The Microbiome Promotes Pancreatic Cancer Progression
See article, p. 403
Gut bacteria migrate to the pancreas to increase immunosuppressive macrophages and suppress T cells.
Bacterial ablation enhances antitumor immunity by disrupting monocytic cell Toll-like receptor activation.
The gut microbiome may potentially be therapeutically targeted to enhance responses to immunotherapy.
The microbiome contributes to oncogenesis in a variety of tumor types in the intestinal tract where the gut microbiome is in direct contact with the tumor site. However, the effect of the microbiome in pancreatic ductal adenocarcinoma (PDA) is not well understood. Pushalkar, Hundeyin, and colleagues investigated the role of gut bacteria in pancreatic tumorigenesis. When administered to wild-type mice via oral gavage, bacteria migrated into the pancreas, suggesting that intestinal bacteria may directly affect the pancreatic microenvironment, and, in both mice and humans, PDA had an increased bacterial abundance compared with normal pancreas. In a mouse model of pancreatic cancer, mice grown in germ-free conditions exhibited reduced disease progression. Similarly, treatment with antibiotics reduced the tumor burden, and repopulation with feces or select bacterial species from untreated PDA-bearing mice accelerated tumorigenesis, suggesting that the PDA microbiome promotes tumor progression. Bacterial ablation reduced immunosuppressive CD206+ M2-like tumor-associated macrophages (TAM) and increased M1-like TAMs to enhance the antitumor immunity. After bacterial ablation, TAMs had an increased ability to activate T cells, and transferring tumor-infiltrating T cells from bacteria-ablated mice reduced the tumor burden in mice with a normal microbiome. Further, bacterial ablation upregulated PD-1 expression and enabled responsiveness to anti–PD-1 immunotherapy. Mechanistically, the microbiome differentially activated select Toll-like receptors in monocytic cells to promote a tolerogenic immune program. In addition to demonstrating that PDA tumors harbor a distinct and abundant microbiome that suppresses innate and adaptive antitumor immunity to support tumor progression, these findings suggest that the microbiome may potentially be therapeutically targeted in PDA.
Heterogeneous Resistance Mutations Reactivate MAPK after BRAF Inhibition
See article, p. 417
ERK inhibition may suppress resistance to BRAF inhibitor combinations in BRAFV600E colorectal cancer.
Analysis of cfDNA from patients identified 14 distinct MAPK pathway mutations that promote resistance.
Including ERK inhibitors with BRAF inhibitor combinations may improve clinical outcomes.
BRAFV600E mutations constitutively activate MAPK signaling and produce poor outcomes in approximately 10% of patients with colorectal cancer. However, single-agent BRAF inhibition (BRAFi) does not produce durable responses in colorectal cancer because reactivation of MAPK signaling by receptor tyrosine kinases such as EGFR drives resistance. BRAFi combination therapies including EGFRi and/or MEKi have produced better response rates, but the clinical benefit is limited by the rapid development of resistance. To explore mechanisms of resistance, Hazar-Rethinam and colleagues performed whole-exome sequencing of paired tumor biopsies and targeted sequencing of cell-free DNA (cfDNA) from four patients with BRAFV600E colorectal cancer treated with BRAFi combinations. In postprogression tumor and cfDNA samples, the original BRAFV600E mutation persisted, but acquired resistance mutations were heterogeneous. Fourteen distinct mutations converged on MAPK signaling, including mutations in KRAS, NRAS, MEK1, and MEK2, and adding ERKi completely suppressed acquired resistance in cells treated with BRAFi/EGFRi. Analysis of cfDNA from the culture media of heterogeneous pooled clones cultured in vitro, with 1% of cells harboring resistance mutations, allowed real-time assessment of clonal outgrowth to model resistance. In this assay rapid resistant outgrowth was observed with BRAFi/EGFRi, BRAF/MEKi, and BRAFi/MEKi/EGFRi, whereas triplet BRAFi/ERKi/EGFRi prevented outgrowth of resistant clones. Further, in tumor xenografts, BRAFi/ERKi/EGFRi induced durable tumor regression, whereas acquired resistance arose following BRAFi/EGFRi, BRAFi/MEKi, ERKi, BRAFi/MEKi/ERKi, and BRAFi/ERKi. Altogether, these findings suggest that ERKi may suppress acquired resistance in BRAFi combination therapies, supporting further investigation of convergent, up-front therapies including ERKi to improve outcomes in patients with BRAFV600E colorectal cancer.
Combined BRAF, EGFR, and MEK Inhibition May Suppress MAPK Reactivation
See article, p. 428
Triplet BRAF, EGFR, MEK inhibition achieves responses in patients with BRAFV600E colorectal cancer.
In patients with BRAFV600E colorectal cancer, inhibition of BRAF, EGFR, and MEK is tolerable.
Triplet BRAF, EGFR, MEK inhibition yields more complete MAPK suppression than dual inhibition.
Patients with BRAFV600-mutant colorectal cancer fail to respond to BRAF inhibitor monotherapy. In preclinical studies, resistance to BRAF inhibition occurs through EGFR-mediated reactivation of MAPK signaling. Dual BRAF and MEK inhibition led to modest improvements in response rates in patients with BRAFV600-mutant colorectal cancer, and Corcoran and colleagues hypothesized that targeting EGFR might further suppress MAPK reactivation to enhance the efficacy of BRAF inhibition. This hypothesis was tested in an open-label phase I trial of 142 patients with BRAFV600-mutant colorectal cancer evaluating the safety and efficacy of BRAF plus EGFR inhibition (in 20 patients receiving dabrafenib and panitumumab; D+P), MEK plus EGFR inhibition (in 31 patients receiving trametinib and panitumumab; T+P), or “triplet” BRAF, MEK, and EGFR inhibition (in 91 patients receiving dabrafenib, panitumumab, and trametinib; D+T+P). The primary endpoint was safety, and secondary endpoints included overall response rate and pharmacodynamics of drug combinations. D+T+P treatment was tolerable, although 18% of patients experienced an adverse event that resulted in therapy discontinuation. The overall response rates were 10% for D+P, 0% for T+P, and 21% for D+T+P, and D+T+P achieved the greatest reduction in MAPK pathway activity. Serial cfDNA analysis revealed that a reduction in BRAFV600E levels was associated with response and identified KRAS and NRAS mutations as putative resistance mechanisms associated with disease progression. Collectively, these findings suggest that combined inhibition of BRAF, EGFR, and MEK may suppress adaptive feedback pathways to improve responses in patients with BRAFV600-mutant colorectal cancer.
ctDNA Analysis Identifies Biomarkers of Resistance in Prostate Cancer
See article, p. 444
ctDNA sequencing was done on samples from patients enrolled in a prospective randomized clinical trial.
ctDNA analysis characterized the impact of common genomic alterations on response to AR inhibitors.
Truncating AR-GSRs may contribute to primary resistance to AR-targeted therapy in patients with mCRPC.
Abiraterone and enzalutamide are androgen receptor (AR)–targeting inhibitors that are currently first-line therapies for patients with metastatic castration-resistant prostate cancer (mCRPC). Analyses of plasma circulating tumor DNA (ctDNA) have identified AR amplification and AR splice variants as potential biomarkers of resistance to abiraterone and enzalutamide. To assess the relative impact of common ctDNA alterations on patient response to enzalutamide and abiraterone for advanced prostate cancer and perform a direct comparison of abiraterone to enzalutamide, Annala and colleagues performed an opportunistic exploratory analysis of baseline ctDNA from 202 patients, randomized to either enzalutamide (101) or abiraterone (101) treatment, by a combination of whole-exome sequencing and/or deep targeted sequencing of a panel of 72 prostate cancer–associated genes. They found that ctDNA recapitulated the somatic landscape of mCRPC and identified two patients with hypermutator phenotypes. Consistent with previous findings described in retrospective analyses, alterations in the p53 and PI3K pathways were independently predictive of poor response to AR-directed therapies, alterations in the homologous recombination repair genes BRCA2 and ATM were associated with primary resistance to AR-directed therapy, and diverse AR-genomic structural rearrangements (AR-GSR) were enriched in patients with AR amplification. Further, ctDNA profiling revealed that a subset of previously unidentified AR-GSRs contributes to primary resistance to AR-targeted therapy. Together, these results show that ctDNA profiling can stratify patients with mCRPC for AR-targeted therapy and identify potential genomic drivers of resistance to AR-targeted therapy.
BRD4 Regulates the Core Transcriptional Program in CLL
See article, p. 458
BRD4 profiling reveals enrichment at transcriptionally active sites and BCR pathway genes in CLL.
PLX51107, a structurally distinct BET inhibitor, reduces CLL growth and expression of CLL driver genes.
Targeting BRD4 may disrupt the transcriptional circuitry driving leukemogenesis in patients with CLL.
Chronic lymphocytic leukemia (CLL) is characterized by aberrant B-cell receptor (BCR) signaling, and the BCR pathway inhibitor ibrutinib has clinical activity in patients with CLL. However, responses are limited by mutations that reactivate BCR signaling; thus, new therapeutic approaches are needed. CLL has a low mutational burden, suggesting epigenetic control of pathogenic gene expression. The BET family protein BRD4 is an essential epigenetic regulator in many tumor types, prompting Ozer and colleagues to perform BRD4 profiling in B cells from four patients with CLL. BRD4 was overexpressed in CLL and enriched at transcriptionally active sites genome-wide in CLL cells compared with normal B cells. Further, BRD4 occupied sites proximal to key BCR signaling pathway genes. These findings suggest the potential for therapeutic targeting of BRD4, and scaffold screening using low-affinity binding assays followed by cocrystallography identified PLX51107 as a BET inhibitor structurally distinct from previous BET inhibitors. PLX51107 was well tolerated in mice, had activity at a 10-fold lower dose than the BET inhibitor OTX015, and had a short half-life, which might reduce toxicity. In vitro, PLX51107 suppressed CLL cell proliferation and modulated expression of CLL driver genes. In vivo, PLX51107 reduced the disease burden and extended survival in mouse models of CLL and also Richter's transformation. Collectively, these findings indicate that BRD4 regulates the core transcriptional programs in CLL and suggest it may be a therapeutic target. Further, these data support clinical investigation of PLX51107 in patients with CLL.
Phosphorylation of MEF2C Induces Chemoresistance in AML
See article, p. 478
MARK-mediated phosphorylation of MEF2C S222 is required for leukemic stem cell maintenance in AML.
MARK inhibition induces apoptosis and reverses chemoresistance in MEF2C-activated AML cells.
Therapeutic targeting of MARK may overcome primary chemoresistance in patients with AML.
Many patients with acute myeloid leukemia (AML) experience resistance to chemotherapy, but the mechanisms underlying chemoresistance are incompletely understood. To identify markers of chemoresistance, Brown and colleagues used high-accuracy mass spectrometry to determine the phospho-signaling profiles of human AML samples collected at diagnosis from patients who experienced chemoresistance. This approach revealed phosphorylation of S222 of MEF2C (MEF2C pS222), a transcription factor involved in hematopoietic self-renewal and differentiation, as a marker of primary chemoresistance. MEF2C pS222 was not required for normal hematopoiesis, but MEF2C phosphorylation was required for leukemogenesis in a mouse model of MLL–AF9-rearranged leukemia. MEF2C pS222 was essential for leukemic stem cell maintenance and cooperated with MLL–AF9 to induce leukemogenesis. Mutating MEF2C S222 to alanine to prevent phosphorylation suppressed the growth of MLL-rearranged leukemia cell xenografts. Further, MEF2C pS222 was required for stem cell maintenance in Runx1−/−;Flt3ITD leukemias lacking MLL–AF9. Mechanistically, MEF2C phosphorylation was required for its maximal transcriptional activation, and MEF2C was phosphorylated by the upstream microtubule affinity regulating kinase (MARK). Thus, MEF2C pS222 conferred sensitivity to MARK inhibitors, which induced apoptosis. MEF2C phosphorylation also induced chemotherapy resistance in vivo, which could be blocked by MARK inhibition. Taken together, these findings demonstrate that MEF2C S222 phosphorylation promotes chemoresistance in AML that may be reversed by MARK kinase inhibition, supporting further investigation of MARK inhibitors for the treatment of patients with AML.
ROS1 Inhibitors Target E-Cadherin–Defective Breast Cancer
See article, p. 498
ROS1 inhibition is synthetically lethal with E-cadherin deficiency in breast cancer cells.
Clinical ROS1 inhibitors show antitumor activity in models of E-cadherin– defective breast cancer.
ROS1 inhibition induces cytokinesis defects in part by further impairing p120 catenin function.
E-cadherin is as a cell adhesion protein that regulates cell–cell contact and contractility of epithelial cells and is frequently inactivated in lobular breast cancers. To identify potentially actionable targets that are synthetically lethal with E-cadherin deficiency, Bajrami and colleagues performed parallel siRNA and small-molecule drug sensitivity screens in breast cancer cells engineered with loss of E-cadherin expression. This integrated approach identified ROS1 inhibition as synthetically lethal with E-cadherin loss, and analysis of data from previous large-scale siRNA screens confirmed that this synthetic lethal effect was operative in a broad panel of molecularly diverse breast cancer cell lines as well as in cell lines derived from other tumor types. E-cadherin–defective breast cancer cells exhibited enhanced sensitivity to several ROS1 inhibitors compared with E-cadherin wild-type cells, and treatment with the clinical ROS1 inhibitors foretinib and crizotinib significantly inhibited tumor growth and prolonged survival in mouse models of E-cadherin–defective breast cancer. Mechanistically, E-cadherin cells exhibited impaired activity of p120 catenin, which controls actomyosin contractility during mitosis; inhibition of ROS1 further reduced p120 phosphorylation and localization at the cleavage furrow during cytokinesis in E-cadherin–defective, but not E-cadherin wild-type, cells. This effect was associated with cytokinesis defects in E-cadherin–defective cells, leading to abnormal mitoses, accumulation of multinucleated cells, and induction of apoptosis. These findings provide preclinical support for the initiation of clinical trials to assess the activity of clinically approved ROS1 inhibitors in patients with E-cadherin–defective breast cancer.
Note: In This Issue is written by Cancer Discovery editorial staff. Readers are encouraged to consult the original articles for full details.