-author of the first study linking prostate cancer to XMRV, discussed his findings on prostate cancer and XMRV. Klein said that XMRV is in urine.  He also gave possible explanations for why some groups haven’t found the retrovirus, including geographical differences, which is what is seen with another human retrovirus, HTLV. Klein stressed that PCR “details” are important in doing studies correctly.

He also discussed how most prostate cancer is relatively benign: though 17 percent of men get the disease, only 3 percent will die from it.  But scientists can’t always predict which prostate cancers need to be treated aggressively, and as it stands now, 90 percent of men with the disease opt for treatment.  Treatment, however, carries it own constellation of possible long-term complications, including incontinence and erectile dysfunction.  Perhaps, Klein said, the XMRV positive patients will prove to be among those who should be treated. Klein said given that many viruses have been shown to cause cancer, “It’s not unreasonable to assume that the retrovirus XMRV could cause cancer.”  Mutations, he said, in RNASE-L and MSR1 genes are associated with an increased risk of prostate cancer. In addition, eunuchs may have a decreased risk of prostate cancer not because of their diminished production of testosterone, but due to the fact that they’re not exposed to pathogens through sex. 


--Dr. Ila Singh of the University of Utah and co-author of a study linking XMRV to prostate cancer voiced that XMRV may become a marker for prostate cancer.  It’s conceivable, she said, that XMRV status may eventually prove to be a better indicator than the current PSA test, and perhaps antiretroviral therapy will be effective in treating the disease. She explained that the XMRV gag (the basic physical infrastructure of the retrovirus) cross-reacts with Moloney mouse MLV.  However, the envelop (the retroviral protein that makes up the outside of the retrovirus) reacts only to XMRV.  Of 233 men with prostate cancer in her study, 27 percent were XMRV positive.  Interestingly, the higher the Gleason score—which is used to determine the prostate-cancer prognosis and ranges from two to 10—the more likely the men were to be infected with XMRV. 

Singh’s CFS study involves 200 healthy controls and 105 patients, all from the Utah area. Singh said she won’t use nested PCR, because in her view contamination can result.  Other ways to reduce contamination:  Handle only one specimen at a time, and have a negative control.  Her tests, she said, are very sensitive at a 97.5 percent detection rate, with a specificity of 100 percent (i.e., no cross reaction with other retroviruses).

--Dr. Maureen Hanson of Cornell University is studying 30 subjects, with a mean age of 39.  Ten of those patients supplied by longtime CFS physician Dr. David Bell were severely ill with CFS, with less than three hours of daily upright activity.  Another ten had recovered from CFS, with 13.5 hours of upright activity daily. And, finally, 10 healthy controls weighed in with more than 15.5 hours of daily upright activity.  Hanson’s findings are consistent, she said, with the Lombardi Science study (which found XMRV in 67 percent of CFS patients) and the recent FDA/NIH/Harvard MLV study (which found XMRV-related MLVs in 86.5 percent of patients). She also said that she’s never worked with mice in her lab, making contamination with a mouse retrovirus less likely. 

--Dr. Judy Mikovits of the Whittemore Peterson Institute and principal investigator of the first study linking XMRV to CFS talked about a London CFS cohort (and then, I believe, it was British researcher Dr. Jonathan Stoye who corrected her and said that the cohort was nearer to Kent) who met the Canadian consensus and suffered from severe cognitive dysfunction, joint pain, vertigo and tender lymph nodes; more than 50 percent were homebound. The predominant MLV was XMRV, and the rate of infection among CFS patients was 70 percent.  Fifty blood donors served as controls. Dr. Jonathan Kerr (who, like Jonathan Stoye, was an author on the second negative British XMRV CFS study) drew their blood. Four percent of the controls were positive for XMRV.

--Dr. Mary Kearney of the National Cancer Institute discussed the assays she developed to detect XMRV, which she said can discriminate between XMRV and mouse retroviruses.  She explained that  mouse retroviruses emit a lower level of florescence than XMRV.  She claimed she can distinguish between the two with 100 percent accuracy. Perhaps her assay will be used to render a final verdict on the mouse-contamination theory.

--Dr. Ross Molinaro of Emory University, co-author on the first study linking XMRV to prostate cancer, said that XMRV encodes a previous unknown Orf3 protein with punctate patterns.  In macaques, he explained, the prostate epithelium is infected early on, followed by the seminal vesicles and epididymis, and finally the stromal fiberblasts. 

--Dr. Francois Villinger of Emory University chronicled the infection of macaques with XMRV.  He said their CD4, CD8 and natural killer cells were infected and was surprised that monocytes were not.  "The viral load in blood was contained very quickly," he said. Chronic infections, Villinger said, were seen in the spleen, epididymis, macrophages in the lung, the seminal vesicles, cervix and vagina. Villinger said he didn't look at the breast or spinal cord.  Someone in the audience brought up screening sperm banks for MLVs.

--Dr. Jonas Blomberg of Uppsala University in Sweden looked at CFS and fibromyalgia in Swedish patients. Judy Mikovits supplied five samples, and Blomberg found three to be positive, so he was able to detect the retrovirus, just not in the Swedish cohort. Of the 35 CFS patients and 15 fibromyalgia patients, all were negative. 

--Dr. Graham Simmons of the Blood Working Group discussed the different phases of study for the blood people.  Phase 1:  assay evaluation. Of all the assays, he said, the CDC’s and the FDA’s were the most sensitive. Phase 2: pilot clinical study.  Phase 3: clinical sensitivity specificity.  Phase 4: blood donor clinical panel. While phase 2 to phase 4 are not complete, “We do think we have begun to figure it out,” Simmons said and indicated that proper collection and processing of blood are vitally important.

--Dr. Nicole Fischer of University Medical Center Hamburg-Eppendorf found XMRV in one out of 300 men with prostate cancer and one out of 70 controls in Germany.

--Dr. Shyh-Ching Lo of the FDA talked about the findings in FDA/NIH/Harvard paper, which he co-authored, and said that different primers yield different results.

--Dr. Brigitte Huber of Tufts University focused on contamination as the cause of XMRV positive samples. Veteran CFS physician Dr. Susan Levine supplied the 111 patients, all of whom met the Fukuda CFS definition; 25 percent were severely disabled. Only one of the 111  patients—less than 1 percent—proved positive. 

Last fall, Huber collected new blood:  three CFS samples and 36 healthy controls.  Using  nested gag PCR, two out of the three CFS patients and 17 of the 36 controls were positive. Twelve samples were negative. Mouse DNA probes detected mouse DNA in those 17 positive samples from CFS patients, and Huber called the correlation “shocking.”  Her conclusion was that a common lab reagent is contaminated with mouse DNA.  Most of her samples contained several MLVs.  Her lecture drew criticism from several members of the audience.  One outspoken critic shouted to Huber that heparin tubes were a “nightmare.  Why are you still using it?” The heparin naysayers used EDTA tubes instead.

--Bill Switzer, lead investigator of the CDC’s negative XMRV CFS study, reported on the agency’s prostate cancer study of 162 patients.  Switzer said that the CDC found a gag sequence in one and pol sequences in three. (Pol proteins synthesize viral DNA.)  Other scientists are looking at RNA, Switzer said, and the CDC is looking at DNA.  Switzer made the point that the Robert Koch Institute in Germany didn’t find XMRV in prostate-cancer patients either. 

--Dr. Frank Ruscetti of the National Cancer Institute and co-author of the first study linking XMRV to CFS said that there are 17 million people in the world with Chronic Fatigue Syndrome, and all of them have contacted Judy Mikovits.  That line drew a huge laugh from the audience.  Single round PCR, he explained, locates only 15 percent of cases, whereas nested PCR finds 69 percent. XMRV reservoirs, he said, have not yet been identified. One patient who was studied had both XMRV and a polytropic MLV.  When I asked if that person was more ill, Judy Mikovits replied yes.


Coming up:  A look at the salient posters presented at the conference.

This article is copyright CFS Central 2010. All Rights Reserved. You may quote up to 150 words from this article as long as you indicate in the body of your post (as opposed to a footnote or an endnote) that the excerpt is by Mindy Kitei for CFS Central. You may not reprint more than 150 words from this article on blogs, forums, websites or any other online or print venue. Instead, refer readers to this blog to read the article.

38 comments:

  1. Mindy, you've given us a cornucopia of new data.
    Teeth-grinding frustration reading your opening paragraphs, but then:
    - wow, 14.5% in Chinese prostate cancer patients AND controls.
    - Eric Klein: eunuchs? ah, now how big was that cohort? ok, not a topic of humour, but I'm laughing all the same. perhaps Dr. Klein prefers cohorts of eunuchs as they'd be no competition whatsoever for his dream of playing opposite Angelina Jolie in 'XMRV: The Movie'? (wink, wink, all in good fun, doctor)
    - Klein and Singh on XMRV as better-than-PSA marker for prostate cancer prognosis, this is a great advance, excellent news.
    - Hanson-Bell cohort and Mikovits UK-cohort, solid positive numbers!! and 4% positive UK controls!
    - Molinaro, 'XMRV is stimulated by inflammation', scary news, whether cause or result of inflammation.
    - Villinger, 'CD4, CD8 and natural killer cells were infected'.
    - Blomberg, sadly negative results, but delighted that he confirmed 3 positives in WPI samples.
    - Ruscetti, such a wag, gotta love him.
    Awaiting your next installment with interest, Mindy. THANK YOU!

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  2. RE: Dr Mikovits and the UK sample

    The patients had their blood collected from a Hospital in Ashford. Dr Stoye is getting confused with Ashford in Kent.

    This Ashford is on the border of South London and Surrey. The postcode is Middlesex.

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  3. Thanks for making the trip, pressing the issue of the CDC CFS definition, which is central to the 'cohort issue' and for this summary. I wish we could hear more about what was discussed (especially the efforts by Ruscetti and Cheney) but hopefully some things will be coming out of embargo soon.

    Thanks as always
    Otis

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  4. Than you for your excellent coverage, Mindy! It was well worth the wait. What would we do without you?

    It is so thrilling to see CFS finally being taken seriously by the medical/scientific community.

    Real data! Debate rooted in data! What a concept!

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  5. Thank you, Mindy for your continuing coverage. This is the first review of the conference that's been available and it was well worth the wait!

    I am 33 years into this illness and it is heart warming to see CFS finally being considered by the medical/scientific community.

    Real data! Debate based on data! What a concept!

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  6. Thanks for this summary of the conference Mindy.

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  7. Thank you, Mindy. Fascinating stuff.

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  8. supercalifragilisticexpedaliocious

    fabulous, stupendous, wonderful....all the words in the language to give you huge praise. this report was like a breath of fresh air after years of breathing noxious fumes. thank you for being there, asking questions and reporting back what you heard and learned.

    i am sooooo grateful to you and all of the hardworking scientists and dr's trying to crack the code on this frackin' disease....

    loved that huber was crticized from the audience for her use of heparin tubes.

    love ruscetti's joke that all 17 million of us cfids sufferers have contacted judy....the beautiful thing abt judy is that she has responded to all of us with concern, empathy and intelligence.

    thank you mindy!!!!!

    p.s. when i tried to log in thru firefox...a message told me i needed to be invited to see your blog. i was allowed to enter through safari...hmmmm.

    anyway i am sooooo glad to have read your post.

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  9. Wow! I didn't expect you would be allowed to publish so much Mindy, great job. We all saw you at the Q and A, and like many, I was shocked by the rudeness of the chair when he declaired 'nobody was looking for a negative result'. Scientists are so naive when it comes to the influence of politics and economics in their trade.

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  10. Thank you so much for being persistent and asking the questions that, apparently, no one wanted asked or answered. I am on the edge of my seat waiting to hear about the poster section.
    Thank YOU!

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  11. Thank you for continuing to cover the developing science and bizarre politics of CFS, XMRV and now PMRVs and for doing so with clarity, depth, objectivity and integrity. Rare!

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  12. Thanks Mindy!

    Sounds like the Cornell study could be a huge confirmation result. Do you know if it was done completely independently of WPI (e.g. no materials from the WPI were used)?

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  13. Excellent, excellent coverage. I am so grateful to you!!!

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  14. Thank you Mindy, this was really interesting reading!

    Anne, Norway

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  15. Otis, I will be reporting on Cheney's poster, and Anonymous, I don't know if the Cornell study used any materials from WPI.

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  16. Thank you for a fascinating piece. I learn a ton from your work.

    And thanks for using a larger font today. My vision is impaired by CFS or whatever and the font size makes big difference in my ability to read.

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  17. Mindy, many many thanks for some excellent journalism and reporting what you saw and heard. All of the comments above are fantastic. It is just about time we are being heard and taken seriously. Thanks for providing a voice.

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  18. Funny fact that I read on your blog post, XMRV and prostate: doctors are worried about maintaining erectile function. XMRV and CFS, patients are concerned about standing up.

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  19. @Mindy: Amazing piece of work, Mindy. You know we all loooove you, don't you? Cannot wait to see the next installment.
    Thank you so much for "being there"!

    @Katie: ROFLOL....you, know if men can't get a hard-on with XMRV, maybe women can't get a wide-on with it, either! Equal opportunity Retro, right?
    Sorry for being so gross, couldn't help myself! Am not usually like this!

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  20. Thank you so much Mindy,
    we so look forward to your reports.
    Wasn't the rude chairperson who said 'none of us is looking for a negative result' one of those who had in fact been involved in a study finding a negative result.
    Rude ignorant man.

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  21. Thanks a lot, Mindy.
    That's exactly what i was looking for, a summary of all the work presented at the workshop (or at least of that part that can be reported on).
    One small question mark, i think the Lombardi et al. Science study used heparin tubes too. And Dr. Vernon criticized the negative CDC study for not using heparin tubes. http://www.cfids.org/xmrv/070110study.asp So there seem to be some conflicting opinions there. But nevertheless i'm not too much worried about contamination because i think both positive studies (Lombardi et al. and Lo et al.) have done a lot to prove that there's no contamination.
    I'm a bit disappointed that Dr. Blomberg couldn't find any XMRV in his subjects, especially as he could find it in the WPI samples. But maybe the viruses in Sweden are different from the ones in the USA (there's quite a distance) and this can explain it.
    Great to hear about the Hanson study.
    Stoye was really unbearable. And then he's even wrong about Ashford... Lol... what a joke.

    Eric

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  22. I wonder what tubes Blomberg used...

    Thanks for all this, Mindy.

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  23. Thanks Mindy. Just to repeat the earlier comment that the Ashford in question is in Middlesex, and not the better known Ashford in Kent as Dr Stoye incorrectly assumed. Patients attending the Ashford blood draw were from London and surrounding areas as Dr Mikovits stated.

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  24. Mindy, did Huber mention amplifying (ie PCR-detecting) any *other* mouse sequences from reagents or humans? Or, even, did she detect MLVs in the reagents alone? What she is claiming is possible - but those sort of results are indisputably (and to a well-trained researcher, quite obviously) what is needed in order to significantly support her hypothesis, and also in order to bring out (in a strong journal) a paper making a significantly strong claim that this is true. Especially when we know that Alter has performed some such assays, attempting to detect mouse mitochondrial DNA, and did not get the results predicted by Huber. Mitochondrial DNA ought to be exquisitely easy to detect, because there is one 'regular' genome in each mammalian cell, vs something like 1,000 mitochondrial ones.

    In my opinion, the Huber hypothesis merits attention, but is fairly unlikely to be true when about eight labs have found a 15-20x difference in HMLV prevalence by PCR, between CFS patients and normals. If the 'goods' are coming from the reagents, there shouldn't be a difference between patients and controls. Unless there is some PCR enhancing agent in DNA preps made from CFS blood, or a lack of some PCR inhibitor! This seems unlikely, but in any case it is simple to test by experiment. Or, one lab might be biased and subconsciously treat the patient and control samples non-identically (hence the desire that the sample tubes, optimally, be marked in a code unknown to the worker). But it is steeply unlikely that so many different labs could be so biased.

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  25. I'm tired please summarise to seven words.

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  26. Erich Jansohn, Excellent questions, but I don't have the answers to most of them. Huber did say she found gag and IAP mouse sequences, and one other sequence that I didn't hear. I did ask Huber when her paper was coming out and in what journal, but she didn't answer those questions.

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  27. > but those sort of results are indisputably (and to a well-trained researcher, quite obviously) what is needed in order to significantly support her hypothesis

    By the way, I didn't mean to imply that Prof. Huber doesn't know left from right. I've heard that her reputation is good - and researchers do not assume another researcher to be highly credible by default! They assume another researcher to be quasi-credible at best, until proven otherwise.

    I did mean to imply that any pro researcher, including her, either would or really really *should* agree that you need the results I mentioned before a hypothesis like hers can be considered quite strong - strong enough to be not just a problem, but a serious problem, for contradictory hypotheses. I'm sure that like 90% of research professors, and probably Huber herself, would agree with me.

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  28. Mindy, thank you so much for that! It was so interesting to read!
    Also - millions of thanks for your arrival to the conference and your great question about the CDC which was brutally rejected by Dr. Stoye. He should be ashamed at himself - because what you have said was the truth.

    By the way, about Huber not answering your question regarding to "in what journal this would be published?" - I think that might be because - as I've heared - at least two journals have turned down her request for publishment of her so-called "research".

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  29. @Eric Johnson: It seems like it would be easy to test Huber's hypothesis that its the reagents that are contaminated. Just test the reagents alone, without putting them anywhere near a blood sample.

    Also, if they suspect that heparin is contaminated, they can do the same.

    I agree though, the discrepancy in results between the CFS patients and control in a blinded study (which the Science one was) makes it unlikely that either of these are the case. If the heparin in the tubes was contaminated, then you should see the same false positive rate in the control population. Same with the reagents. They use the same reagents and tubes on both CFS and controls.

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  30. Mindy, thanks so much for the reporting and all your hard work. It is much appreciated.

    I haven't seen any information about Ellen Sparger's talk and am very curious to know what she said. There shouldn't have been anything embargoed there (I presume)...?

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  31. theparsley, I didn't take many notes during Ellen Sparger's talk because vaccine development is such a pipe dream at this point. Look at HIV vaccine development after all these years of working on a vaccine. Sparger talked about feline leukemia virus--for which there is a vaccine--and its similarity to MLV. She explained that newborn kittens are more susceptible to FLV and adult cats eliminate the virus more easily.

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  32. Mindy,
    Did anyone address testing non-blood tissue? Since the rhesus macaque studies showed the viral load clearing quickly from the blood, but the retrovirus lodging indefinitely in the tissue, particularly reproductive organs. Needle biopsies, anyone? Why or why not?
    Otherwise, thank you for calling a spade a spade.

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  33. Mindy,
    Is there any word on when Dr. Kearney's (NCI) assays will be available for clinical use? (Tails are wagging in anticipation!) Did she mention expense? Has to be less $$$ than a tissue sample, yes?

    Thank you for the summary and for speaking up on our behalf at the Q&A!

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  34. I saw your question during the Q&A. A brave attempt. Although the moderator tried to shame you off the face of the Earth, your question was perfectly relevant to "the science." Even apart from fraud or deliberate "trying not to find the virus," in the moderator's derisive phrase, investigator bias or "expectation bias" is a standard concern in science -- as Q&A remarks referring to "independent" labs implicitly acknowledged.

    According to "A selected history of expectation bias in physics" (Am. J. Phys. 74(7), July 2006 (PDF available on request from me at lgilman [at] myfairpoint [dot] net), expectation bias is not only "well known and widely discussed in psychology and medicine" but affects physics, too, that hardest-core of sciences. In a well-known example of one kind of expectation bias, the "bandwagon effect," typical measurements of the speed of light jumped suddenly in 1900 to a lower value "many error bars from the previously accepted value."

    The ways in which expectation or desire can influence results are often obscure, because science is complex and human behavior is subtle: in the case of the speed-of-light bandwagon, how investigator bias was translated into numerical error is still not clear (though it is not thought to have been fraud). In elaborate, long-chain observational efforts such as virus hunts, there are scores of opportunities for observer bias to creep in. Persons who do not expect to find the virus will, to some extent, be more likely to not find it: and vice versa. This is exactly why independent testing, blinding of samples, etc. are employed.

    In the case of the CDC, where an institutional bias of extraordinary intensity has been manifest over many years, one must logically anticipate a particularly strong expectation bias. One would therefore be obliged to scrutinize the CDC's methods with particular care -- as you were trying to do.

    What was nonscientific was not your question, but the moderator's angry pretense that bias is not part of scientific process.

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  35. I second Larry Gilman's outstanding comment.

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  36. Mindy,

    Do you know if Maureen Hanson's study used the WPI/VIPdx for testing or if the testing for that study was done in house? To me one of the biggest steps is for more outside labs to replicate the findings independantly; it doesn't seem quite as robust if everything has to go through WPI/VIPdx (or even Redlabs, which uses the WPI's protocol), which I imagine Cheney's, DeMeirleir's and Mikovits's XMRV+ studies did.

    Thanks a bunch, great reporting.

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  37. @Martha, you say, "My vision is impaired by CFS or whatever and the font size makes big difference in my ability to read. "

    Did you know that you can get your browser to enlarge the display for you, or even enlarge just the text? In Firefox the command is under the View menu, use the Zoom submenu. Don't know about other browsers.

    @Mindy - sorry to hijack your thread, but couldn't find another way of getting hold of Martha. Or not one that the Brain Fog would let me see!

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  38. Thank you Mindy from the bottom of my heart for such consise and thorough reporting...us in the UK are waivering in hope and after reading this I shall be printing it out, waving it in front of my doctors face. The truth will out thanks to you, Judy and all those other heroes tireslessly working on behalf of those who are too sick to try. xxx

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