To systematically define molecular features in human tumour cells which determine their degree of sensitivity to human allogeneic natural killer (NK) cells, we quantified the NK cell responsiveness of hundreds of molecularly-annotated "DNA-barcoded" solid tumour cell lines in multiplexed format and applied genome-scale CRISPR-based gene editing screens in several solid tumour cell lines to functionally interrogate which genes in tumour cells regulate the response to NK cells. In these orthogonal studies, NK-sensitive tumour cells tend to exhibit "mesenchymal-like" transcriptional programs; high transcriptional signature for chromatin remodeling complexes; high levels of B7-H6 (NCR3LG1); low levels of HLA-E/antigen presentation genes. Importantly, transcriptional signatures of NK cell-sensitive tumour cells correlate with immune checkpoint inhibitor (ICI) resistance in clinical samples. This study provides a comprehensive map of mechanisms regulating tumour cell responses to NK cells, with implications for future biomarker-driven applications of NK cell immunotherapies.

The extent and importance of functional heterogeneity and crosstalk between tumor cells is poorly understood. Here, we describe the generation of clonal populations from a patient-derived ovarian clear cell carcinoma model which forms malignant ascites and solid peritoneal tumors upon intraperitoneal transplantation in mice. The clonal populations are engineered with secreted Gaussia luciferase to monitor tumor growth dynamics and tagged with a unique DNA barcode to track their fate in multiclonal mixtures during tumor progression. Only one clone, CL31, grows robustly, generating exclusively malignant ascites. However, multiclonal mixtures form large solid peritoneal metastases, populated almost entirely by CL31, suggesting that transient cooperative interclonal interactions are sufficient to promote metastasis of CL31. CL31 uniquely harbors ERBB2 amplification, and its acquired metastatic activity in clonal mixtures is dependent on transient exposure to amphiregulin, which is exclusively secreted by non-tumorigenic clones. Amphiregulin enhances CL31 mesothelial clearance, a prerequisite for metastasis. These findings demonstrate that transient, ostensibly innocuous tumor subpopulations can promote metastases via “hit-and-run” commensal interactions.

KB-0742 is an orally available, potent, and selective inhibitor of cyclin-dependent kinase 9 (CDK9). Sensitivity profiling across a panel of 800 adherent and suspension immortalized pan-cancer cell lines using the Broad Institute PRISM platform had previously demonstrated MYC amplification as being a driver of KB-0742 sensitivity. We sought to further determine the activity of KB-0742 in breast cancer cell lines, patient-derived cell and organoid cultures, and patient-derived xenograft (PDX) models. KB-0742 decreased the viability of cell lines derived from primary and metastatic breast cancers, including triple-negative breast cancers (TNBCs). In a panel of 17 breast cancer cell lines, including 2 human epidermal growth factor receptor 2-positive (HER2+), 7 estrogen-receptor positive (ER+), 1 HER2+/ER+ and 7 TNBCs, the effect of KB-0742 on viability was either cytostatic or cytotoxic over the duration of the culture period. KB-0742 was also tested in patient-derived cell lines derived from primary and metastatic tumors including TNBC, and in 2 TNBC MYC-high expressing patient-derived organoid cultures. KB-0742 demonstrated cytostatic and cytotoxic effects on cell growth and superior inhibitory effects on cell growth in the TNBC organoid cultures as compared with paclitaxel and gemcitabine. Since sensitivity to KB-0742 was observed in breast cancer cell lines and patient-derived models, including TNBCs, we evaluated efficacy and target engagement in several MYC-high expressing TNBC PDX animal models. Treatment with KB-0742 using an intermittent dosing schedule (3-days on/4-days off) was well tolerated and resulted in anti-tumor activity comparable to that observed with standard of care chemotherapeutic agents. Tumors collected at 2 and 8 hours post-final dose displayed a decrease in phosphorylated serine residue 2 (pSER2) in the C-terminal domain of RNA polymerase II (RNA pol-II) and decreased MYC protein levels consistent with CDK9 inhibition. These data demonstrate the efficacy of KB-0742 in preclinical models of breast cancer and supports clinical testing in TNBC patients. KB-0742 is currently being evaluated in a phase I dose-escalation trial in patients with relapsed or refractory solid tumors or non-Hodgkin lymphoma (NCT04718675). Citation Format: Douglas C. Saffran, Melinda A.L. Day, Nathalie Rioux, Tom Chen, Christina Lee, Suha Naffar-Abu Amara, David B. Freeman, Tressa Hood, Charles Y. Lin, Pavan Kumar, Jorge DiMartino. Preclinical activity of KB-0742, an oral, highly selective, CDK9 inhibitor, in cell lines and in MYC-high expressing, patient-derived models of multiple breast cancer subtypes [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-08-05.