Tumor innate immunity primed by specific interferon-stimulated endogenous retroviruses.

Mesenchymal tumor subpopulations secrete pro-tumorigenic cytokines and promote treatment resistance1-4. This phenomenon has been implicated in chemorefractory small cell lung cancer and resistance to targeted therapies5-8, but remains incompletely defined. Here, we identify a subclass of endogenous retroviruses (ERVs) that engages innate immune signaling in these cells. Stimulated 3 prime antisense retroviral coding sequences (SPARCS) are oriented inversely in 3' untranslated regions of specific genes enriched for regulation by STAT1 and EZH2. Derepression of these loci results in double-stranded RNA generation following IFN-γ exposure due to bi-directional transcription from the STAT1-activated gene promoter and the 5' long terminal repeat of the antisense ERV. Engagement of MAVS and STING activates downstream TBK1, IRF3, and STAT1 signaling, sustaining a positive feedback loop. SPARCS induction in human tumors is tightly associated with major histocompatibility complex class 1 expression, mesenchymal markers, and downregulation of chromatin modifying enzymes, including EZH2. Analysis of cell lines with high inducible SPARCS expression reveals strong association with an AXL/MET-positive mesenchymal cell state. While SPARCS-high tumors are immune infiltrated, they also exhibit multiple features of an immune-suppressed microenviroment. Together, these data unveil a subclass of ERVs whose derepression triggers pathologic innate immune signaling in cancer, with important implications for cancer immunotherapy

Characterizing genomic alterations in cancer by complementary functional associations.

Systematic efforts to sequence the cancer genome have identified large numbers of mutations and copy number alterations in human cancers. However, elucidating the functional consequences of these variants, and their interactions to drive or maintain oncogenic states, remains a challenge in cancer research. We developed REVEALER, a computational method that identifies combinations of mutually exclusive genomic alterations correlated with functional phenotypes, such as the activation or gene dependency of oncogenic pathways or sensitivity to a drug treatment. We used REVEALER to uncover complementary genomic alterations associated with the transcriptional activation of β-catenin and NRF2, MEK-inhibitor sensitivity, and KRAS dependency. REVEALER successfully identified both known and new associations, demonstrating the power of combining functional profiles with extensive characterization of genomic alterations in cancer genomes

Interaction of the Retinoblastoma Protein with Orc1 and Its Recruitment to Human Origins of DNA Replication

Background: The retinoblastoma protein (Rb) is a crucial regulator of cell cycle progression by binding with E2F transcription factor and repressing the expression of a variety of genes required for the G1-S phase transition. Methodology/Principal Findings: Here we show that Rb and E2F1 directly participate in the control of initiation of DNA replication in human HeLa, U2OS and T98G cells by specifically binding to origins of DNA replication in a cell cycle regulated manner. We show that, both in vitro and inside the cells, the largest subunit of the origin recognition complex (Orc1) specifically binds hypo-phosphorylated Rb and that this interaction is competitive with the binding of Rb to E2F1. The displacement of Rb-bound Orc1 by E2F1 at origins of DNA replication marks the progression of the G1 phase of the cell cycle toward the G1-S border. Conclusions/Significance: The participation of Rb and E2F1 in the formation of the multiprotein complex that binds origins of DNA replication in mammalian cells appears to represent an effective mechanism to couple the expression of gene

SIGNATURE: A workbench for gene expression signature analysis

<p>Abstract</p> <p>Background</p> <p>The biological phenotype of a cell, such as a characteristic visual image or behavior, reflects activities derived from the expression of collections of genes. As such, an ability to measure the expression of these genes provides an opportunity to develop more precise and varied sets of phenotypes. However, to use this approach requires computational methods that are difficult to implement and apply, and thus there is a critical need for intelligent software tools that can reduce the technical burden of the analysis. Tools for gene expression analyses are unusually difficult to implement in a user-friendly way because their application requires a combination of biological data curation, statistical computational methods, and database expertise.</p> <p>Results</p> <p>We have developed SIGNATURE, a web-based resource that simplifies gene expression signature analysis by providing software, data, and protocols to perform the analysis successfully. This resource uses Bayesian methods for processing gene expression data coupled with a curated database of gene expression signatures, all carried out within a GenePattern web interface for easy use and access.</p> <p>Conclusions</p> <p>SIGNATURE is available for public use at <url>http://genepattern.genome.duke.edu/signature/</url>.</p

Endocrine Therapy Synergizes with SMAC Mimetics to Potentiate Antigen Presentation and Tumor Regression in Hormone Receptor-Positive Breast Cancer.

UNLABELLED: Immunotherapies have yet to demonstrate significant efficacy in the treatment of hormone receptor-positive (HR+) breast cancer. Given that endocrine therapy (ET) is the primary approach for treating HR+ breast cancer, we investigated the effects of ET on the tumor immune microenvironment (TME) in HR+ breast cancer. Spatial proteomics of primary HR+ breast cancer samples obtained at baseline and after ET from patients enrolled in a neoadjuvant clinical trial (NCT02764541) indicated that ET upregulated β2-microglobulin and influenced the TME in a manner that promotes enhanced immunogenicity. To gain a deeper understanding of the underlying mechanisms, the intrinsic effects of ET on cancer cells were explored, which revealed that ET plays a crucial role in facilitating the chromatin binding of RelA, a key component of the NF-κB complex. Consequently, heightened NF-κB signaling enhanced the response to interferon-gamma, leading to the upregulation of β2-microglobulin and other antigen presentation-related genes. Further, modulation of NF-κB signaling using a SMAC mimetic in conjunction with ET augmented T-cell migration and enhanced MHC-I-specific T-cell-mediated cytotoxicity. Remarkably, the combination of ET and SMAC mimetics, which also blocks prosurvival effects of NF-κB signaling through the degradation of inhibitors of apoptosis proteins, elicited tumor regression through cell autonomous mechanisms, providing additional support for their combined use in HR+ breast cancer. SIGNIFICANCE: Adding SMAC mimetics to endocrine therapy enhances tumor regression in a cell autonomous manner while increasing tumor immunogenicity, indicating that this combination could be an effective treatment for HR+ patients with breast cancer

BAY61-3606 Affects the Viability of Colon Cancer Cells in a Genotype-Directed Manner

Background: K-RAS mutation poses a particularly difficult problem for cancer therapy. Activating mutations in K-RAS are common in cancers of the lung, pancreas, and colon and are associated with poor response to therapy. As such, targeted therapies that abrogate K-RAS-induced oncogenicity would be of tremendous value. Methods: We searched for small molecule kinase inhibitors that preferentially affect the growth of colorectal cancer cells expressing mutant K-RAS. The mechanism of action of one inhibitor was explored using chemical and genetic approaches. Results: We identified BAY61-3606 as an inhibitor of proliferation in colorectal cancer cells expressing mutant forms of K-RAS, but not in isogenic cells expressing wild-type K-RAS. In addition to its anti-proliferative effects in mutant cells, BAY61-3606 exhibited a distinct biological property in wild-type cells in that it conferred sensitivity to inhibition of RAF. In this context, BAY61-3606 acted by inhibiting MAP4K2 (GCK), which normally activates NFκβ signaling in wild-type cells in response to inhibition of RAF. As a result of MAP4K2 inhibition, wild-type cells became sensitive to AZ-628, a RAF inhibitor, when also treated with BAY61-3606. Conclusions: These studies indicate that BAY61-3606 exerts distinct biological activities in different genetic contexts

Treatment-Induced Tumor Dormancy through YAP-Mediated Transcriptional Reprogramming of the Apoptotic Pathway

Eradicating tumor dormancy that develops following epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) treatment of EGFR-mutant non-small cell lung cancer, is an attractive therapeutic strategy but the mechanisms governing this process are poorly understood. Blockade of ERK1/2 reactivation following EGFR TKI treatment by combined EGFR/MEK inhibition uncovers cells that survive by entering a senescence-like dormant state characterized by high YAP/TEAD activity. YAP/TEAD engage the epithelial-to-mesenchymal transition transcription factor SLUG to directly repress pro-apoptotic BMF, limiting drug-induced apoptosis. Pharmacological co-inhibition of YAP and TEAD, or genetic deletion of YAP1, all deplete dormant cells by enhancing EGFR/MEK inhibition-induced apoptosis. Enhancing the initial efficacy of targeted therapies could ultimately lead to prolonged treatment responses in cancer patients

Essential Role of Cdc42 in Ras-Induced Transformation Revealed by Gene Targeting

The ras proto-oncogene is one of the most frequently mutated genes in human cancer. However, given the prevalence of activating mutations in Ras and its association with aggressive forms of cancer, attempts to therapeutically target aberrant Ras signaling have been largely disappointing. This lack of progress highlights the deficiency in our understanding of cellular pathways required for Ras-mediated tumorigenesis and suggests the importance of identifying new molecular pathways associated with Ras-driven malignancies. Cdc42 is a Ras-related small GTPase that is known to play roles in oncogenic processes such as cell growth, survival, invasion, and migration. A pan-dominant negative mutant overexpression approach to suppress Cdc42 and related pathways has previously shown a requirement for Cdc42 in Ras-induced anchorage-independent cell growth, however the lack of specificity of such approaches make it difficult to determine if effects are directly related to changes in Cdc42 activity or other Rho family members. Therefore, in order to directly and unambiguously address the role of Cdc42 in Ras-mediated transformation, tumor formation and maintenance, we have developed a model of conditional cdc42 gene in Ras-transformed cells. Loss of Cdc42 drastically alters the cell morphology and inhibits proliferation, cell cycle progression and tumorigenicity of Ras-transformed cells, while non-transformed cells or c-Myc transformed cells are largely unaffected. The loss of Cdc42 in Ras-transformed cells results in reduced Akt signaling, restoration of which could partially rescues the proliferation defects associated with Cdc42 loss. Moreover, disruption of Cdc42 function in established tumors inhibited continued tumor growth. These studies implicate Cdc42 in Ras-driven tumor growth and suggest that targeting Cdc42 is beneficial in Ras-mediated malignancies

TBK1 Kinase Addiction in Lung Cancer Cells Is Mediated via Autophagy of Tax1bp1/Ndp52 and Non-Canonical NF-kappa B Signalling

K-Ras dependent non-small cell lung cancer (NSCLC) cells are 'addicted' to basal autophagy that reprograms cellular metabolism in a lysosomal-sensitive manner. Here we demonstrate that the xenophagy-associated kinase TBK1 drives basal autophagy, consistent with its known requirement in K-Ras-dependent NSCLC proliferation. Furthermore, basal autophagy in this context is characterised by sequestration of the xenophagy cargo receptor Ndp52 and its paralogue Tax1bp1, which we demonstrate here to be a bona fide cargo receptor. Autophagy of these cargo receptors promotes non-canonical NF-κB signalling. We propose that this TBK1-dependent mechanism for NF-κB signalling contributes to autophagy addiction in K-Ras driven NSCLC