Poly(ADP-ribose) polymerase inhibitors (PARPi) are selectively active in cells with homologous recombination (HR) deficiency (HRD) caused by mutations in BRCA1, BRCA2, and other pathway members. We sought small molecules that induce HRD in HR-competent cells to induce synthetic lethality with PARPi and extend the utility of PARPi. We demonstrated that inhibition of bromodomain containing 4 (BRD4) induced HRD and sensitized cells across multiple tumor lineages to PARPi regardless of BRCA1/2, TP53, RAS, or BRAF mutation status through depletion of the DNA double-stand break resection protein CtIP (C-terminal binding protein interacting protein). Importantly, BRD4 inhibitor (BRD4i) treatment reversed multiple mechanisms of resistance to PARPi. Furthermore, PARPi and BRD4i are synergistic in multiple in vivo models.

The functional impact of the vast majority of cancer somatic mutations remains unknown, representing a critical knowledge gap for implementing precision oncology. Here, we report the development of a moderate-throughput functional genomic platform consisting of efficient mutant generation, sensitive viability assays using two growth factor-dependent cell models, and functional proteomic profiling of signaling effects for select aberrations. We apply the platform to annotate >1,000 genomic aberrations, including gene amplifications, point mutations, indels, and gene fusions, potentially doubling the number of driver mutations characterized in clinically actionable genes. Further, the platform is sufficiently sensitive to identify weak drivers. Our data are accessible through a user-friendly, public data portal. Our study will facilitate biomarker discovery, prediction algorithm improvement, and drug development.

Expression of the PTEN tumor suppressor is frequently lost in breast cancer in the absence of mutation or promoter methylation through as yet undetermined mechanisms. In this study, we demonstrate that the Rak tyrosine kinase physically interacts with PTEN and phosphorylates PTEN on Tyr336. Knockdown of Rak enhanced the binding of PTEN to its E3 ligase NEDD4-1 and promoted PTEN polyubiquitination, leading to PTEN protein degradation. Notably, ectopic expression of Rak effectively suppressed breast cancer cell proliferation, invasion, and colony formation in vitro and tumor growth in vivo. Furthermore, Rak knockdown was sufficient to transform normal mammary epithelial cells. Therefore, Rak acts as a bona fide tumor suppressor gene through the mechanism of regulating PTEN protein stability and function.

Dysregulation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway occurs frequently in human cancer. PTEN tumor suppressor or PIK3CA oncogene mutations both direct PI3K-dependent tumorigenesis largely through activation of the AKT/PKB kinase. However, here we show through phosphoprotein profiling and functional genomic studies that many PIK3CA mutant cancer cell lines and human breast tumors exhibit only minimal AKT activation and a diminished reliance on AKT for anchorage-independent growth. Instead, these cells retain robust PDK1 activation and membrane localization and exhibit dependency on the PDK1 substrate SGK3. SGK3 undergoes PI3K- and PDK1-dependent activation in PIK3CA mutant cancer cells. Thus, PI3K may promote cancer through both AKT-dependent and AKT-independent mechanisms. Knowledge of differential PI3K/PDK1 signaling could inform rational therapeutics in cancers harboring PIK3CA mutations.

Perturbation biology is a powerful approach to modeling quantitative cellular behaviors and understanding detailed disease mechanisms. However, large-scale protein response resources of cancer cell lines to perturbations are not available, resulting in a critical knowledge gap. Here we generated and compiled perturbed expression profiles of ∼210 clinically relevant proteins in >12,000 cancer cell line samples in response to ∼170 drug compounds using reverse-phase protein arrays. We show that integrating perturbed protein response signals provides mechanistic insights into drug resistance, increases the predictive power for drug sensitivity, and helps identify effective drug combinations. We build a systematic map of "protein-drug" connectivity and develop a user-friendly data portal for community use. Our study provides a rich resource to investigate the behaviors of cancer cells and the dependencies of treatment responses, thereby enabling a broad range of biomedical applications.

Small noncoding miRNAs represent underexplored targets of genomic aberrations and emerging therapeutic targets. The 3q26.2 amplicon is among the most frequent genomic aberrations in multiple cancer lineages including ovarian and breast cancers. We demonstrate that hsa-miR-569 (hereafter designated as miR569), which is overexpressed in a subset of ovarian and breast cancers, at least in part due to the 3q26.2 amplicon, alters cell survival and proliferation. Downregulation of TP53INP1 expression by miR569 is required for the effects of miR569 on survival and proliferation. Targeting miR569 sensitizes ovarian and breast cancer cells overexpressing miR569 to cisplatin by increasing cell death both in vitro and in vivo. Thus targeting miR569 could potentially benefit patients with the 3q26.2 amplicon and subsequent miR569 elevation.

Adenosine-to-inosine (A-to-I) RNA editing is a widespread post-transcriptional mechanism, but its genomic landscape and clinical relevance in cancer have not been investigated systematically. We characterized the global A-to-I RNA editing profiles of 6,236 patient samples of 17 cancer types from The Cancer Genome Atlas and revealed a striking diversity of altered RNA-editing patterns in tumors relative to normal tissues. We identified an appreciable number of clinically relevant editing events, many of which are in noncoding regions. We experimentally demonstrated the effects of several cross-tumor nonsynonymous RNA editing events on cell viability and provide the evidence that RNA editing could selectively affect drug sensitivity. These results highlight RNA editing as an exciting theme for investigating cancer mechanisms, biomarkers, and treatments.

Lysophosphatidic acid (LPA) acts through high-affinity G protein-coupled receptors to mediate a plethora of physiological and pathological activities associated with tumorigenesis. LPA receptors and autotaxin (ATX/LysoPLD), the primary enzyme producing LPA, are aberrantly expressed in multiple cancer lineages. However, the role of ATX and LPA receptors in the initiation and progression of breast cancer has not been evaluated. We demonstrate that expression of ATX or each edg family LPA receptor in mammary epithelium of transgenic mice is sufficient to induce a high frequency of late-onset, estrogen receptor (ER)-positive, invasive, and metastatic mammary cancer. Thus, ATX and LPA receptors can contribute to the initiation and progression of breast cancer.

PIK3R1 (p85α regulatory subunit of PI3K) is frequently mutated across cancer lineages. Herein, we demonstrate that the most common recurrent PIK3R1 mutation PIK3R1(R348∗) and a nearby mutation PIK3R1(L370fs), in contrast to wild-type and mutations in other regions of PIK3R1, confers an unexpected sensitivity to MEK and JNK inhibitors in vitro and in vivo. Consistent with the response to inhibitors, PIK3R1(R348∗) and PIK3R1(L370fs) unexpectedly increase JNK and ERK phosphorylation. Surprisingly, p85α R348(∗) and L370fs localize to the nucleus where the mutants provide a scaffold for multiple JNK pathway components facilitating nuclear JNK pathway activation. Our findings uncover an unexpected neomorphic role for PIK3R1(R348∗) and neighboring truncation mutations in cellular signaling, providing a rationale for therapeutic targeting of these mutant tumors.

We showed in this study that cells deficient of the BRCA1-associated BACH1 helicase, also known as BRIP1, failed to elicit homologous recombination (HR) after DNA double-stranded breaks (DSBs). BACH1-deficient cells were also sensitive to mitomycin C (MMC) and underwent MMC-induced chromosome instability. Moreover, we identified a homozygous nonsense mutation in BACH1 in a FA-J patient-derived cell line and could not detect BACH1 protein in this cell line. Expression of wild-type BACH1 in this cell line reduced the accumulation of cells at G2/M phases following exposure to DNA crosslinkers, a characteristic of Fanconi anemia (FA) cells. These results support the conclusion that BACH1 is FANCJ.

We demonstrate that concurrent administration of poly(ADP-ribose) polymerase (PARP) and WEE1 inhibitors is effective in inhibiting tumor growth but poorly tolerated. Concurrent treatment with PARP and WEE1 inhibitors induces replication stress, DNA damage, and abrogates the G2 DNA damage checkpoint in both normal and malignant cells. Following cessation of monotherapy with PARP or WEE1 inhibitors, effects of these inhibitors persist suggesting that sequential administration of PARP and WEE1 inhibitors could maintain efficacy while ameliorating toxicity. Strikingly, while sequential administration mirrored concurrent therapy in cancer cells that have high basal replication stress, low basal replication stress in normal cells protected them from DNA damage and toxicity, thus improving tolerability while preserving efficacy in ovarian cancer xenograft and patient-derived xenograft models.

Adenosine (A) to inosine (I) RNA editing introduces many nucleotide changes in cancer transcriptomes. However, due to the complexity of post-transcriptional regulation, the contribution of RNA editing to proteomic diversity in human cancers remains unclear. Here, we performed an integrated analysis of TCGA genomic data and CPTAC proteomic data. Despite limited site diversity, we demonstrate that A-to-I RNA editing contributes to proteomic diversity in breast cancer through changes in amino acid sequences. We validate the presence of editing events at both RNA and protein levels. The edited COPA protein increases proliferation, migration, and invasion of cancer cells in vitro. Our study suggests an important contribution of A-to-I RNA editing to protein diversity in cancer and highlights its translational potential.

Cancer cell lines are major model systems for mechanistic investigation and drug development. However, protein expression data linked to high-quality DNA, RNA, and drug-screening data have not been available across a large number of cancer cell lines. Using reverse-phase protein arrays, we measured expression levels of ∼230 key cancer-related proteins in >650 independent cell lines, many of which have publically available genomic, transcriptomic, and drug-screening data. Our dataset recapitulates the effects of mutated pathways on protein expression observed in patient samples, and demonstrates that proteins and particularly phosphoproteins provide information for predicting drug sensitivity that is not available from the corresponding mRNAs. We also developed a user-friendly bioinformatic resource, MCLP, to help serve the biomedical research community.

S1P has been proposed to contribute to cancer progression by regulating tumor proliferation, invasion, and angiogenesis. We developed a biospecific monoclonal antibody to S1P to investigate its role in tumorigenesis. The anti-S1P mAb substantially reduced tumor progression and in some cases eliminated measurable tumors in murine xenograft and allograft models. Tumor growth inhibition was attributed to antiangiogenic and antitumorigenic effects of the antibody. The anti-S1P mAb blocked EC migration and resulting capillary formation, inhibited blood vessel formation induced by VEGF and bFGF, and arrested tumor-associated angiogenesis. The anti-S1P mAb also neutralized S1P-induced proliferation, release of proangiogenic cytokines, and the ability of S1P to protect tumor cells from apoptosis in several tumor cell lines, validating S1P as a target for therapy.

Checkpoint inhibition immunotherapy has revolutionized cancer treatment, but many patients show resistance. Here we perform integrative transcriptomic and proteomic analyses on emerging immuno-oncology targets across multiple clinical cohorts of melanoma under anti-PD-1 treatment, on both bulk and single-cell levels. We reveal a surprising role of tumor-intrinsic SIRPA in enhancing antitumor immunity, in contrast to its well-established role as a major inhibitory immune modulator in macrophages. The loss of SIRPA expression is a marker of melanoma dedifferentiation, a key phenotype linked to immunotherapy efficacy. Inhibition of SIRPA in melanoma cells abrogates tumor killing by activated CD8+ T cells in a co-culture system. Mice bearing SIRPA-deficient melanoma tumors show no response to anti-PD-L1 treatment, whereas melanoma-specific SIRPA overexpression significantly enhances immunotherapy response. Mechanistically, SIRPA is regulated by its pseudogene, SIRPAP1. Our results suggest a complicated role of SIRPA in the tumor ecosystem, highlighting cell-type-dependent antagonistic effects of the same target on immunotherapy.