Purpose: In cancer cells, the epigenome is often deregulated, and inhibition of the bromodomain and extra-terminal (BET) family of bromodomain-containing proteins is a novel epigenetic therapeutic approach. Preliminary results of an ongoing phase I trial have reported promising activity and tolerability with the new BET bromodomain inhibitor OTX015.Experimental Design: We assessed the preclinical activity of OTX015 as single agent and in combination in mature B-cell lymphoma models and performed in vitro and in vivo experiments to identify the mechanism of action and the genetic features associated with sensitivity to the compound.Results: OTX015 showed antiproliferative activity in a large panel of cell lines derived from mature B-cell lymphoid tumors with median IC 50 of 240 nmol/L, without significant differences among the different histotypes. In vitro and in vivo experiments showed that OTX015 targeted NFKB/TLR/JAK/STAT signaling pathways, MYC-and E2F1-regulated genes, cell-cycle regulation, and chromatin structure. OTX015 presented in vitro synergism with several anticancer agents, especially with mTOR and BTK inhibitors. Gene expression signatures associated with different degrees of sensitivity to OTX015 were identified. Although OTX015 was mostly cytostatic, the compound induced apoptosis in a genetically defined subgroup of cells, derived from activated B-cell-like diffuse large B-cell lymphoma, bearing wtTP53, mutations in MYD88, and CD79B or CARD11.Conclusions: Together with the data coming from the ongoing phase I study, the in vitro and in vivo data presented here provide the basis for further clinical investigation of OTX015 as single agent and in combination therapies.

The involvement of the MET oncogene in de novo and acquired resistance of non-small cell lung cancers (NSCLC) to tyrosine kinase inhibitors (TKIs) has been reported, but the precise mechanism by which MET overexpression contributes to TKI-resistant NSCLC remains unclear. MicroRNAs (miRNAs) negatively regulate gene expression and their dysregulation has been implicated in tumorigenesis. To understand the role of microRNAs in TKI-resistant NSCLC, we examined TK receptor-mediated microRNA changes. Here we report that miR-30b/c and miR-221/222, modulated by both EGF and MET receptors, and miR-103, -203, controlled only by MET, play important roles in gefitinib-induced apoptosis and epithelial-mesenchymal transition (EMT) of NSCLC cells, in vitro and in vivo, by inhibiting the expression of Bim, APAF-1, PKC-ε and SRC genes. The finding suggests that modulation of specific microRNAs may provide a therapeutic approach for future treatment of NSCLC.

SummaryMicroRNA deregulation is frequent in human colorectal cancers (CRCs), but little is known as to whether it represents a bystander event or actually drives tumor progression in vivo. We show that miR-135b overexpression is triggered in mice and humans by APC loss, PTEN/PI3K pathway deregulation, and SRC overexpression and promotes tumor transformation and progression. We show that miR-135b upregulation is common in sporadic and inflammatory bowel disease-associated human CRCs and correlates with tumor stage and poor clinical outcome. Inhibition of miR-135b in CRC mouse models reduces tumor growth by controlling genes involved in proliferation, invasion, and apoptosis. We identify miR-135b as a key downsteam effector of oncogenic pathways and a potential target for CRC treatment.

Triple negative breast cancer (TNBC) is a heterogeneous disease at the molecular, pathologic and clinical levels. To stratify TNBCs, we determined microRNA (miRNA) expression profiles, as well as expression profiles of a cancer-focused mRNA panel, in tumor, adjacent non-tumor (normal) and lymph node metastatic lesion (mets) tissues, from 173 women with TNBCs; we linked specific miRNA signatures to patient survival and used miRNA/mRNA anti-correlations to identify clinically and genetically different TNBC subclasses. We also assessed miRNA signatures as potential regulators of TNBC subclass-specific gene expression networks defined by expression of canonical signal pathways.Tissue specific miRNAs and mRNAs were identified for normal vs tumor vs mets comparisons. miRNA signatures correlated with prognosis were identified and predicted anti-correlated targets within the mRNA profile were defined. Two miRNA signatures (miR-16, 155, 125b, 374a and miR-16, 125b, 374a, 374b, 421, 655, 497) predictive of overall survival (P = 0.05) and distant-disease free survival (P = 0.009), respectively, were identified for patients 50 yrs of age or younger. By multivariate analysis the risk signatures were independent predictors for overall survival and distant-disease free survival. mRNA expression profiling, using the cancer-focused mRNA panel, resulted in clustering of TNBCs into 4 molecular subclasses with different expression signatures anti-correlated with the prognostic miRNAs.Our findings suggest that miRNAs play a key role in triple negative breast cancer through their ability to regulate fundamental pathways such as: cellular growth and proliferation, cellular movement and migration, Extra Cellular Matrix degradation. The results define miRNA expression signatures that characterize and contribute to the phenotypic diversity of TNBC and its metastasis.

Acute graft-versus-host disease (aGVHD) remains a major complication of allogeneic hematopoietic stem cell transplant (alloHSCT), underscoring the need to further elucidate its mechanisms and develop novel treatments. Based on recent observations that microRNA-155 (miR-155) is up-regulated during T-cell activation, we hypothesized that miR-155 is involved in the modulation of aGVHD.Here we show that miR-155 expression was up-regulated in T cells from mice developing aGVHD after alloHSCT. Mice receiving miR-155-deficient donor lymphocytes had markedly reduced lethal aGVHD, whereas lethal aGVHD developed rapidly in mice recipients of miR-155 overexpressing T cells. Blocking miR-155 expression using a synthetic antimiR-155 after alloHSCT decreased aGVHD severity and prolonged survival in mice. Finally, miR-155 up-regulation was shown in specimens from patients with pathologic evidence of intestinal aGVHD. Altogether, our data indicate a role for miR-155 in the regulation of GVHD and point to miR-155 as a novel target for therapeutic intervention in this disease.

It is generally assumed that recurrent mutations within a given cancer driver gene elicit similar drug responses. Cancer genome studies have identified recurrent but divergent missense mutations in the substrate recognition domain of the ubiquitin ligase adaptor SPOP in endometrial and prostate cancer. Their therapeutic implications remain incompletely understood. Here, we analyzed changes in the ubiquitin landscape induced by endometrial cancer-associated SPOP mutations and identified BRD2, BRD3 and BRD4 proteins (BETs) as SPOP-CUL3 substrates that are preferentially degraded by endometrial SPOP mutants. The resulting reduction of BET protein levels sensitized cancer cells to BET inhibitors. Conversely, prostate cancer-specific SPOP mutants impaired degradation of BETs, promoting resistance against their pharmacologic inhibition. These results uncover an oncogenomics paradox, whereby mutations within the same domain evoke opposing drug susceptibilities. Specifically, we provide a molecular rationale for the use of BET inhibitors to treat endometrial but not prostate cancer patients with SPOP mutations.

Significance Cell survival after DNA damage relies on DNA repair, the abrogation of which causes genomic instability and development of cancer. DNA double-strand breaks are lesions induced by ionizing radiation (IR) and can be efficiently repaired by DNA homologous recombination, a system that requires RAD51 recombinase (RAD51). Here we show that overexpression of miR-155 in human breast cancer cells reduces the levels of RAD51 and affects the cellular response to IR. High miR-155 levels were associated with lower RAD51 expression and with better overall survival of patients in a large series of triple-negative breast cancers. Testing triple-negative breast cancer patients for miR-155 expression may be a useful prognostic tool to identify who will benefit from an IR-based therapeutic approach.

MicroRNAs (miRNAs) have an important role in the development of chemosensitivity or chemoresistance in different types of cancer. Activation of the ERK1/2 pathway is a major determinant of diverse cellular processes and cancer development and is responsible for the transcription of several important miRNAs. Here we show a link between the ERK1/2 pathway and BIM expression through miR-494. We blocked ERK1/2 nuclear activity through the overexpression of an ERK1/2 natural interactor, the protein PED/PEA15, and we performed a microRNA expression profile. miR-494 was the most down-regulated microRNA after ERK1/2 inactivation. Moreover, we found that miR-494 induced Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) resistance in non–small-cell lung cancer (NSCLC) through the down-modulation of BIM. Elucidation of this undiscovered ERK1/2 pathway that regulates apoptosis and cell proliferation through miR-494 in NSCLC will greatly enhance our understanding of the mechanisms responsible for TRAIL resistance and will provide an additional arm for the development of anticancer therapies.