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Identification of replication competent murine gammaretroviruses in commonly used prostate cancer cell lines.

Sfanos KS, Aloia AL, Hicks JL, Esopi DM, Steranka JP, Shao W, Sanchez-Martinez S, Yegnasubramanian S, Burns KH, Rein A, De Marzo AM - PLoS ONE (2011)

Bottom Line: We also identified a gammaretrovirus in the non-small-cell lung carcinoma cell line EKVX.Prostate cancer cell lines appear to have a propensity for infection with murine gammaretroviruses, and we propose that this may be in part due to cell line establishment by xenograft passage in immunocompromised mice.Importantly, our results suggest a need for regular screening of cancer cell lines for retroviral "contamination", much like routine mycoplasma testing.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America. ksfanos@jhmi.edu

ABSTRACT
A newly discovered gammaretrovirus, termed XMRV, was recently reported to be present in the prostate cancer cell line CWR22Rv1. Using a combination of both immunohistochemistry with broadly-reactive murine leukemia virus (MLV) anti-sera and PCR, we determined if additional prostate cancer or other cell lines contain XMRV or MLV-related viruses. Our study included a total of 72 cell lines, which included 58 of the 60 human cancer cell lines used in anticancer drug screens and maintained at the NCI-Frederick (NCI-60). We have identified gammaretroviruses in two additional prostate cancer cell lines: LAPC4 and VCaP, and show that these viruses are replication competent. Viral genome sequencing identified the virus in LAPC4 and VCaP as nearly identical to another known xenotropic MLV, Bxv-1. We also identified a gammaretrovirus in the non-small-cell lung carcinoma cell line EKVX. Prostate cancer cell lines appear to have a propensity for infection with murine gammaretroviruses, and we propose that this may be in part due to cell line establishment by xenograft passage in immunocompromised mice. It is unclear if infection with these viruses is necessary for cell line establishment, or what confounding role they may play in experiments performed with these commonly used lines. Importantly, our results suggest a need for regular screening of cancer cell lines for retroviral "contamination", much like routine mycoplasma testing.

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CWR22Rv1, LAPC4, VCaP and EKVX produce replication competent virus as indicated by Gaussia luciferase production in iGLuc-DERSE cells.10 µl of media was assayed 2 days after exposure to 24 hr. cell culture supernatant from CWR22Rv1, LAPC4 and VCaP, and 13 days (and 3 passages) after exposure to supernatant from EKVX, DU145, MycCaP (murine prostate cancer cell line) and PC3. Red line indicates background luciferase expression as determined by a mock-infected control. Error bars represent standard error of the mean for 3 experiments.
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pone-0020874-g003: CWR22Rv1, LAPC4, VCaP and EKVX produce replication competent virus as indicated by Gaussia luciferase production in iGLuc-DERSE cells.10 µl of media was assayed 2 days after exposure to 24 hr. cell culture supernatant from CWR22Rv1, LAPC4 and VCaP, and 13 days (and 3 passages) after exposure to supernatant from EKVX, DU145, MycCaP (murine prostate cancer cell line) and PC3. Red line indicates background luciferase expression as determined by a mock-infected control. Error bars represent standard error of the mean for 3 experiments.

Mentions: To verify that the commonly used prostate cancer cell lines LAPC4 and VCaP contain integrated and replication-competent virus, we performed a viral infectivity assay. The indicator cell line, iGLuc-DERSE cells, will only secrete the Gaussia luciferase enzyme following infection with a replication-competent MLV. Gaussia luciferase activity was detected 48 hours after infection of iGLuc-DERSE cells with supernatant from the CWR22Rv1, LAPC4 and VCaP cell lines (Figure 3). Interestingly, a triplicate infection of iGLuc-DERSE cells with supernatant from the EKVX cell line did not show Gaussia luciferase activity until 13 days (and 3 passages) after infection (Figure 3). Further, only 2 of the 3 infections showed luciferase activity above background. Thus, the virus from EKVX cells apparently has very low infectivity, at least for the iGLuc-DERSE cell line. No Gaussia luciferase activity was detected from cells (passaged for a total of 13 days) exposed to supernatant from the DU145, MycCaP (a mouse prostate cancer cell line obtained from the C. L. Sawyers lab) or PC3 cell lines (Figure 3). CWR22Rv1 was previously reported to produce replication competent XMRV [4] but it has not been reported that LAPC4, VCaP and EKVX produce replication competent virus.


Identification of replication competent murine gammaretroviruses in commonly used prostate cancer cell lines.

Sfanos KS, Aloia AL, Hicks JL, Esopi DM, Steranka JP, Shao W, Sanchez-Martinez S, Yegnasubramanian S, Burns KH, Rein A, De Marzo AM - PLoS ONE (2011)

CWR22Rv1, LAPC4, VCaP and EKVX produce replication competent virus as indicated by Gaussia luciferase production in iGLuc-DERSE cells.10 µl of media was assayed 2 days after exposure to 24 hr. cell culture supernatant from CWR22Rv1, LAPC4 and VCaP, and 13 days (and 3 passages) after exposure to supernatant from EKVX, DU145, MycCaP (murine prostate cancer cell line) and PC3. Red line indicates background luciferase expression as determined by a mock-infected control. Error bars represent standard error of the mean for 3 experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3117837&req=5

pone-0020874-g003: CWR22Rv1, LAPC4, VCaP and EKVX produce replication competent virus as indicated by Gaussia luciferase production in iGLuc-DERSE cells.10 µl of media was assayed 2 days after exposure to 24 hr. cell culture supernatant from CWR22Rv1, LAPC4 and VCaP, and 13 days (and 3 passages) after exposure to supernatant from EKVX, DU145, MycCaP (murine prostate cancer cell line) and PC3. Red line indicates background luciferase expression as determined by a mock-infected control. Error bars represent standard error of the mean for 3 experiments.
Mentions: To verify that the commonly used prostate cancer cell lines LAPC4 and VCaP contain integrated and replication-competent virus, we performed a viral infectivity assay. The indicator cell line, iGLuc-DERSE cells, will only secrete the Gaussia luciferase enzyme following infection with a replication-competent MLV. Gaussia luciferase activity was detected 48 hours after infection of iGLuc-DERSE cells with supernatant from the CWR22Rv1, LAPC4 and VCaP cell lines (Figure 3). Interestingly, a triplicate infection of iGLuc-DERSE cells with supernatant from the EKVX cell line did not show Gaussia luciferase activity until 13 days (and 3 passages) after infection (Figure 3). Further, only 2 of the 3 infections showed luciferase activity above background. Thus, the virus from EKVX cells apparently has very low infectivity, at least for the iGLuc-DERSE cell line. No Gaussia luciferase activity was detected from cells (passaged for a total of 13 days) exposed to supernatant from the DU145, MycCaP (a mouse prostate cancer cell line obtained from the C. L. Sawyers lab) or PC3 cell lines (Figure 3). CWR22Rv1 was previously reported to produce replication competent XMRV [4] but it has not been reported that LAPC4, VCaP and EKVX produce replication competent virus.

Bottom Line: We also identified a gammaretrovirus in the non-small-cell lung carcinoma cell line EKVX.Prostate cancer cell lines appear to have a propensity for infection with murine gammaretroviruses, and we propose that this may be in part due to cell line establishment by xenograft passage in immunocompromised mice.Importantly, our results suggest a need for regular screening of cancer cell lines for retroviral "contamination", much like routine mycoplasma testing.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America. ksfanos@jhmi.edu

ABSTRACT
A newly discovered gammaretrovirus, termed XMRV, was recently reported to be present in the prostate cancer cell line CWR22Rv1. Using a combination of both immunohistochemistry with broadly-reactive murine leukemia virus (MLV) anti-sera and PCR, we determined if additional prostate cancer or other cell lines contain XMRV or MLV-related viruses. Our study included a total of 72 cell lines, which included 58 of the 60 human cancer cell lines used in anticancer drug screens and maintained at the NCI-Frederick (NCI-60). We have identified gammaretroviruses in two additional prostate cancer cell lines: LAPC4 and VCaP, and show that these viruses are replication competent. Viral genome sequencing identified the virus in LAPC4 and VCaP as nearly identical to another known xenotropic MLV, Bxv-1. We also identified a gammaretrovirus in the non-small-cell lung carcinoma cell line EKVX. Prostate cancer cell lines appear to have a propensity for infection with murine gammaretroviruses, and we propose that this may be in part due to cell line establishment by xenograft passage in immunocompromised mice. It is unclear if infection with these viruses is necessary for cell line establishment, or what confounding role they may play in experiments performed with these commonly used lines. Importantly, our results suggest a need for regular screening of cancer cell lines for retroviral "contamination", much like routine mycoplasma testing.

Show MeSH
Related in: MedlinePlus