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closeInfectious virus in VCaP cells
Posted by DustyMiller on 19 Jun 2011 at 07:17 GMT
In their Results, the authors state,
"CWR22Rv1 has previously reported to produce replication competent XMRV [4] but it has not been reported that LAPC4, VCaP and EKVX produce replication competent virus."
And in the first paragraph of their Discussion, the authors state,
"Although the infection of human cell lines with murine gammaretroviruses has been previously described in the literature, what makes our current study particularly important is that prior to this study it was not known that multiple commonly used prostate cancer cell lines are contaminated with MLVs."
These statements are not correct. My collaborators and I have shown that the VCaP prostate cancer cell line produces an infectious retrovirus by two reliable methods, an S+L− assay and a marker rescue assay (Knouf et al., J Virol 83: 7353, 2009; ref. [4] in the current report). In the marker rescue assay, a cell line carrying a retroviral vector was exposed to filtered medium from VCaP cells, was passaged for 2 weeks to allow retrovirus spread, and was then analyzed for vector production. In this assay, the retroviral vector was produced at a titer of 5 × 10^6 infectious particles per ml of medium, a quite respectable titer that could only have resulted from the presence of an infectious retrovirus in the medium from the VCaP cells. We also showed that the VCaP virus was relatively poorly able to infect and replicate in human cells, unlike XMRV, indicating that the VCaP virus is not XMRV. Our conclusion that the VCaP virus is not XMRV is consistent with the virus sequencing results presented in the current report.
Elsewhere in their Results, the authors state,
"LAPC4 and VCaP are not known to contain retroviruses. Interestingly, in a previous study, cell culture media from VCaP was anecdotally reported to display viral activity [4]."
I ask the authors, in what sense are our results anecdotal compared to the present results, especially when the iGLuc-DERSE assay the authors used to detect virus activity is new, unpublished and relatively uncharacterized?
I do agree that the authors' identification of infectious viruses in the LAPC4 and EKVX cancer cell lines, and the determination of the sequences of the LAPC4, VCaP and EKVX viruses, are very useful contributions, and help sort out the origin of retrovirus infections found in human prostate cancer cell lines.
RE: Infectious virus in VCaP cells
ksfanos replied to DustyMiller on 06 Jul 2011 at 02:44 GMT
We appreciate the comments by Dr. Miller regarding a number of statements in our manuscript. We recognize that the word anecdotal may not have been the most appropriate choice of words when describing the findings reported by Knouf et al. (Knouf et al., J Virol 83: 7353-56, 2009) in reference to their identification of an infectious retrovirus in the supernatants of cultured VCaP cells. We did not intend to indicate that these results were based on “anecdotal evidence”. Rather, we intended to convey that, although Knouf et al. performed a series of highly specific infectivity assays which showed for the first time (to our knowledge) that supernatant from cultured VCaP cells contained a replication competent gammaretrovirus, this was not a main focus of their study. In our manuscript we have confirmed this finding and have comprehensively extended it by performing a series of experiments to help to define the exact nature of this virus. These experiments included the following: i) sequencing the viral genome; ii) performing phylogenetic analyses; iii) confirming that the distributed VCaP cell line obtained directly from the ATCC is positive for virus; and iv) mapping multiple integration sites for the virus, which we determined to be Bxv-1 (XMV43).