Abstract 1595: Human mammary tumor virus (HMTV) detection in surgical specimens and metastases of human breast cancer by the Nanostring nCounter system and FISH analysis

Abstract

Abstract The purpose of this investigation was to confirm previous PCR studies that showed the presence of Human Mammary Tumor Virus (HMTV) in human breast cancer. HMTV a retrovirus 90-98% homologous to Mouse Mammary tumor Virus (MMTV), the etiological agent of murine mammary cancer, has been detected by PCR in 40% of American women's breast cancers, but not in healthy tissues of the same breast (Cancer Research 1995, 55:5173-79). A complete 9.9kb HMTV proviral sequence has been detected from breast cancer genomic DNA and found to be inserted at different chromosomes (Cancer Res 2001,61:1754-9). HMTV viral particles with betaretrovirus characteristics and RNA sequence 85-95% homologous to MMTV have been isolated from primary cultures of metastatic breast cancer established from pleural and ascitic effusions of patients (MSSM cells). Expression of HMTV proteins has been detected in MSSM cells but not in normal mammary epithelial cells by western blot, fluorescence-activated cell sorting analysis, and immunofluorescence assays (J Virol Methods 2010, 163(1): 157-61). PCR is a sensitive molecular technique susceptible of contamination, which due to amplification, can result in false positive detection. Although our samples were tested for cox2 sequences indicative of murine mitochondrial DNA and Intracisternal A type Particle (IAP) sequences characteristic of mouse genomic DNA, the possibility still exists that laboratory contamination accounted for mistaken HMTV detection. This possibility of contamination has led to doubt that murine-like viruses exist as human pathogens. We now report two new methods for HMTV detection to reinforce previous findings. Using fluorescence in situ hybridization (FISH) assays with a HMTV probe, HMTV proviral DNA has been visualized in the nuclei of surgical samples of breast cancer as well as MSSM cells. The Nanostring nCounter system, a new technology using specific 100mer HMTV oligonucleotides as probes bound to reporter molecules, detected HMTV gag, env and LTR sequences in the absence of cox2 and IAP in a single reaction without amplification. Results from FISH analysis and Nanostring nCounter reactions show the presence of HMTV DNA in surgical cancer specimens and in MSSM metastatic cells. The data exclude contamination and confirm the authenticity of HMTV in human breast cancer. Studies are being pursued to determine if HMTV, like MMTV, plays a role in causation. Citation Format: Stella M. Melana, Polly Etkind, Teiko Nartey, Yetunde Agbaje, James F. Holland, Beatriz G-T Pogo. Human mammary tumor virus (HMTV) detection in surgical specimens and metastases of human breast cancer by the Nanostring nCounter system and FISH analysis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1595. doi:10.1158/1538-7445.AM2014-1595