Publications by authors named "Elisa de Stanchina"

170 Publications

Loss of FBXO31-mediated degradation of DUSP6 dysregulates ERK and PI3K-AKT signaling and promotes prostate tumorigenesis.

Cell Rep 2021 Oct;37(3):109870

Department of Biochemistry and Molecular Pharmacology, NYU Grossman School of Medicine, The Alexandria Center for Life Science, New York, NY 10016, USA; Laura and Isaac Perlmutter Cancer Center, NYU Grossman School of Medicine, The Alexandria Center for Life Science, New York, NY 10016, USA; Howard Hughes Medical Institute, NYU Grossman School of Medicine, The Alexandria Center for Life Science, New York, NY 10016, USA. Electronic address:

FBXO31 is the substrate receptor of one of many CUL1-RING ubiquitin ligase (CRL1) complexes. Here, we show that low FBXO31 mRNA levels are associated with high pre-operative prostate-specific antigen (PSA) levels and Gleason grade in human prostate cancer. Mechanistically, the ubiquitin ligase CRL1 promotes the ubiquitylation-mediated degradation of DUSP6, a dual specificity phosphatase that dephosphorylates and inactivates the extracellular-signal-regulated kinase-1 and -2 (ERK1/2). Depletion of FBXO31 stabilizes DUSP6, suppresses ERK signaling, and activates the PI3K-AKT signaling cascade. Moreover, deletion of FBXO31 promotes tumor development in a mouse orthotopic model of prostate cancer. Treatment with BCI, a small molecule inhibitor of DUSP6, suppresses AKT activation and prevents tumor formation, suggesting that the FBXO31 tumor suppressor activity is dependent on DUSP6. Taken together, our studies highlight the relevance of the FBXO31-DUSP6 axis in the regulation of ERK- and PI3K-AKT-mediated signaling pathways, as well as its therapeutic potential in prostate cancer.
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http://dx.doi.org/10.1016/j.celrep.2021.109870DOI Listing
October 2021

Comprehensive molecular characterization of lung tumors implicates AKT and MYC signaling in adenocarcinoma to squamous cell transdifferentiation.

J Hematol Oncol 2021 Oct 16;14(1):170. Epub 2021 Oct 16.

Department of Medicine, Thoracic Oncology Service, Memorial Sloan Kettering Cancer Center, 408 East 69th Street, ZRC-1731, New York, NY, 10021, USA.

Background: Lineage plasticity, the ability to transdifferentiate among distinct phenotypic identities, facilitates therapeutic resistance in cancer. In lung adenocarcinomas (LUADs), this phenomenon includes small cell and squamous cell (LUSC) histologic transformation in the context of acquired resistance to targeted inhibition of driver mutations. LUAD-to-LUSC transdifferentiation, occurring in up to 9% of EGFR-mutant patients relapsed on osimertinib, is associated with notably poor prognosis. We hypothesized that multi-parameter profiling of the components of mixed histology (LUAD/LUSC) tumors could provide insight into factors licensing lineage plasticity between these histologies.

Methods: We performed genomic, epigenomics, transcriptomics and protein analyses of microdissected LUAD and LUSC components from mixed histology tumors, pre-/post-transformation tumors and reference non-transformed LUAD and LUSC samples. We validated our findings through genetic manipulation of preclinical models in vitro and in vivo and performed patient-derived xenograft (PDX) treatments to validate potential therapeutic targets in a LUAD PDX model acquiring LUSC features after osimertinib treatment.

Results: Our data suggest that LUSC transdifferentiation is primarily driven by transcriptional reprogramming rather than mutational events. We observed consistent relative upregulation of PI3K/AKT, MYC and PRC2 pathway genes. Concurrent activation of PI3K/AKT and MYC induced squamous features in EGFR-mutant LUAD preclinical models. Pharmacologic inhibition of EZH1/2 in combination with osimertinib prevented relapse with squamous-features in an EGFR-mutant patient-derived xenograft model, and inhibition of EZH1/2 or PI3K/AKT signaling re-sensitized resistant squamous-like tumors to osimertinib.

Conclusions: Our findings provide the first comprehensive molecular characterization of LUSC transdifferentiation, suggesting putative drivers and potential therapeutic targets to constrain or prevent lineage plasticity.
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http://dx.doi.org/10.1186/s13045-021-01186-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8520275PMC
October 2021

Signatures of plasticity, metastasis, and immunosuppression in an atlas of human small cell lung cancer.

Cancer Cell 2021 Oct 12. Epub 2021 Oct 12.

Department of Medicine, Thoracic Oncology Service, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA; Weill Cornell Medical College, New York, NY 10065, USA.

Small cell lung cancer (SCLC) is an aggressive malignancy that includes subtypes defined by differential expression of ASCL1, NEUROD1, and POU2F3 (SCLC-A, -N, and -P, respectively). To define the heterogeneity of tumors and their associated microenvironments across subtypes, we sequenced 155,098 transcriptomes from 21 human biospecimens, including 54,523 SCLC transcriptomes. We observe greater tumor diversity in SCLC than lung adenocarcinoma, driven by canonical, intermediate, and admixed subtypes. We discover a PLCG2-high SCLC phenotype with stem-like, pro-metastatic features that recurs across subtypes and predicts worse overall survival. SCLC exhibits greater immune sequestration and less immune infiltration than lung adenocarcinoma, and SCLC-N shows less immune infiltrate and greater T cell dysfunction than SCLC-A. We identify a profibrotic, immunosuppressive monocyte/macrophage population in SCLC tumors that is particularly associated with the recurrent, PLCG2-high subpopulation.
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http://dx.doi.org/10.1016/j.ccell.2021.09.008DOI Listing
October 2021

INK4 tumor suppressor proteins mediate resistance to CDK4/6 kinase inhibitors.

Cancer Discov 2021 Sep 20. Epub 2021 Sep 20.

Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center

Cyclin-dependent kinases 4 and 6 (CDK4/6), represent a major therapeutic vulnerability for breast cancer. The kinases are clinically targeted via ATP competitive inhibitors (CDK4/6i); however, drug resistance commonly emerges over time. To understand CDK4/6i resistance, we surveyed over 1,300 breast cancers and identify several genetic alterations (e.g. FAT1, PTEN or ARID1A loss) converging on upregulation of CDK6. Mechanistically, we demonstrate CDK6 causes resistance by inducing and binding CDK inhibitor INK4 proteins (e.g. p18INK4C). In vitro binding and kinase assays together with physical modeling reveal that the p18INK4C/D-cyclin/CDK6 complex occludes CDK4/6i binding while only weakly suppressing ATP binding. Suppression of INK4 expression or its binding to CDK6 restores CDK4/6i sensitivity. To overcome this constraint, we developed bifunctional degraders conjugating palbociclib with E3 ligands. Two resulting lead compounds potently degraded CDK4/6, leading to substantial antitumor effects in vivo, demonstrating the promising therapeutic potential for retargeting CDK4/6 despite CDK4/6i resistance.
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http://dx.doi.org/10.1158/2159-8290.CD-20-1726DOI Listing
September 2021

Defining the therapeutic selective dependencies for distinct subtypes of PI3K pathway-altered prostate cancers.

Nat Commun 2021 08 20;12(1):5053. Epub 2021 Aug 20.

Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA.

Previous studies have suggested that PTEN loss is associated with p110β signaling dependency, leading to the clinical development of p110β-selective inhibitors. Here we use a panel pre-clinical models to reveal that PI3K isoform dependency is not governed by loss of PTEN and is impacted by feedback inhibition and concurrent PIK3CA/PIK3CB alterations. Furthermore, while pan-PI3K inhibition in PTEN-deficient tumors is efficacious, upregulation of Insulin Like Growth Factor 1 Receptor (IGF1R) promotes resistance. Importantly, we show that this resistance can be overcome through targeting AKT and we find that AKT inhibitors are superior to pan-PI3K inhibition in the context of PTEN loss. However, in the presence of wild-type PTEN and PIK3CA-activating mutations, p110α-dependent signaling is dominant and selectively inhibiting p110α is therapeutically superior to AKT inhibition. These discoveries reveal a more nuanced understanding of PI3K isoform dependency and unveil novel strategies to selectively target PI3K signaling nodes in a context-specific manner.
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http://dx.doi.org/10.1038/s41467-021-25341-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8379232PMC
August 2021

Rapid interrogation of cancer cell of origin through CRISPR editing.

Proc Natl Acad Sci U S A 2021 Aug;118(32)

Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065;

The increasing complexity of different cell types revealed by single-cell analysis of tissues presents challenges in efficiently elucidating their functions. Here we show, using prostate as a model tissue, that primary organoids and freshly isolated epithelial cells can be CRISPR edited ex vivo using Cas9-sgRNA (guide RNA) ribotnucleoprotein complex technology, then orthotopically transferred in vivo into immunocompetent or immunodeficient mice to generate cancer models with phenotypes resembling those seen in traditional genetically engineered mouse models. Large intrachromosomal (∼2 Mb) or multigenic deletions can be engineered efficiently without the need for selection, including in isolated subpopulations to address cell-of-origin questions.
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http://dx.doi.org/10.1073/pnas.2110344118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8364185PMC
August 2021

Bispecific CAR T Cells against EpCAM and Inducible ICAM-1 Overcome Antigen Heterogeneity and Generate Superior Antitumor Responses.

Cancer Immunol Res 2021 Oct 2;9(10):1158-1174. Epub 2021 Aug 2.

Molecular Imaging Innovations Institute, Department of Radiology, Weill Cornell Medicine, New York, New York.

Adoptive transfer of chimeric antigen receptor (CAR) T cells has demonstrated unparalleled responses in hematologic cancers, yet antigen escape and tumor relapse occur frequently. CAR T-cell therapy for patients with solid tumors faces even greater challenges due to the immunosuppressive tumor environment and antigen heterogeneity. Here, we developed a bispecific CAR to simultaneously target epithelial cell adhesion molecule (EpCAM) and intercellular adhesion molecule 1 (ICAM-1) to overcome antigen escape and to improve the durability of tumor responses. ICAM-1 is an adhesion molecule inducible by inflammatory cytokines and elevated in many types of tumors. Our study demonstrates superior efficacy of bispecific CAR T cells compared with CAR T cells targeting a single primary antigen. Bispecific CAR T achieved more durable antitumor responses in tumor models with either homogenous or heterogenous expression of EpCAM. We also showed that the activation of CAR T cells against EpCAM in tumors led to upregulation of ICAM-1, which rendered tumors more susceptible to ICAM-1 targeting by bispecific CAR T cells. Our strategy of additional targeting of ICAM-1 may have broad applications in augmenting the activity of CAR T cells against primary tumor antigens that are prone to antigen loss or downregulation.
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http://dx.doi.org/10.1158/2326-6066.CIR-21-0062DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8492509PMC
October 2021

ASN007 is a selective ERK1/2 inhibitor with preferential activity against RAS-and RAF-mutant tumors.

Cell Rep Med 2021 Jul 21;2(7):100350. Epub 2021 Jul 21.

Asana BioSciences, Lawrenceville, NJ, USA.

Inhibition of the extracellular signal-regulated kinases ERK1 and ERK2 (ERK1/2) offers a promising therapeutic strategy in cancers harboring activated RAS/RAF/MEK/ERK signaling pathways. Here, we describe an orally bioavailable and selective ERK1/2 inhibitor, ASN007, currently in clinical development for the treatment of cancer. In preclinical studies, ASN007 shows strong antiproliferative activity in tumors harboring mutations in BRAF and RAS (KRAS, NRAS, and HRAS). ASN007 demonstrates activity in a BRAF mutant melanoma tumor model that is resistant to BRAF and MEK inhibitors. The PI3K inhibitor copanlisib enhances the antiproliferative activity of ASN007 both and due to dual inhibition of RAS/MAPK and PI3K survival pathways. Our data provide a rationale for evaluating ASN007 in RAS/RAF-driven tumors as well as a mechanistic basis for combining ASN007 with PI3K inhibitors.
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http://dx.doi.org/10.1016/j.xcrm.2021.100350DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8324497PMC
July 2021

Multi-omic analysis of lung tumors defines pathways activated in neuroendocrine transformation.

Cancer Discov 2021 Jun 21. Epub 2021 Jun 21.

Memorial Sloan Kettering Cancer Center.

Lineage plasticity is implicated in treatment resistance in multiple cancers. In lung adenocarcinomas (LUADs) amenable to targeted therapy, transformation to small cell lung cancer (SCLC) is a recognized resistance mechanism. Defining molecular mechanisms of neuroendocrine (NE) transformation in lung cancer has been limited by a paucity of pre-/post-transformation clinical samples. Detailed genomic, epigenomic, transcriptomic, and protein characterization of combined LUAD/SCLC tumors, as well as pre-/post-transformation samples, support that NE transformation is primarily driven by transcriptional reprogramming rather than mutational events. We identify genomic contexts in which NE transformation is favored, including frequent loss of the 3p chromosome arm. We observed enhanced expression of genes involved in PRC2 complex and PI3K/AKT and NOTCH pathways. Pharmacological inhibition of the PI3K/AKT pathway delayed tumor growth and NE transformation in an EGFR-mutant patient-derived xenograft model. Our findings define a novel landscape of potential drivers and therapeutic vulnerabilities of neuroendocrine transformation in lung cancer.
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http://dx.doi.org/10.1158/2159-8290.CD-20-1863DOI Listing
June 2021

Tim-4 cavity-resident macrophages impair anti-tumor CD8 T cell immunity.

Cancer Cell 2021 Jul 10;39(7):973-988.e9. Epub 2021 Jun 10.

Departments of Surgery and Pathology, Center of Excellence for Cancer Immunology and Immunotherapy, University of Michigan Rogel Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA.

Immune checkpoint blockade (ICB) has been a remarkable clinical advance for cancer; however, the majority of patients do not respond to ICB therapy. We show that metastatic disease in the pleural and peritoneal cavities is associated with poor clinical outcomes after ICB therapy. Cavity-resident macrophages express high levels of Tim-4, a receptor for phosphatidylserine (PS), and this is associated with reduced numbers of CD8 T cells with tumor-reactive features in pleural effusions and peritoneal ascites from patients with cancer. We mechanistically demonstrate that viable and cytotoxic anti-tumor CD8 T cells upregulate PS and this renders them susceptible to sequestration away from tumor targets and proliferation suppression by Tim-4 macrophages. Tim-4 blockade abrogates this sequestration and proliferation suppression and enhances anti-tumor efficacy in models of anti-PD-1 therapy and adoptive T cell therapy in mice. Thus, Tim-4 cavity-resident macrophages limit the efficacy of immunotherapies in these microenvironments.
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http://dx.doi.org/10.1016/j.ccell.2021.05.006DOI Listing
July 2021

Co-inhibition of SMAD and MAPK signaling enhances 124I uptake in BRAF-mutant thyroid cancers.

Endocr Relat Cancer 2021 May 18;28(6):391-402. Epub 2021 May 18.

Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, New York, USA.

Constitutive MAPK activation silences genes required for iodide uptake and thyroid hormone biosynthesis in thyroid follicular cells. Accordingly, most BRAFV600E papillary thyroid cancers (PTC) are refractory to radioiodide (RAI) therapy. MAPK pathway inhibitors rescue thyroid-differentiated properties and RAI responsiveness in mice and patient subsets with BRAFV600E-mutant PTC. TGFB1 also impairs thyroid differentiation and has been proposed to mediate the effects of mutant BRAF. We generated a mouse model of BRAFV600E-PTC with thyroid-specific knockout of the Tgfbr1 gene to investigate the role of TGFB1 on thyroid-differentiated gene expression and RAI uptake in vivo. Despite appropriate loss of Tgfbr1, pSMAD levels remained high, indicating that ligands other than TGFB1 were engaging in this pathway. The activin ligand subunits Inhba and Inhbb were found to be overexpressed in BRAFV600E-mutant thyroid cancers. Treatment with follistatin, a potent inhibitor of activin, or vactosertib, which inhibits both TGFBR1 and the activin type I receptor ALK4, induced a profound inhibition of pSMAD in BRAFV600E-PTCs. Blocking SMAD signaling alone was insufficient to enhance iodide uptake in the setting of constitutive MAPK activation. However, combination treatment with either follistatin or vactosertib and the MEK inhibitor CKI increased 124I uptake compared to CKI alone. In summary, activin family ligands converge to induce pSMAD in Braf-mutant PTCs. Dedifferentiation of BRAFV600E-PTCs cannot be ascribed primarily to activation of SMAD. However, targeting TGFβ/activin-induced pSMAD augmented MAPK inhibitor effects on iodine incorporation into BRAF tumor cells, indicating that these two pathways exert interdependent effects on the differentiation state of thyroid cancer cells.
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http://dx.doi.org/10.1530/ERC-21-0017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8183640PMC
May 2021

Novel Preclinical Patient-Derived Lung Cancer Models Reveal Inhibition of HER3 and MTOR Signaling as Therapeutic Strategies for NRG1 Fusion-Positive Cancers.

J Thorac Oncol 2021 07 8;16(7):1149-1165. Epub 2021 Apr 8.

Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, New York; Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, New York.

Introduction: NRG1 rearrangements produce chimeric ligands that subvert the ERBB pathway to drive tumorigenesis. A better understanding of the signaling networks that mediate transformation by NRG1 fusions is needed to inform effective therapeutic strategies. Unfortunately, this has been hampered by a paucity of patient-derived disease models that faithfully recapitulate this molecularly defined cancer subset.

Methods: Patient-derived xenograft (PDX) and cell line models were established from NRG1-rearranged lung adenocarcinoma samples. Transcriptomic, proteomic, and biochemical analyses were performed to identify activated pathways. Efficacy studies were conducted to evaluate HER3- and MTOR-directed therapies.

Results: We established a pair of PDX and cell line models of invasive mucinous lung adenocarcinoma (LUAD) (LUAD-0061AS3, SLC3A2-NRG1), representing the first reported paired in vitro and in vivo model of NRG1-driven tumors. Growth of LUAD-0061AS3 models was reduced by the anti-HER3 antibody GSK2849330. Transcriptomic profiling revealed activation of the MTOR pathway in lung tumor samples with NRG1 fusions. Phosphorylation of several MTOR effectors (S6 and 4EBP1) was higher in LUAD-0061AS3 cells compared with human bronchial epithelial cells and the breast cancer cell line MDA-MB-175-VII (DOC4-NRG1 fusion). Accordingly, LUAD-0061AS3 cells were more sensitive to MTOR inhibitors than MDA-MB-175-VII cells and targeting the MTOR pathway with rapamycin blocked growth of LUAD-0061AS3 PDX tumors in vivo. In contrast, MDA-MB-175-VII breast cancer cells had higher MAPK pathway activation and were more sensitive to MEK inhibition.

Conclusions: We identify the MTOR pathway as a candidate vulnerability in NRG1 fusion-positive lung adenocarcinoma that may warrant further preclinical evaluation, with the eventual goal of finding additional therapeutic options for patients in whom ERBB-directed therapy fails. Moreover, our results uncover heterogeneity in downstream oncogenic signaling among NRG1-rearranged cancers, possibly tumor type-dependent, the therapeutic significance of which requires additional investigation.
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http://dx.doi.org/10.1016/j.jtho.2021.03.013DOI Listing
July 2021

Mutations in and differentially affect the tumor microenvironment and response to checkpoint blockade immunotherapy.

Nat Cancer 2021 Dec 16;1(12):1188-1203. Epub 2020 Nov 16.

Dept. of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY 10065.

Immune checkpoint blockade (ICB) has improved outcomes for patients with advanced cancer, but the determinants of response remain poorly understood. Here we report differential effects of mutations in the homologous recombination genes BRCA1 and BRCA2 on response to ICB in mouse and human tumors, and further show that truncating mutations in BRCA2 are associated with superior response compared to those in BRCA1. Mutations in BRCA1 and BRCA2 result in distinct mutational landscapes and differentially modulate the tumor-immune microenvironment, with gene expression programs related to both adaptive and innate immunity enriched in BRCA2-deficient tumors. Single-cell RNA sequencing further revealed distinct T cell, natural killer, macrophage, and dendritic cell populations enriched in BRCA2-deficient tumors. Taken together, our findings reveal the divergent effects of BRCA1 and BRCA2-deficiency on ICB outcome, and have significant implications for elucidating the genetic and microenvironmental determinants of response to immunotherapy.
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http://dx.doi.org/10.1038/s43018-020-00139-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8023400PMC
December 2021

The Anti-HER3 mAb Seribantumab Effectively Inhibits Growth of Patient-Derived and Isogenic Cell Line and Xenograft Models with Oncogenic Fusions.

Clin Cancer Res 2021 Jun 6;27(11):3154-3166. Epub 2021 Apr 6.

Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, New York.

Purpose: Oncogenic fusions involving the () gene are found in approximately 0.2% of cancers of diverse histologies. The resulting chimeric NRG1 proteins bind predominantly to HER3, leading to HER3-HER2 dimerization and activation of downstream growth and survival pathways. HER3 is, therefore, a rational target for therapy in NRG1 fusion-driven cancers.

Experimental Design: We developed novel patient-derived and isogenic models of NRG1-rearranged cancers and examined the effect of the anti-HER3 antibody, seribantumab, on growth and activation of signaling networks and .

Results: Seribantumab inhibited NRG1-stimulated growth of MCF-7 cells and growth of patient-derived breast (MDA-MB-175-VII, fusion) and lung (LUAD-0061AS3, fusion) cancer cells harboring fusions or amplification (HCC-95). In addition, seribantumab inhibited growth of isogenic HBEC cells expressing a fusion (HBECp53-CD74-NRG1) and induced apoptosis in MDA-MB-175-VII and LUAD-0061AS3 cells. Induction of proapoptotic proteins and reduced expression of the cell-cycle regulator, cyclin D1, were observed in seribantumab-treated cells. Treatment of MDA-MB-175-VII, LUAD-0061AS3, and HBECp53-CD74-NRG1 cells with seribantumab reduced phosphorylation of EGFR, HER2, HER3, HER4, and known downstream signaling molecules, such as AKT and ERK1/2. Significantly, administration of seribantumab to mice bearing LUAD-0061AS3 patient-derived xenograft (PDX) and OV-10-0050 (ovarian cancer with fusion) PDX tumors induced regression of tumors by 50%-100%. Afatinib was much less effective at blocking tumor growth.

Conclusions: Seribantumab treatment blocked activation of the four ERBB family members and of downstream signaling, leading to inhibition of fusion-dependent tumorigenesis and in breast, lung, and ovarian patient-derived cancer models.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-3605DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8172458PMC
June 2021

Design of BET Inhibitor Bottlebrush Prodrugs with Superior Efficacy and Devoid of Systemic Toxicities.

J Am Chem Soc 2021 03 19;143(12):4714-4724. Epub 2021 Mar 19.

Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, United States.

Prodrugs engineered for preferential activation in diseased versus normal tissues offer immense potential to improve the therapeutic indexes (TIs) of preclinical and clinical-stage active pharmaceutical ingredients that either cannot be developed otherwise or whose efficacy or tolerability it is highly desirable to improve. Such approaches, however, often suffer from trial-and-error design, precluding predictive synthesis and optimization. Here, using bromodomain and extra-terminal (BET) protein inhibitors (BETi)-a class of epigenetic regulators with proven anticancer potential but clinical development hindered in large part by narrow TIs-we introduce a macromolecular prodrug platform that overcomes these challenges. Through tuning of traceless linkers appended to a "bottlebrush prodrug" scaffold, we demonstrate correlation of prodrug activation kinetics with tumor pharmacokinetics, enabling the predictive design of novel BETi prodrugs with enhanced antitumor efficacies and devoid of dose-limiting toxicities in a syngeneic triple-negative breast cancer murine model. This work may have immediate clinical implications, introducing a platform for predictive prodrug design and potentially overcoming hurdles in drug development.
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http://dx.doi.org/10.1021/jacs.1c00312DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8317017PMC
March 2021

Targeting eIF4A-Dependent Translation of KRAS Signaling Molecules.

Cancer Res 2021 04 25;81(8):2002-2014. Epub 2021 Feb 25.

Cancer Biology and Genetics Program, Memorial Sloan-Kettering Cancer Center, New York, New York.

Pancreatic adenocarcinoma (PDAC) epitomizes a deadly cancer driven by abnormal KRAS signaling. Here, we show that the eIF4A RNA helicase is required for translation of key KRAS signaling molecules and that pharmacological inhibition of eIF4A has single-agent activity against murine and human PDAC models at safe dose levels. EIF4A was uniquely required for the translation of mRNAs with long and highly structured 5' untranslated regions, including those with multiple G-quadruplex elements. Computational analyses identified these features in mRNAs encoding KRAS and key downstream molecules. Transcriptome-scale ribosome footprinting accurately identified eIF4A-dependent mRNAs in PDAC, including critical KRAS signaling molecules such as PI3K, RALA, RAC2, MET, MYC, and YAP1. These findings contrast with a recent study that relied on an older method, polysome fractionation, and implicated redox-related genes as eIF4A clients. Together, our findings highlight the power of ribosome footprinting in conjunction with deep RNA sequencing in accurately decoding translational control mechanisms and define the therapeutic mechanism of eIF4A inhibitors in PDAC. SIGNIFICANCE: These findings document the coordinate, eIF4A-dependent translation of RAS-related oncogenic signaling molecules and demonstrate therapeutic efficacy of eIF4A blockade in pancreatic adenocarcinoma.
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http://dx.doi.org/10.1158/0008-5472.CAN-20-2929DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8137674PMC
April 2021

The serine hydroxymethyltransferase-2 (SHMT2) initiates lymphoma development through epigenetic tumor suppressor silencing.

Nat Cancer 2020 22;1:653-664. Epub 2020 Jun 22.

Cancer Biology and Genetics, Memorial Sloan-Kettering Cancer Center, New York, NY, USA.

Cancer cells adapt their metabolic activities to support growth and proliferation. However, increased activity of metabolic enzymes is not usually considered an initiating event in the malignant process. Here, we investigate the possible role of the enzyme serine hydroxymethyltransferase-2 (SHMT2) in lymphoma initiation. localizes to the most frequent region of copy number gains at chromosome 12q14.1 in lymphoma. Elevated expression of cooperates with in lymphoma development; loss or inhibition of impairs lymphoma cell survival. SHMT2 catalyzes the conversion of serine to glycine and produces an activated one-carbon unit that can be used to support -adenosyl methionine synthesis. SHMT2 induces changes in DNA and histone methylation patterns leading to promoter silencing of previously uncharacterized mutational genes, such as and Together, our findings reveal that amplification of in cooperation with is sufficient in the initiation of lymphomagenesis through epigenetic tumor suppressor silencing.
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http://dx.doi.org/10.1038/s43018-020-0080-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7872152PMC
June 2020

NRF2 Activation Confers Resistance to eIF4A Inhibitors in Cancer Therapy.

Cancers (Basel) 2021 Feb 5;13(4). Epub 2021 Feb 5.

Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.

Inhibition of the eIF4A RNA helicase with silvestrol and related compounds is emerging as a powerful anti-cancer strategy. We find that a synthetic silvestrol analogue (CR-1-31 B) has nanomolar activity across many cancer cell lines. It is especially active against aggressive MYC/BCL2 B cell lymphomas and this likely reflects the eIF4A-dependent translation of both MYC and BCL2. We performed a genome-wide CRISPR/Cas9 screen and identified mechanisms of resistance to this new class of therapeutics. We identify three negative NRF2 regulators (KEAP1, CUL3, CAND1) whose inactivation is sufficient to cause CR1-31-B resistance. NRF2 is known to alter the oxidation state of translation factors and cause a broad increase in protein production. We find that NRF2 activation particularly increases the translation of some eIF4A-dependent mRNAs and restores MYC and BCL2 production. We know that NRF2 functions depend on removal of sugar adducts by the frutosamine-3-kinase (FN3K). Accordingly, loss of FN3K results in NRF2 hyper-glycation and inactivation and resensitizes cancer cells to eIF4A inhibition. Together, our findings implicate NRF2 in the translation of eIF4A-dependent mRNAs and point to FN3K inhibition as a new strategy to block NRF2 functions in cancer.
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http://dx.doi.org/10.3390/cancers13040639DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7915661PMC
February 2021

Direct genome editing of patient-derived xenografts using CRISPR-Cas9 enables rapid functional genomics.

Nat Cancer 2020 Mar 9;1(3):359-369. Epub 2020 Mar 9.

Perlmutter Cancer Center, New York University Langone Health, New York, NY, USA.

Patient-derived xenografts are high fidelity tumor models that accurately reflect many key aspects of human cancer. In contrast to either cancer cell lines or genetically engineered mouse models, the utility of PDXs has been limited by the inability to perform targeted genome editing of these tumors. To address this limitation, we have developed methods for CRISPR-Cas9 editing of PDXs using a tightly regulated, inducible Cas9 vector that does not require culture for selection of transduced cells. We demonstrate the utility of this platform in PDXs (1) to analyze genetic dependencies by targeted gene disruption and (2) to analyze mechanisms of acquired drug resistance by site-specific gene editing using templated homology-directed repair. This flexible system has broad application to other explant models and substantially augments the utility of PDXs as genetically programmable models of human cancer.
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http://dx.doi.org/10.1038/s43018-020-0040-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7745982PMC
March 2020

Comparative Effectiveness Research Needs to Consider Optimal Dosing and Scheduling.

J Clin Oncol 2021 01 17;39(3):253-254. Epub 2020 Dec 17.

Andrew D. Seidman, MD, Elisa de Stanchina, PhD, and Larry Norton, MD, Memorial Sloan Kettering Cancer Center, New York, NY and Aki Morikawa, MD, University of Michigan, Ann Arbor, MI.

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http://dx.doi.org/10.1200/JCO.20.02355DOI Listing
January 2021

NTRK kinase domain mutations in cancer variably impact sensitivity to type I and type II inhibitors.

Commun Biol 2020 12 16;3(1):776. Epub 2020 Dec 16.

Department of Pediatrics, Oregon Health & Science University, Portland, OR, USA.

Tyrosine kinase domains dynamically fluctuate between two main structural forms that are referred to as type I (DFG-in) or type II (DFG-out) conformations. Comprehensive data comparing type I and type II inhibitors are currently lacking for NTRK fusion-driven cancers. Here we used a type II NTRK inhibitor, altiratinib, as a model compound to investigate its inhibitory potential for larotrectinib (type I inhibitor)-resistant mutations in NTRK. Our study shows that a subset of larotrectinib-resistant NTRK1 mutations (V573M, F589L and G667C) retains sensitivity to altiratinib, while the NTRK1 and xDFG motif NTRK1 mutations are highly sensitive to type II inhibitors, including altiratinib, cabozantinib and foretinib. Moreover, molecular modeling suggests that the introduction of a sulfur moiety in the binding pocket, via methionine or cysteine substitutions, specifically renders the mutant kinase hypersensitive to type II inhibitors. Future precision treatment strategies may benefit from selective targeting of these kinase mutants based on our findings.
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http://dx.doi.org/10.1038/s42003-020-01508-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7745027PMC
December 2020

RET inhibition in novel patient-derived models of RET-fusion positive lung adenocarcinoma reveals a role for MYC upregulation.

Dis Model Mech 2020 Dec 14. Epub 2020 Dec 14.

Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, NY, USA

Multi-kinase RET inhibitors, such as cabozantinib and RXDX-105, are active in lung cancer patients with fusions; however, the overall response rates to these two drugs are unsatisfactory compared to other targeted therapy paradigms. Moreover, these inhibitors may have different efficacies against rearrangements depending on the upstream fusion partner. A comprehensive preclinical analysis of the efficacy of RET inhibitors is lacking due to a paucity of disease models harboring rearrangements. Here we generated two new patient-derived xenograft (PDX) models, one new patient-derived cell line, one PDX-derived cell line, and several isogenic cell lines with RET fusions. Using these models, we re-examined the efficacy and mechanism of action of cabozantinib and found that this RET inhibitor was effective at blocking growth of cell lines, activating caspase 3/7 and inhibiting activation of ERK and AKT. Cabozantinib treatment of mice bearing RET-fusion-positive cell line xenografts and two PDXs significantly reduced tumor proliferation without adverse toxicity. Moreover, cabozantinib was effective at reducing growth of a lung cancer PDX that was not responsive to RXDX-105. Transcriptomic analysis of lung tumors and cell lines with RET alterations showed activation of a MYC signature and this was suppressed by treatment of cell lines with cabozantinib. MYC protein levels were rapidly depleted following cabozantinib treatment. Taken together, our results demonstrate that cabozantinib is an effective agent in preclinical models harboring rearrangements with three different 5' fusion partners (, and ). Notably, we identify MYC as a protein that is upregulated by RET expression and down-regulated by cabozantinib treatment, opening up potentially new therapeutic avenues for combinatorial targeting RET-fusion driven lung cancers. The novel RET fusion-dependent preclinical models described herein represent valuable tools for further refinement of current therapies and the evaluation of novel therapeutic strategies.
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http://dx.doi.org/10.1242/dmm.047779DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7888717PMC
December 2020

SWI/SNF Complex Mutations Promote Thyroid Tumor Progression and Insensitivity to Redifferentiation Therapies.

Cancer Discov 2021 05 14;11(5):1158-1175. Epub 2020 Dec 14.

Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, New York.

Mutations of subunits of the SWI/SNF chromatin remodeling complexes occur commonly in cancers of different lineages, including advanced thyroid cancers. Here we show that thyroid-specific loss of , or in mouse BRAF-mutant tumors promotes disease progression and decreased survival, associated with lesion-specific effects on chromatin accessibility and differentiation. As compared with normal thyrocytes, BRAF-mutant mouse papillary thyroid cancers have decreased lineage transcription factor expression and accessibility to their target DNA binding sites, leading to impairment of thyroid-differentiated gene expression and radioiodine incorporation, which is rescued by MAPK inhibition. Loss of individual SWI/SNF subunits in BRAF tumors leads to a repressive chromatin state that cannot be reversed by MAPK pathway blockade, rendering them insensitive to its redifferentiation effects. Our results show that SWI/SNF complexes are central to the maintenance of differentiated function in thyroid cancers, and their loss confers radioiodine refractoriness and resistance to MAPK inhibitor-based redifferentiation therapies. SIGNIFICANCE: Reprogramming cancer differentiation confers therapeutic benefit in various disease contexts. Oncogenic BRAF silences genes required for radioiodine responsiveness in thyroid cancer. Mutations in SWI/SNF genes result in loss of chromatin accessibility at thyroid lineage specification genes in -mutant thyroid tumors, rendering them insensitive to the redifferentiation effects of MAPK blockade..
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http://dx.doi.org/10.1158/2159-8290.CD-20-0735DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8102308PMC
May 2021

Concurrent Mutations in STK11 and KEAP1 Promote Ferroptosis Protection and SCD1 Dependence in Lung Cancer.

Cell Rep 2020 12;33(9):108444

Graduate Program in Pharmacology, Weill Cornell Medicine, New York, NY 10021, USA; Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA; Louis V. Gerstner Jr. Graduate School of Biomedical Sciences, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA; Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Electronic address:

Concurrent loss-of-function mutations in STK11 and KEAP1 in lung adenocarcinoma (LUAD) are associated with aggressive tumor growth, resistance to available therapies, and early death. We investigated the effects of coordinate STK11 and KEAP1 loss by comparing co-mutant with single mutant and wild-type isogenic counterparts in multiple LUAD models. STK11/KEAP1 co-mutation results in significantly elevated expression of ferroptosis-protective genes, including SCD and AKR1C1/2/3, and resistance to pharmacologically induced ferroptosis. CRISPR screening further nominates SCD (SCD1) as selectively essential in STK11/KEAP1 co-mutant LUAD. Genetic and pharmacological inhibition of SCD1 confirms the essentiality of this gene and augments the effects of ferroptosis induction by erastin and RSL3. Together these data identify SCD1 as a selective vulnerability and a promising candidate for targeted drug development in STK11/KEAP1 co-mutant LUAD.
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http://dx.doi.org/10.1016/j.celrep.2020.108444DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7722473PMC
December 2020

Therapeutic Potential of NTRK3 Inhibition in Desmoplastic Small Round Cell Tumor.

Clin Cancer Res 2021 02 23;27(4):1184-1194. Epub 2020 Nov 23.

Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, New York.

Purpose: Desmoplastic small round cell tumor (DSRCT) is a highly lethal intra-abdominal sarcoma of adolescents and young adults. DSRCT harbors a t(11;22)(p13:q12) that generates the EWSR1-WT1 chimeric transcription factor, the key oncogenic driver of DSRCT. EWSR1-WT1 rewires global gene expression networks and activates aberrant expression of targets that together mediate oncogenesis. EWSR1-WT1 also activates a neural gene expression program.

Experimental Design: Among these neural markers, we found prominent expression of neurotrophic tyrosine kinase receptor 3 (NTRK3), a druggable receptor tyrosine kinase. We investigated the regulation of NTRK3 by EWSR1-WT1 and its potential as a therapeutic target and , the latter using novel patient-derived models of DSRCT.

Results: We found that EWSR1-WT1 binds upstream of and activates its transcription. NTRK3 mRNA is highly expressed in DSRCT compared with other major chimeric transcription factor-driven sarcomas and most DSRCTs are strongly immunoreactive for NTRK3 protein. Remarkably, expression of kinase domain mRNA in DSRCT is also higher than in cancers with fusions. Abrogation of NTRK3 expression by RNAi silencing reduces growth of DSRCT cells and pharmacologic targeting of NTRK3 with entrectinib is effective in both and models of DSRCT.

Conclusions: Our results indicate that EWSR1-WT1 directly activates NTRK3 expression in DSRCT cells, which are dependent on its expression and activity for growth. Pharmacologic inhibition of NTRK3 by entrectinib significantly reduces growth of DSRCT cells both and , providing a rationale for clinical evaluation of NTRK3 as a therapeutic target in DSRCT.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-2585DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8212565PMC
February 2021

Targeting Germline- and Tumor-Associated Nucleotide Excision Repair Defects in Cancer.

Clin Cancer Res 2021 Apr 16;27(7):1997-2010. Epub 2020 Nov 16.

Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, New York.

Purpose: Nucleotide excision repair (NER) gene alterations constitute potential cancer therapeutic targets. We explored the prevalence of NER gene alterations across cancers and putative therapeutic strategies targeting these vulnerabilities.

Experimental Design: We interrogated our institutional dataset with mutational data from more than 40,000 patients with cancer to assess the frequency of putative deleterious alterations in four key NER genes. Gene-edited isogenic pairs of wild-type and mutant or cell lines were created and used to assess response to several candidate drugs.

Results: We found that putative damaging germline and somatic alterations in NER genes were present with frequencies up to 10% across multiple cancer types. Both and studies showed significantly enhanced sensitivity to the sesquiterpene irofulven in cells harboring specific clinically observed heterozygous mutations in or . Sensitivity of NER mutants to irofulven was greater than to a current standard-of-care agent, cisplatin. Hypomorphic -mutant cells had impaired ability to repair irofulven-induced DNA damage. Transcriptomic profiling of tumor tissues suggested codependencies between DNA repair pathways, indicating a potential benefit of combination therapies, which were confirmed by studies.

Conclusions: These findings provide novel insights into a synthetic lethal relationship between clinically observed NER gene deficiencies and sensitivity to irofulven and its potential synergistic combination with other drugs..
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http://dx.doi.org/10.1158/1078-0432.CCR-20-3322DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8191507PMC
April 2021

Functional Genomics In Vivo Reveal Metabolic Dependencies of Pancreatic Cancer Cells.

Cell Metab 2021 01 4;33(1):211-221.e6. Epub 2020 Nov 4.

Laboratory of Metabolic Regulation and Genetics, The Rockefeller University, New York, NY, USA. Electronic address:

Pancreatic ductal adenocarcinoma (PDAC) cells require substantial metabolic rewiring to overcome nutrient limitations and immune surveillance. However, the metabolic pathways necessary for pancreatic tumor growth in vivo are poorly understood. To address this, we performed metabolism-focused CRISPR screens in PDAC cells grown in culture or engrafted in immunocompetent mice. While most metabolic gene essentialities are unexpectedly similar under these conditions, a small fraction of metabolic genes are differentially required for tumor progression. Among these, loss of heme synthesis reduces tumor growth due to a limiting role of heme in vivo, an effect independent of tissue origin or immune system. Our screens also identify autophagy as a metabolic requirement for pancreatic tumor immune evasion. Mechanistically, autophagy protects cancer cells from CD8+ T cell killing through TNFα-induced cell death in vitro. Altogether, this resource provides metabolic dependencies arising from microenvironmental limitations and the immune system, nominating potential anti-cancer targets.
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http://dx.doi.org/10.1016/j.cmet.2020.10.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7790894PMC
January 2021

Discovery of IPN60090, a Clinical Stage Selective Glutaminase-1 (GLS-1) Inhibitor with Excellent Pharmacokinetic and Physicochemical Properties.

J Med Chem 2020 11 29;63(21):12957-12977. Epub 2020 Oct 29.

Institute for Applied Cancer Science (IACS), The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, United States.

Inhibition of glutaminase-1 (GLS-1) hampers the proliferation of tumor cells reliant on glutamine. Known glutaminase inhibitors have potential limitations, and in vivo exposures are potentially limited due to poor physicochemical properties. We initiated a GLS-1 inhibitor discovery program focused on optimizing physicochemical and pharmacokinetic properties, and have developed a new selective inhibitor, compound (IPN60090), which is currently in phase 1 clinical trials. Compound attains high oral exposures in preclinical species, with strong in vivo target engagement, and should robustly inhibit glutaminase in humans.
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http://dx.doi.org/10.1021/acs.jmedchem.0c01398DOI Listing
November 2020
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