Combination therapies to improve the efficacy of immunotherapy in triple-negative breast cancer [RNA-seq]

Immune checkpoint inhibitors combined with chemotherapy represent a promising treatment option in triple-negative breast cancer (TNBC). However, response rates are still relatively low necessitating the design of novel therapeutic strategies to improve clinical outcomes. Here, we describe a triple combination of anti-PDL-1 immune checkpoint blockade, epigenetic modulation thorough BET bromodomain inhibition, and chemotherapy with paclitaxel that effectively inhibits both primary and metastatic tumor growth in two different syngeneic murine breast cancer models. Detailed cellular and molecular profiling of tumors from single and combination treatment arms revealed increased T and B cell infiltration and macrophage reprogramming from M1 to a M2 phenotype in mice treated with triple combination. Overall design: 5- to 6-week-old BALBc mice were injected with 200,000 EMT-6 mammary tumors cells (ATTC, CRL-2755). Both inguinal mammary fat pads were injected in each mouse. Tumor development was monitored by palpation and once tumors were palpable, usually about 5-7 days after cancer cell inoculation, treatment was initiate. JQ1 was administered as single agents or in combination at the following doses: JQ1 at 50mg/kg by daily gavage,, anti-PD-L1 antibody (clone 10F.9G2, BioXCell) and paclitaxel each at 10mg/kg by i.p. injection twice per week. Tumor volumes were measured by caliper every three days until the end of the experiment. Once control tumors reached the study endpoint, all mice were euthanized, and primary tumors and various organs were collected. Tumors were then dissociated into single cells and separated into CD45+ and CD45- cell fractions using anti-CD45 antibody coupled to Dynabeads to enrich for leukocytes and tumor cells, respectively. Cells were counted and about 50,000 cells for each fraction were frozen for bulk ATAC-seq. The remaining cells were used for RNA extraction and bulk RNA-seq.