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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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LAPC4, VCaP and EKVX cancer cell lines are infected with a murine leukemia virus which is not XMRV.(A) Examples of MLV-negative (DU145) and positive (CWR22Rv1, LAPC4, VCaP and EKVX) cancer cell lines stained with MLV30 and MLV70 antisera. (B) Positive staining in LAPC4, VCaP and EKVX does not represent XMRV. Lanes 1–7, PCR with MLV-specific primers (1 =  CWR22Rv1, 2 =  LAPC4, 3 =  VCaP, 4 =  DU145, 5 =  LNCaP, 6 =  PC3, 7 =  EKVX), Lanes 8–11, PCR with XMRV-specific primers (8 =  CWR22Rv1, 9 =  LAPC4, 10 =  VCaP, 11 =  EKVX). Difference in product size between CWR22Rv1 (lane 1) and LAPC4, VCaP or EKVX (lanes 2,3,7) is due to the XMRV-specific 24-nt deletion [13]. L =  molecular weight ladder.
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pone-0020874-g001: LAPC4, VCaP and EKVX cancer cell lines are infected with a murine leukemia virus which is not XMRV.(A) Examples of MLV-negative (DU145) and positive (CWR22Rv1, LAPC4, VCaP and EKVX) cancer cell lines stained with MLV30 and MLV70 antisera. (B) Positive staining in LAPC4, VCaP and EKVX does not represent XMRV. Lanes 1–7, PCR with MLV-specific primers (1 =  CWR22Rv1, 2 =  LAPC4, 3 =  VCaP, 4 =  DU145, 5 =  LNCaP, 6 =  PC3, 7 =  EKVX), Lanes 8–11, PCR with XMRV-specific primers (8 =  CWR22Rv1, 9 =  LAPC4, 10 =  VCaP, 11 =  EKVX). Difference in product size between CWR22Rv1 (lane 1) and LAPC4, VCaP or EKVX (lanes 2,3,7) is due to the XMRV-specific 24-nt deletion [13]. L =  molecular weight ladder.

Mentions: As an initial screen for murine gammaretroviruses in human cell lines, we utilized a tissue microarray (TMA) containing 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) (Table S1). As shown in Figure 1A, IHC with broadly reactive antisera for Moloney murine leukemia virus (Mo-MLV) p30CA (MLV30) or gp70SU (MLV70) [3] identified three prostate cancer cell lines (CWR22Rv1, LAPC4 [17], VCaP [18]) as positive for virus. While CWR22Rv1 was previously reported to be infected with replication competent XMRV [4], 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]. We also identified a non-small-cell lung carcinoma cell line, EKVX, as being positive for 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)

LAPC4, VCaP and EKVX cancer cell lines are infected with a murine leukemia virus which is not XMRV.(A) Examples of MLV-negative (DU145) and positive (CWR22Rv1, LAPC4, VCaP and EKVX) cancer cell lines stained with MLV30 and MLV70 antisera. (B) Positive staining in LAPC4, VCaP and EKVX does not represent XMRV. Lanes 1–7, PCR with MLV-specific primers (1 =  CWR22Rv1, 2 =  LAPC4, 3 =  VCaP, 4 =  DU145, 5 =  LNCaP, 6 =  PC3, 7 =  EKVX), Lanes 8–11, PCR with XMRV-specific primers (8 =  CWR22Rv1, 9 =  LAPC4, 10 =  VCaP, 11 =  EKVX). Difference in product size between CWR22Rv1 (lane 1) and LAPC4, VCaP or EKVX (lanes 2,3,7) is due to the XMRV-specific 24-nt deletion [13]. L =  molecular weight ladder.
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pone-0020874-g001: LAPC4, VCaP and EKVX cancer cell lines are infected with a murine leukemia virus which is not XMRV.(A) Examples of MLV-negative (DU145) and positive (CWR22Rv1, LAPC4, VCaP and EKVX) cancer cell lines stained with MLV30 and MLV70 antisera. (B) Positive staining in LAPC4, VCaP and EKVX does not represent XMRV. Lanes 1–7, PCR with MLV-specific primers (1 =  CWR22Rv1, 2 =  LAPC4, 3 =  VCaP, 4 =  DU145, 5 =  LNCaP, 6 =  PC3, 7 =  EKVX), Lanes 8–11, PCR with XMRV-specific primers (8 =  CWR22Rv1, 9 =  LAPC4, 10 =  VCaP, 11 =  EKVX). Difference in product size between CWR22Rv1 (lane 1) and LAPC4, VCaP or EKVX (lanes 2,3,7) is due to the XMRV-specific 24-nt deletion [13]. L =  molecular weight ladder.
Mentions: As an initial screen for murine gammaretroviruses in human cell lines, we utilized a tissue microarray (TMA) containing 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) (Table S1). As shown in Figure 1A, IHC with broadly reactive antisera for Moloney murine leukemia virus (Mo-MLV) p30CA (MLV30) or gp70SU (MLV70) [3] identified three prostate cancer cell lines (CWR22Rv1, LAPC4 [17], VCaP [18]) as positive for virus. While CWR22Rv1 was previously reported to be infected with replication competent XMRV [4], 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]. We also identified a non-small-cell lung carcinoma cell line, EKVX, as being positive for 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