Pharmacokinetic and safety profile of raltegravir and ribavirin, when dosed separately and together, in healthy volunteers

Results: No statistically significant differences in PK parameters were observed for raltegravir between phases 2 and 3. A statistically significant decrease in maximum plasma concentration (Cmax) and an increase in time to maximum plasma concentration (Tmax) were observed for ribavirin in phase 3 compared with phase 1 [GMR (95% confidence interval) 0.79 (0.62 –1.00) and 1.39 (1.08 –1.78), respectively], whereas no significant differences in other ribavirin PK parameters were observed between study phases. No clinically significant safety concerns were reported. Conclusions: The PK profile of ribavirin is altered when administered with raltegravir (reduced Cmax and increased Tmax), with no safety concerns identified. This is unlikely to be of clinical significance or have an impact on the antiviral effects of ribavirin in HIV-1- and HCV-co-infected subjects

No Evidence of XMRV or MuLV Sequences in Prostate Cancer, Diffuse Large B-Cell Lymphoma, or the UK Blood Donor Population

Xenotropic murine leukaemia virus-related virus (XMRV) is a recently described retrovirus which has been claimed to infect humans and cause associated pathology. Initially identified in the US in patients with prostate cancer and subsequently in patients with chronic fatigue syndrome, doubt now exists that XMRV is a human pathogen. We studied the prevalence of genetic sequences of XMRV and related MuLV sequences in human prostate cancer, from B cell lymphoma patients and from UK blood donors. Nucleic acid was extracted from fresh prostate tissue biopsies, formalin-fixed paraffin-embedded (FFPE) prostate tissue and FFPE B-cell lymphoma. The presence of XMRV-specific LTR or MuLV generic gag-like sequences was investigated by nested PCR. To control for mouse DNA contamination, a PCR that detected intracisternal A-type particle (IAP) sequences was included. In addition, DNA and RNA were extracted from whole blood taken from UK blood donors and screened for XMRV sequences by real-time PCR. XMRV or MuLV-like sequences were not amplified from tissue samples. Occasionally MuLV gag and XMRV-LTR sequences were amplified from Indian prostate cancer samples, but were always detected in conjunction with contaminating murine genomic DNA. We found no evidence of XMRV or MuLV infection in the UK blood donors

Investigation into the Presence of and Serological Response to XMRV in CFS Patients

The novel human gammaretrovirus xenotropic murine leukemia virus-related virus (XMRV), originally described in prostate cancer, has also been implicated in chronic fatigue syndrome (CFS). When later reports failed to confirm the link to CFS, they were often criticised for not using the conditions described in the original study. Here, we revisit our patient cohort to investigate the XMRV status in those patients by means of the original PCR protocol which linked the virus to CFS. In addition, sera from our CFS patients were assayed for the presence of xenotropic virus envelope protein, as well as a serological response to it. The results further strengthen our contention that there is no evidence for an association of XMRV with CFS, at least in the UK

A cautionary tale of virus and disease

The recent identification of the gammaretrovirus XMRV and a second gammaretrovirus of a different subtype in chronic fatigue syndrome has aroused much interest, not least among sufferers. However, it remains highly controversial whether the detection of these viruses represents true infection or laboratory artifacts

Detection of a gammaretrovirus, XMRV, in the human population: Open questions and implications for xenotransplantation

XMRV (xenotropic murine leukaemia virus-related virus) is a gammaretrovirus that has been detected in human patients with prostate carcinoma, chronic fatigue syndrome (CFS) and also in a small percentage of clinically healthy individuals. It is not yet clear whether the distribution of this virus is primarily limited to the USA or whether it is causally associated with human disease. If future investigations confirm a broad distribution of XMRV and its association with disease, this would have an impact on xenotransplantation of porcine tissues and organs. Xenotransplantation is currently being developed to compensate for the increasing shortage of human material for the treatment of tissue and organ failure but could result in the transmission of porcine pathogens. Maintenance of pathogen-free donor animals will dramatically reduce this risk, but some of the porcine endogenous retroviruses (PERVs) found in the genome of all pigs, can produce infectious virus and infect cultured human cells. PERVs are closely related to XMRV so it is critical to develop tests that discriminate between them. Since recombination can occur between viruses, and recombinants can exhibit synergism, recipients should be tested for XMRV before xenotransplantation

Multiple Sources of Contamination in Samples from Patients Reported to Have XMRV Infection

Xenotropic murine leukemia virus (MLV)-related retrovirus (XMRV) was reported to be associated with prostate cancer by Urisman, et al. in 2006 and chronic fatigue syndrome (CFS) by Lombardi, et al. in 2009. To investigate this association, we independently evaluated plasma samples from 4 patients with CFS reported by Lombardi, et al. to have XMRV infection and from 5 healthy controls reported to be XMRV uninfected. We also analyzed viral sequences obtained from supernatants of cell cultures found to contain XMRV after coculture with 9 clinical samples from 8 patients. A qPCR assay capable of distinguishing XMRV from endogenous MLVs showed that the viral sequences detected in the CFS patient plasma behaved like endogenous MLVs and not XMRV. Single-genome sequences (N = 89) from CFS patient plasma were indistinguishable from endogenous MLVs found in the mouse genome that are distinct from XMRV. By contrast, XMRV sequences were detected by qPCR in 2 of the 5 plasma samples from healthy controls (sequencing of the qPCR product confirmed XMRV not MLV). Single-genome sequences (N = 234) from the 9 culture supernatants reportedly positive for XMRV were indistinguishable from XMRV sequences obtained from 22Rv1 and XMRV-contaminated 293T cell-lines. These results indicate that MLV DNA detected in the plasma samples from CFS patients evaluated in this study was from contaminating mouse genomic DNA and that XMRV detected in plasma samples from healthy controls and in cultures of patient samples was due to cross-contamination with XMRV (virus or nucleic acid)

False negative results from using common PCR reagents

Background\ud The sensitivity of the PCR reaction makes it ideal for use when identifying potentially novel viral infections in human disease. Unfortunately, this same sensitivity also leaves this popular technique open to potential contamination with previously amplified PCR products, or "carry-over" contamination. PCR product carry-over contamination can be prevented with uracil-DNA-glycosylase (UNG), and it is for this reason that it is commonly included in many commercial PCR master-mixes. While testing the sensitivity of PCR assays to detect murine DNA contamination in human tissue samples, we inadvertently discovered that the use of this common PCR reagent may lead to the production of false-negative PCR results.\ud \ud Findings\ud We show here that contamination with minute quantities of UNG-digested PCR product or any negative control PCR reactions containing primer-dimers regardless of UNG presence can completely block amplification from as much as 60 ng of legitimate target DNA.\ud \ud Conclusions\ud These findings could potentially explain discrepant results from laboratories attempting to amplify MLV-related viruses including XMRV from human samples, as none of the published reports used internal-tube controls for amplification. The potential for false negative results needs to be considered and carefully controlled in PCR experiments, especially when the target copy number may be low - just as the potential for false positive results already is

No Evidence of Murine Leukemia Virus-Related Viruses in Live Attenuated Human Vaccines

The association of xenotropic murine leukemia virus (MLV)-related virus (XMRV) in prostate cancer and chronic fatigue syndrome reported in previous studies remains controversial as these results have been questioned by recent data. Nonetheless, concerns have been raised regarding contamination of human vaccines as a possible source of introduction of XMRV and MLV into human populations. To address this possibility, we tested eight live attenuated human vaccines using generic PCR for XMRV and MLV sequences. Viral metagenomics using deep sequencing was also done to identify the possibility of other adventitious agents.All eight live attenuated vaccines, including Japanese encephalitis virus (JEV) (SA-14-14-2), varicella (Varivax), measles, mumps, and rubella (MMR-II), measles (Attenuvax), rubella (Meruvax-II), rotavirus (Rotateq and Rotarix), and yellow fever virus were negative for XMRV and highly related MLV sequences. However, residual hamster DNA, but not RNA, containing novel endogenous gammaretrovirus sequences was detected in the JEV vaccine using PCR. Metagenomics analysis did not detect any adventitious viral sequences of public health concern. Intracisternal A particle sequences closest to those present in Syrian hamsters and not mice were also detected in the JEV SA-14-14-2 vaccine. Combined, these results are consistent with the production of the JEV vaccine in Syrian hamster cells.We found no evidence of XMRV and MLV in eight live attenuated human vaccines further supporting the safety of these vaccines. Our findings suggest that vaccines are an unlikely source of XMRV and MLV exposure in humans and are consistent with the mounting evidence on the absence of these viruses in humans