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Although poly(ADP-ribose) polymerase (PARP) inhibitors are active in homologous recombination (HR)-deficient cancers, their utility is limited by acquired resistance after restoration of HR. Here, we report that dinaciclib, an inhibitor of cyclin-dependent kinases (CDKs) 1, 2, 5, and 9, additionally has potent activity against CDK12, a transcriptional regulator of HR. In BRCA-mutated triple-negative breast cancer (TNBC) cells and patient-derived xenografts (PDXs), dinaciclib ablates restored HR and reverses PARP inhibitor resistance. Additionally, we show that de novo resistance to PARP inhibition in BRCA1-mutated cell lines and a PDX derived from a PARP-inhibitor-naive BRCA1carrier is mediated by residual HR and is reversed by CDK12 inhibition. Finally, dinaciclib augments the degree of response in a PARP-inhibitor-sensitive model, converting tumor growth inhibition to durable regression. These results highlight the significance of HR disruption as a therapeutic strategy and support the broad use of combined CDK12 and PARP inhibition in TNBC.

Signal transduction mediated by ErbB/HER receptor tyrosine kinases is crucial for the development and maintenance of epithelial tissues, and aberrant signaling is frequently associated with malignancies of epithelial origin. This review focuses on the roles played by the Hsp90 chaperone machinery in the regulation of signaling through the ErbB/HER network, and discusses potential therapeutic strategies that disrupt chaperone functions. Hsp90 and its associated co-chaperones regulate ErbB signal transduction through multiple mechanisms. The chaperone system controls the stability of the nascent forms of both ErbB-1 (EGF-receptor) and ErbB-2/HER2, while regulation of the mature form is restricted to ErbB-2. Regulation by the Hsp90 complex extends to downstream effectors of ErbB signaling, namely Raf-1, Pdk-1 and Akt/PKB. Disrupting the function of Hsp90 results in the degradation of both the receptors and their effectors, thereby inhibiting tumor cell growth. The importance of an Hsp90-recognition motif located within the kinase domain of ErbB-2 is discussed, as well as a direct role for Hsp90 in regulating tyrosine kinase activity. In light of recent observations, we emphasize the ability of specific tyrosine kinase inhibitors to selectively target ErbB-2 to the chaperone-mediated degradation pathway. ErbB-specific drugs are already used to treat cancers, and clinical trials are underway for additional compounds that intercept ErbB signaling, including drugs that target Hsp90. Hence, the dependence of ErbB-2 upon Hsp90 reveals an Achilles heel, which opens a window of opportunity for combating cancers driven by the ErbB/HER signaling network.

Cells deficient in ataxia telangiectasia mutated (ATM) are hypersensitive to ionizing radiation and other anti-cancer agents that induce double-strand DNA breaks. ATM inhibitors may therefore sensitize cancer cells to these agents. Some cancers may also have underlying genetic defects predisposing them to an ATM inhibitor monotherapy response. We have conducted a genome-wide CRISPR screen to identify genetic vulnerabilities that sensitize lung cancer cells to ATM inhibitors. Knockout of genes in the Fanconi anemia (FA)/BRCA pathway results in hypersensitivity to the ATM inhibitor M3541. Knockdown of either an FA gene or of ATM results in reduced double-strand break end resection, enhanced non-homologous end joining (NHEJ) repair, and decreased homologous recombination repair. Knockout of both the FA/BRCA pathway and ATM strongly inhibits end resection and generates toxic levels of NHEJ, thereby elucidating a mechanism of cellular death by synthetic lethality. ATM inhibitors may therefore be useful for the treatment of tumors with a defective FA/BRCA pathway.

The molecular mechanisms responsible for opposing oncogenic and tumor-suppressor activities of NF-kB are obscure. Semi-quantitative immunohistochemistry of primary breast tumors using antibodies to RelA, the pleiotropic NF-kB factor, and Ki67 revealed a negative correlation between RelA levels and Ki67-index among ER+/HER2− tumors [1]. Similarly, expression of AURKA, a marker for proliferation, negatively correlates with expression of NFKBIA, a surrogate for RelA expression and activity, in ER+/HER2− tumors analyzed by The Cancer Genome Atlas [2–4]. Furthermore, conditional expression of RelA using a Tetracycline-inducible system in Human Mammary Epithelial Cells (HRA cells) caused proliferation arrest while withdrawal of Doxycycline (Dox) and suppression of RelA expression in arrested cells restored cell cycle progression [1]. To identify genes responsible for the negative relationship between RelA levels and proliferation, we performed genome-wide gene expression analysis of HRA cells under the following conditions: RelA un-induced, No Dox (ND); Dox induced for 24h; Dox induced for 72h; Dox induced for 24h then Dox withdrawn for 48h. The expression data was submitted to Gene Expression Ominibus (GEO) and the accession number is GSE65040. Analysis of the data identified cross-talk between basal RelA activity and the Interferon pathway mediated by IRF1, a target of RelA [5]. Activation of the Interferon pathway lead to down-regulation of CDK4 expression resulting in RB1 hypo-phosphorylation and suppression of cell cycle progression. The tumor-suppressor activity of NF-kB, specifically RelA, may stem from cross-talk with the Interferon pathway.

BRCA2 (also known as FANCD1), a core component of the Fanconi pathway, suppresses transformation of immature T-cell progenitors in mice. However, whether the Fanconi pathway contributes to human T-cell acute lymphoblastic leukemia (T-ALL) pathogenesis is unknown. Using targeted exon sequencing of childhood T-ALL diagnostic specimens, we found heterozygous mutations of Fanconi pathway genes in 23% of 40 cases analyzed. Array comparative genomic hybridization (CGH) analysis revealed that an additional 15% of these cases harbored large heterozygous deletions involving Fanconi pathway genes. The majority of these mutations appear to be somatic although in 2 of 8 available remission samples, the mutations were present, suggestive that these mutations are likely to be germline. Fanconi genes are typically considered two-hit tumor suppressors, where breast or ovarian cancers arising in heterozygous carriers often lose activity of the wild-type allele, leading to genomic instability. However, T-ALL cases with heterozygous Fanconi mutations lacked evidence of increased genomic instability based on array CGH analysis, were not hypersensitive to the PARP inhibitor olaparib, and did not demonstrate increased radial chromosome formation in response to mitomycin C (MMC), indicating that these cases retained activity of the wild-type allele. Complementation studies of Fanconi mutant cells revealed that most of the identified Fanconi mutations encoded pathogenic alleles, as evidenced by their impaired ability to rescue Fanconi-deficient cells from mitomycin C-induced radial chromosome formation and cytotoxicity. Treatment of a patient-derived xenograft sample harboring a heterozygous BRCA2 mutation with mitomycin C revealed that these cells had an intermediate increase in sensitivity to mitomycin C-induced cytotoxicity compared to Fanconi wild-type T-ALLs, suggesting that Fanconi haploinsufficiency is pathogenic in T-ALL. Together, these findings implicate a tumor suppressor role for Fanconi pathway haploinsufficiency in childhood T-ALL.