Abstract P5-04-19: JAK2 copy number and targeted JAK2 inhibition of TNBC cell lines

Abstract

Abstract Background A subset of triple negative breast cancers (TNBC) has somatic amplification of chromosome 9p24.1, encoding PD-L1, PD-L2, and JAK2 (the "PDJ" amplicon). The JAK/STAT pathway promotes malignant transformation and proliferation, and the JAK1/JAK2 inhibitor ruxolitinib is being evaluated in early phase clinical trials in breast cancer. This study was designed to measure the frequency of PDJ copy number gain in existing TNBC cell lines, and determine the impact of targeted JAK2 inhibition on cellular proliferation. Methods: Copy number alterations were measured in nine TNBC cell lines (MDA-MB-453, MDA-MB-436, BT549, MDA-MB231, MFM-223, MDA-MB-468, H5578T, MDA-MB-157, HCC1937) by array comparative genomic hybridization (aCGH). JAK2 and pSTAT3 expression was measured by immunoblot in four of the TNBC lines (HCC1937, MDA-MB-468, MDA-MB-231, MDA-MB-436) and compared with MCF10A, SK-BR3, and T47D. To selectively inhibit JAK2, lentiviral vectors encoding five different shRNA targeting JAK2 were generated. Ruxolitinib treatment was performed at 25 uM for 48 hrs and 72 hrs. Cell proliferation was measured by CellTiter-Glo. Results: All TNBC cell lines had an elevated level of baseline protein expression of JAK2 and pSTAT3 compared to SK-BR3, T47D, and MCF10A. Of the nine TNBC cell lines, only MDA-MB-436 and BT549 had copy number gain of 9p24.1 (log2ratio<1). Four independent shRNA efficiently silenced over 90% of JAK2 expression in MDA-MB-231. The inhibition of JAK2 in MDA-MB-231 was associated with a decrease in both JAK2 and pSTAT3 protein expression and reduction in cell proliferation compared to control shRNA after 96 hrs (p<0.0001). Inhibition of proliferation was also observed for both MDA-MB-231 and MDA-MB-436 after ruxolitinib treatment. Conclusion These results demonstrate that existing TNBC cell lines have varying copy number alterations of chromosome 9p24.1 encoding JAK2, and targeted JAK2 inhibition in TNBC inhibits cell proliferation. Citation Format: Chen M, Andreozzi M, Gonzalez-Malerva L, Eaton S, Pockaj B, Barrett MT, Anderson KS. JAK2 copy number and targeted JAK2 inhibition of TNBC cell lines. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P5-04-19.