Publications by authors named "Raul Rabadan"

186 Publications

Transcriptional control of CBX5 by the RNA binding proteins RBMX and RBMXL1 maintains chromatin state in myeloid leukemia.

Nat Cancer 2021 Jul 5;2:741-757. Epub 2021 Jul 5.

Molecular Pharmacology Program and Center for Cell Engineering, Center for Stem Cell Biology, Center for Experimental Therapeutics, Center for Hematologic Malignancies, Memorial Sloan Kettering Cancer Center, New York, NY, USA.

RNA binding proteins (RBPs) are key arbiters of post-transcriptional regulation and are found to be found dysregulated in hematological malignancies. Here, we identify the RBP RBMX and its retrogene RBMXL1 to be required for murine and human myeloid leukemogenesis. RBMX/L1 are overexpressed in acute myeloid leukemia (AML) primary patients compared to healthy individuals, and RBMX/L1 loss delayed leukemia development. RBMX/L1 loss lead to significant changes in chromatin accessibility, as well as chromosomal breaks and gaps. We found that RBMX/L1 directly bind to mRNAs, affect transcription of multiple loci, including (HP1α), and control the nascent transcription of the locus. Forced CBX5 expression rescued the RBMX/L1 depletion effects on cell growth and apoptosis. Overall, we determine that RBMX/L1 control leukemia cell survival by regulating chromatin state through their downstream target . These findings identify a mechanism for RBPs directly promoting transcription and suggest RBMX/L1, as well as CBX5, as potential therapeutic targets in myeloid malignancies.
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http://dx.doi.org/10.1038/s43018-021-00220-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8388313PMC
July 2021

TOP2B Enzymatic Activity on Promoters and Introns Modulates Multiple Oncogenes in Human Gliomas.

Clin Cancer Res 2021 Oct 25;27(20):5669-5680. Epub 2021 Aug 25.

Department of Neurosurgery, Feinberg School of Medicine, Northwestern University and Northwestern Medicine Malnati Brain Tumor Institute of the Robert H. Lurie Comprehensive Cancer Center, Feinberg School of Medicine, Northwestern University, Chicago, Illinois.

Purpose: The epigenetic mechanisms involved in transcriptional regulation leading to malignant phenotype in gliomas remains poorly understood. Topoisomerase IIB (TOP2B), an enzyme that decoils and releases torsional forces in DNA, is overexpressed in a subset of gliomas. Therefore, we investigated its role in epigenetic regulation in these tumors.

Experimental Design: To investigate the role of TOP2B in epigenetic regulation in gliomas, we performed paired chromatin immunoprecipitation sequencing for TOP2B and RNA-sequencing analysis of glioma cell lines with and without TOP2B inhibition and in human glioma specimens. These experiments were complemented with assay for transposase-accessible chromatin using sequencing, gene silencing, and mouse xenograft experiments to investigate the function of TOP2B and its role in glioma phenotypes.

Results: We discovered that TOP2B modulates transcription of multiple oncogenes in human gliomas. TOP2B regulated transcription only at sites where it was enzymatically active, but not at all native binding sites. In particular, TOP2B activity localized in enhancers, promoters, and introns of PDGFRA and MYC, facilitating their expression. TOP2B levels and genomic localization was associated with and expression across glioma specimens, which was not seen in nontumoral human brain tissue. , TOP2B knockdown of human glioma intracranial implants prolonged survival and downregulated .

Conclusions: Our results indicate that TOP2B activity exerts a pleiotropic role in transcriptional regulation of oncogenes in a subset of gliomas promoting a proliferative phenotype.
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http://dx.doi.org/10.1158/1078-0432.CCR-21-0312DOI Listing
October 2021

A persistent look at how tumours evade therapy.

Nature 2021 08;596(7873):491-493

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http://dx.doi.org/10.1038/d41586-021-02117-1DOI Listing
August 2021

Recombination and lineage-specific mutations linked to the emergence of SARS-CoV-2.

Genome Med 2021 08 6;13(1):124. Epub 2021 Aug 6.

Program for Mathematical Genomics, Columbia University, New York, NY, USA.

Background: The emergence of SARS-CoV-2 underscores the need to better understand the evolutionary processes that drive the emergence and adaptation of zoonotic viruses in humans. In the betacoronavirus genus, which also includes SARS-CoV and MERS-CoV, recombination frequently encompasses the receptor binding domain (RBD) of the Spike protein, which is responsible for viral binding to host cell receptors. In this work, we reconstruct the evolutionary events that have accompanied the emergence of SARS-CoV-2, with a special emphasis on the RBD and its adaptation for binding to its receptor, human ACE2.

Methods: By means of phylogenetic and recombination analyses, we found evidence of a recombination event in the RBD involving ancestral linages to both SARS-CoV and SARS-CoV-2. We then assessed the effect of this recombination at protein level by reconstructing the RBD of the closest ancestors to SARS-CoV-2, SARS-CoV, and other Sarbecoviruses, including the most recent common ancestor of the recombining clade. The resulting information was used to measure and compare, in silico, their ACE2-binding affinities using the physics-based trRosetta algorithm.

Results: We show that, through an ancestral recombination event, SARS-CoV and SARS-CoV-2 share an RBD sequence that includes two insertions (positions 432-436 and 460-472), as well as the variants 427N and 436Y. Both 427N and 436Y belong to a helix that interacts directly with the human ACE2 (hACE2) receptor. Reconstruction of ancestral states, combined with protein-binding affinity analyses, suggests that the recombination event involving ancestral strains of SARS-CoV and SARS-CoV-2 led to an increased affinity for hACE2 binding and that alleles 427N and 436Y significantly enhanced affinity as well.

Conclusions: We report an ancestral recombination event affecting the RBD of both SARS-CoV and SARS-CoV-2 that was associated with an increased binding affinity to hACE2. Structural modeling indicates that ancestors of SARS-CoV-2 may have acquired the ability to infect humans decades ago. The binding affinity with the human receptor would have been subsequently boosted in SARS-CoV and SARS-CoV-2 through further mutations in RBD.
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http://dx.doi.org/10.1186/s13073-021-00943-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8343217PMC
August 2021

A Phase I Study of the Combination of Pexidartinib and Sirolimus to Target Tumor-Associated Macrophages in Unresectable Sarcoma and Malignant Peripheral Nerve Sheath Tumors.

Clin Cancer Res 2021 Oct 28;27(20):5519-5527. Epub 2021 Jul 28.

Division of Hematology and Oncology, Columbia University Irving Medical Center, New York, New York.

Purpose: To evaluate the safety and tolerability in phase I first-in-human combination therapy with pexidartinib, an inhibitor of colony-stimulating factor-1 receptor, and sirolimus, an mTOR inhibitor, to target tumor-associated macrophage (TAM) polarization in soft tissue sarcomas (STS).

Patients And Methods: This multicenter phase I study used the time-to-event continual reassessment method (TITE-CRM) to study the combination of sirolimus, doses ranging from 2 to 6 mg, with pexidartinib, doses ranging from 600 to 1,000 mg, both provided continuously on a 28-day cycle, in patients with advanced sarcoma. A total of 24 patients [8 malignant peripheral nerve sheath tumor, 3 tenosynovial giant cell tumor (TGCT), 5 leiomyosarcoma, and 8 with other sarcoma subtypes] were enrolled. The median age was 46 years, 56% were male, and 61% had >2 prior lines of therapy.

Results: The recommended phase II dose was 2 mg of sirolimus combined with 1,000 mg of pexidartinib daily. Of the 18 evaluable subjects, 5 experienced dose-limiting toxicities (2 elevated aspartate aminotransferase/alanine aminotransferase, 2 elevated sirolimus trough levels, and 1 grade 5 dehydration). Most common grade 2 or higher treatment-related adverse events included anemia, fatigue, neutropenia, and lymphopenia. Clinical benefit was observed in 12 of 18 (67%) evaluable subjects with 3 partial responses (all in TGCT) and 9 stable disease. Tissue staining indicated a decreased proportion of activated M2 macrophages within tumor samples with treatment.

Conclusions: Pexidartinib can be safely administered with sirolimus. These findings support further investigation of this combination to determine clinical efficacy. Clinicaltrials.gov identifier NCT02584647.
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http://dx.doi.org/10.1158/1078-0432.CCR-21-1779DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8530953PMC
October 2021

iPLA2β-mediated lipid detoxification controls p53-driven ferroptosis independent of GPX4.

Nat Commun 2021 06 15;12(1):3644. Epub 2021 Jun 15.

Institute for Cancer Genetics, and Department of Pathology and Cell Biology, and Herbert Irving Comprehensive Cancer Center,Vagelos College of Physicians & Surgeons, Columbia University, New York, NY, 10032, USA.

Here, we identify iPLA2β as a critical regulator for p53-driven ferroptosis upon reactive oxygen species (ROS)-induced stress. The calcium-independent phospholipase iPLA2β is known to cleave acyl tails from the glycerol backbone of lipids and release oxidized fatty acids from phospholipids. We found that iPLA2β-mediated detoxification of peroxidized lipids is sufficient to suppress p53-driven ferroptosis upon ROS-induced stress, even in GPX4-null cells. Moreover, iPLA2β is overexpressed in human cancers; inhibition of endogenous iPLA2β sensitizes tumor cells to p53-driven ferroptosis and promotes p53-dependent tumor suppression in xenograft mouse models. These results demonstrate that iPLA2β acts as a major ferroptosis repressor in a GPX4-independent manner. Notably, unlike GPX4, loss of iPLA2β has no obvious effect on normal development or cell viability in normal tissues but iPLA2β plays an essential role in regulating ferroptosis upon ROS-induced stress. Thus, our study suggests that iPLA2β is a promising therapeutic target for activating ferroptosis-mediated tumor suppression without serious toxicity concerns.
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http://dx.doi.org/10.1038/s41467-021-23902-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8206155PMC
June 2021

A MYC and RAS co-activation signature in localized prostate cancer drives bone metastasis and castration resistance.

Nat Cancer 2020 Nov 19;1(11):1082-1096. Epub 2020 Oct 19.

Department of Molecular Pharmacology and Therapeutics, Columbia University Irving Medical Center, New York, NY, USA.

Understanding the intricacies of lethal prostate cancer poses specific challenges due to difficulties in accurate modeling of metastasis in vivo. Here we show that mice (for -) develop prostate cancer with a high penetrance of metastasis to bone, thereby enabling detection and tracking of bone metastasis in vivo and ex vivo. Transcriptomic and whole-exome analyses of bone metastasis from these mice revealed distinct molecular profiles conserved between human and mouse and specific patterns of subclonal branching from the primary tumor. Integrating bulk and single-cell transcriptomic data from mouse and human datasets with functional studies in vivo unravels a unique MYC/RAS co-activation signature associated with prostate cancer metastasis. Finally, we identify a gene signature with prognostic value for time to metastasis and predictive of treatment response in human patients undergoing androgen receptor therapy across clinical cohorts, thus uncovering conserved mechanisms of metastasis with potential translational significance.
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http://dx.doi.org/10.1038/s43018-020-00125-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8171279PMC
November 2020

Genetic mechanisms of HLA-I loss and immune escape in diffuse large B cell lymphoma.

Proc Natl Acad Sci U S A 2021 Jun;118(22)

Institute for Cancer Genetics, Columbia University, New York, NY 10032;

Fifty percent of diffuse large B cell lymphoma (DLBCL) cases lack cell-surface expression of the class I major histocompatibility complex (MHC-I), thus escaping recognition by cytotoxic T cells. Here we show that, across B cell lymphomas, loss of MHC-I, but not MHC-II, is preferentially restricted to DLBCL. To identify the involved mechanisms, we performed whole exome and targeted HLA deep-sequencing in 74 DLBCL samples, and found somatic inactivation of and the loci in 80% (34 of 42) of MHC-I tumors. Furthermore, 70% (22 of 32) of MHC-I DLBCLs harbored monoallelic HLA-I genetic alterations (MHC-I), indicating allele-specific inactivation. MHC-I and MHC-I cases harbored significantly higher mutational burden and inferred neoantigen load, suggesting potential coselection of loss and sustained neoantigen production. Notably, the analysis of >500,000 individuals across different cancer types revealed common germline homozygosity, preferentially in DLBCL. In mice, germinal-center B cells lacking HLA-I expression did not progress to lymphoma and were counterselected in the context of oncogene-driven lymphomagenesis, suggesting that additional events are needed to license immune evasion. These results suggest a multistep process of loss in DLBCL development including both germline and somatic events, and have direct implications for the pathogenesis and immunotherapeutic targeting of this disease.
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http://dx.doi.org/10.1073/pnas.2104504118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8179151PMC
June 2021

Single-cell characterization of macrophages in glioblastoma reveals MARCO as a mesenchymal pro-tumor marker.

Genome Med 2021 May 19;13(1):88. Epub 2021 May 19.

Department of Systems Biology, Columbia University Irving Medical Center, New York, NY, USA.

Background: Macrophages are the most common infiltrating immune cells in gliomas and play a wide variety of pro-tumor and anti-tumor roles. However, the different subpopulations of macrophages and their effects on the tumor microenvironment remain poorly understood.

Methods: We combined new and previously published single-cell RNA-seq data from 98,015 single cells from a total of 66 gliomas to profile 19,331 individual macrophages.

Results: Unsupervised clustering revealed a pro-tumor subpopulation of bone marrow-derived macrophages characterized by the scavenger receptor MARCO, which is almost exclusively found in IDH1-wild-type glioblastomas. Previous studies have implicated MARCO as an unfavorable marker in melanoma and non-small cell lung cancer; here, we find that bulk MARCO expression is associated with worse prognosis and mesenchymal subtype. Furthermore, MARCO expression is significantly altered over the course of treatment with anti-PD1 checkpoint inhibitors in a response-dependent manner, which we validate with immunofluorescence imaging.

Conclusions: These findings illustrate a novel macrophage subpopulation that drives tumor progression in glioblastomas and suggest potential therapeutic targets to prevent their recruitment.
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http://dx.doi.org/10.1186/s13073-021-00906-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8136167PMC
May 2021

Promotion of cholangiocarcinoma growth by diverse cancer-associated fibroblast subpopulations.

Cancer Cell 2021 Jun 29;39(6):866-882.e11. Epub 2021 Apr 29.

Liver Cancer Translational Research Laboratory, Institut d'Investigacions Biomèdiques August Pi i Sunyer, Hospital Clínic, Universitat de Barcelona, 08036 Barcelona, Spain; Mount Sinai Liver Cancer Program, Divisions of Liver Diseases, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Institució Catalana de Recerca i Estudis Avançats, Barcelona, Spain.

Cancer-associated fibroblasts (CAF) are a poorly characterized cell population in the context of liver cancer. Our study investigates CAF functions in intrahepatic cholangiocarcinoma (ICC), a highly desmoplastic liver tumor. Genetic tracing, single-cell RNA sequencing, and ligand-receptor analyses uncovered hepatic stellate cells (HSC) as the main source of CAF and HSC-derived CAF as the dominant population interacting with tumor cells. In mice, CAF promotes ICC progression, as revealed by HSC-selective CAF depletion. In patients, a high panCAF signature is associated with decreased survival and increased recurrence. Single-cell RNA sequencing segregates CAF into inflammatory and growth factor-enriched (iCAF) and myofibroblastic (myCAF) subpopulations, displaying distinct ligand-receptor interactions. myCAF-expressed hyaluronan synthase 2, but not type I collagen, promotes ICC. iCAF-expressed hepatocyte growth factor enhances ICC growth via tumor-expressed MET, thus directly linking CAF to tumor cells. In summary, our data demonstrate promotion of desmoplastic ICC growth by therapeutically targetable CAF subtype-specific mediators, but not by type I collagen.
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http://dx.doi.org/10.1016/j.ccell.2021.03.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8241235PMC
June 2021

induces NF-κB signaling-driven peripheral T cell lymphoma.

Nat Cancer 2021 Jan 13;2(1):98-113. Epub 2021 Jan 13.

Institute for Cancer Genetics, Columbia University, New York, NY, 10032, USA.

Angioimmunoblastic T cell lymphoma (AITL) and peripheral T cell lymphoma not-otherwise-specified (PTCL, NOS) have poor prognosis and lack driver actionable targets for directed therapies in most cases. Here we identify as a recurrent oncogenic gene fusion in AITL and PTCL, NOS tumors. Mechanistically, we show that FYN-TRAF3IP2 leads to aberrant NF-κB signaling downstream of T cell receptor activation. Consistent with a driver oncogenic role, FYN-TRAF3IP2 expression in hematopoietic progenitors induces NF-κB-driven T cell transformation in mice and cooperates with loss of the tumor suppressor in PTCL development. Moreover, abrogation of NF-κB signaling in -induced tumors with IκB kinase inhibitors delivers strong anti-lymphoma effects and . These results demonstrate an oncogenic and pharmacologically targetable role for FYN-TRAF3IP2 in PTCLs and call for the clinical testing of anti-NF-κB targeted therapies in these diseases.
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http://dx.doi.org/10.1038/s43018-020-00161-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8081346PMC
January 2021

Mutational and functional genetics mapping of chemotherapy resistance mechanisms in relapsed acute lymphoblastic leukemia.

Nat Cancer 2020 Nov 19;1(11):1113-1127. Epub 2020 Oct 19.

Institute for Cancer Genetics, Columbia University, New York, NY, USA.

Multi-agent combination chemotherapy can be curative in acute lymphoblastic leukemia (ALL). Still, patients with primary refractory disease or with relapsed leukemia have a very poor prognosis. Here we integrate an in-depth dissection of the mutational landscape across diagnostic and relapsed pediatric and adult ALL samples with genome-wide CRISPR screen analysis of gene-drug interactions across seven ALL chemotherapy drugs. By combining these analyses, we uncover diagnostic and relapse-specific mutational mechanisms as well as genetic drivers of chemoresistance. Functionally, our data identifies common and drug-specific pathways modulating chemotherapy response and underscores the effect of drug combinations in restricting the selection of resistance-driving genetic lesions. In addition, by identifying actionable targets for the reversal of chemotherapy resistance, these analyses open novel therapeutic opportunities for the treatment of relapse and refractory disease.
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http://dx.doi.org/10.1038/s43018-020-00124-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8011577PMC
November 2020

PDK1 Is Required for Maintenance of CD4 Foxp3 Regulatory T Cell Function.

J Immunol 2021 04 31;206(8):1776-1783. Epub 2021 Mar 31.

Department of Microbiology and Immunology, Vagelos College of Physicians and Surgeons, Columbia University Irving Medical Center, New York, NY 10032;

Regulatory T (Treg) cells have an essential role in maintaining immune homeostasis, in part by suppressing effector T cell functions. Phosphoinositide-dependent kinase 1 (PDK1) is a pleiotropic kinase that acts as a key effector downstream of PI3K in many cell types. In T cells, PDK1 has been shown to be critical for activation of NF-κB and AKT signaling upon TCR ligation and is therefore essential for effector T cell activation, proliferation, and cytokine production. Using Treg cell-specific conditional deletion, we now demonstrate that PDK1 is also essential for Treg cell suppressive activity in vivo. Ablation of specifically in Treg cells led to systemic, lethal, -like inflammation in mice. Genome-wide analysis confirmed that PDK1 is essential for the regulation of key Treg cell signature gene expression and, further, suggested that PDK1 acts primarily to control Treg cell gene expression through regulation of the canonical NF-κB pathway. Consistent with these results, the like phenotype of mice lacking PDK1 in Treg cells was rescued by enforced activation of NF-κB downstream of PDK1. Therefore, PDK1-mediated activation of the NF-κB signaling pathway is essential for regulation of Treg cell signature gene expression and suppressor function.
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http://dx.doi.org/10.4049/jimmunol.2000051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8026672PMC
April 2021

Functional interrogation of DNA damage response variants with base editing screens.

Cell 2021 02;184(4):1081-1097.e19

Department of Genetics and Development, Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY 10032, USA. Electronic address:

Mutations in DNA damage response (DDR) genes endanger genome integrity and predispose to cancer and genetic disorders. Here, using CRISPR-dependent cytosine base editing screens, we identify > 2,000 sgRNAs that generate nucleotide variants in 86 DDR genes, resulting in altered cellular fitness upon DNA damage. Among those variants, we discover loss- and gain-of-function mutants in the Tudor domain of the DDR regulator 53BP1 that define a non-canonical surface required for binding the deubiquitinase USP28. Moreover, we characterize variants of the TRAIP ubiquitin ligase that define a domain, whose loss renders cells resistant to topoisomerase I inhibition. Finally, we identify mutations in the ATM kinase with opposing genome stability phenotypes and loss-of-function mutations in the CHK2 kinase previously categorized as variants of uncertain significance for breast cancer. We anticipate that this resource will enable the discovery of additional DDR gene functions and expedite studies of DDR variants in human disease.
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http://dx.doi.org/10.1016/j.cell.2021.01.041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8018281PMC
February 2021

Global Patterns of Recombination across Human Viruses.

Mol Biol Evol 2021 05;38(6):2520-2531

Program for Mathematical Genomics, Departments of Systems Biology and Biomedical Informatics, Columbia University, New York, NY, USA.

Viral recombination is a major evolutionary mechanism driving adaptation processes, such as the ability of host-switching. Understanding global patterns of recombination could help to identify underlying mechanisms and to evaluate the potential risks of rapid adaptation. Conventional approaches (e.g., those based on linkage disequilibrium) are computationally demanding or even intractable when sequence alignments include hundreds of sequences, common in viral data sets. We present a comprehensive analysis of recombination across 30 genomic alignments from viruses infecting humans. In order to scale the analysis and avoid the computational limitations of conventional approaches, we apply newly developed topological data analysis methods able to infer recombination rates for large data sets. We show that viruses, such as ZEBOV and MARV, consistently displayed low levels of recombination, whereas high levels of recombination were observed in Sarbecoviruses, HBV, HEV, Rhinovirus A, and HIV. We observe that recombination is more common in positive single-stranded RNA viruses than in negatively single-stranded RNA ones. Interestingly, the comparison across multiple viruses suggests an inverse correlation between genome length and recombination rate. Positional analyses of recombination breakpoints along viral genomes, combined with our approach, detected at least 39 nonuniform patterns of recombination (i.e., cold or hotspots) in 18 viral groups. Among these, noteworthy hotspots are found in MERS-CoV and Sarbecoviruses (at spike, Nucleocapsid and ORF8). In summary, we have developed a fast pipeline to measure recombination that, combined with other approaches, has allowed us to find both common and lineage-specific patterns of recombination among viruses with potential relevance in viral adaptation.
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http://dx.doi.org/10.1093/molbev/msab046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8136497PMC
May 2021

Comprehensive characterization of protein-protein interactions perturbed by disease mutations.

Nat Genet 2021 03 8;53(3):342-353. Epub 2021 Feb 8.

Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.

Technological and computational advances in genomics and interactomics have made it possible to identify how disease mutations perturb protein-protein interaction (PPI) networks within human cells. Here, we show that disease-associated germline variants are significantly enriched in sequences encoding PPI interfaces compared to variants identified in healthy participants from the projects 1000 Genomes and ExAC. Somatic missense mutations are also significantly enriched in PPI interfaces compared to noninterfaces in 10,861 tumor exomes. We computationally identified 470 putative oncoPPIs in a pan-cancer analysis and demonstrate that oncoPPIs are highly correlated with patient survival and drug resistance/sensitivity. We experimentally validate the network effects of 13 oncoPPIs using a systematic binary interaction assay, and also demonstrate the functional consequences of two of these on tumor cell growth. In summary, this human interactome network framework provides a powerful tool for prioritization of alleles with PPI-perturbing mutations to inform pathobiological mechanism- and genotype-based therapeutic discovery.
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http://dx.doi.org/10.1038/s41588-020-00774-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8237108PMC
March 2021

Computing the Role of Alternative Splicing in Cancer.

Trends Cancer 2021 04 23;7(4):347-358. Epub 2021 Jan 23.

Program for Mathematical Genomics, Columbia University, New York, NY 10032, USA; Departments of Systems Biology and Biomedical Informatics, Columbia University, New York, NY 10032, USA. Electronic address:

Most human genes undergo alternative splicing (AS), and dysregulation of alternative splicing contributes to tumor initiation and progression. Computational analysis of genomic and transcriptomic data enables the systematic characterization of alternative splicing and its functional role in cancer. In this review, we summarize the latest computational approaches to studying alternative splicing in cancer and the current limitations of the most popular tools in this field. Finally, we describe some of the current computational challenges in the characterization of the role of alternative splicing in cancer.
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http://dx.doi.org/10.1016/j.trecan.2020.12.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7969404PMC
April 2021

Dose-response modeling in high-throughput cancer drug screenings: an end-to-end approach.

Biostatistics 2021 Jan 7. Epub 2021 Jan 7.

Data Science Institute, Columbia University and Columbia University Medical Center, New York, NY, USA, Department of Applied Physics and Applied Mathematics, Columbia University and Columbia University Medical Center, New York, NY, USA and Department of Systems Biology, Columbia University and Columbia University Medical Center, New York, NY, USA.

Personalized cancer treatments based on the molecular profile of a patient's tumor are an emerging and exciting class of treatments in oncology. As genomic tumor profiling is becoming more common, targeted treatments for specific molecular alterations are gaining traction. To discover new potential therapeutics that may apply to broad classes of tumors matching some molecular pattern, experimentalists and pharmacologists rely on high-throughput, in vitro screens of many compounds against many different cell lines. We propose a hierarchical Bayesian model of how cancer cell lines respond to drugs in these experiments and develop a method for fitting the model to real-world high-throughput screening data. Through a case study, the model is shown to capture nontrivial associations between molecular features and drug response, such as requiring both wild type TP53 and overexpression of MDM2 to be sensitive to Nutlin-3(a). In quantitative benchmarks, the model outperforms a standard approach in biology, with $\approx20\%$ lower predictive error on held out data. When combined with a conditional randomization testing procedure, the model discovers markers of therapeutic response that recapitulate known biology and suggest new avenues for investigation. All code for the article is publicly available at https://github.com/tansey/deep-dose-response.
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http://dx.doi.org/10.1093/biostatistics/kxaa047DOI Listing
January 2021

The UVSSA complex alleviates MYC-driven transcription stress.

J Cell Biol 2021 02;220(2)

Institute for Cancer Genetics, Columbia University Medical Center, New York, NY.

Cancer cells develop strong genetic dependencies, enabling survival under oncogenic stress. MYC is a key oncogene activated across most cancers, and identifying associated synthetic lethality or sickness can provide important clues about its activity and potential therapeutic strategies. On the basis of previously conducted genome-wide screenings in MCF10A cells expressing MYC fused to an estrogen receptor fragment, we identified UVSSA, a gene involved in transcription-coupled repair, whose knockdown or knockout decreased cell viability when combined with MYC expression. Synthetic sick interactions between MYC expression and UVSSA down-regulation correlated with ATM/CHK2 activation, suggesting increased genome instability. We show that the synthetic sick interaction is diminished by attenuating RNA polymerase II (RNAPII) activity; yet, it is independent of UV-induced damage repair, suggesting that UVSSA has a critical function in regulating RNAPII in the absence of exogenous DNA damage. Supporting this hypothesis, RNAPII ChIP-seq revealed that MYC-dependent increases in RNAPII promoter occupancy are reduced or abrogated by UVSSA knockdown, suggesting that UVSSA influences RNAPII dynamics during MYC-dependent transcription. Taken together, our data show that the UVSSA complex has a significant function in supporting MYC-dependent RNAPII dynamics and maintaining cell survival during MYC addiction. While the role of UVSSA in regulating RNAPII has been documented thus far only in the context of UV-induced DNA damage repair, we propose that its activity is also required to cope with transcriptional changes induced by oncogene activation.
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http://dx.doi.org/10.1083/jcb.201807163DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7791342PMC
February 2021

Genomic characterization of HIV-associated plasmablastic lymphoma identifies pervasive mutations in the JAK-STAT pathway.

Blood Cancer Discov 2020 Jul;1(1):112-125

Department of Haematology and Molecular Medicine, National Health Laboratory Service, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.

Plasmablastic lymphoma (PBL) is an aggressive B-cell non-Hodgkin lymphoma associated with immunodeficiency in the context of Human Immunodeficiency Virus (HIV) infection or iatrogenic immunosuppression. While a rare disease in general, the incidence is dramatically increased in regions of the world with high HIV prevalence. The molecular pathogenesis of this disease is poorly characterized. Here, we defined the genomic features of PBL in a cohort of 110 patients from South Africa (15 by whole exome sequencing and 95 by deep targeted sequencing). We identified recurrent mutations in genes of the JAK-STAT signaling pathway, including (42%), (14%) and (10%), leading to its constitutive activation. Moreover, 24% of cases harbored gain-of-function mutations in RAS family members ( and ). Comparative analysis with other B-cell malignancies uncovered PBL-specific somatic mutations and transcriptional programs. We also found recurrent copy number gains encompassing the CD44 gene (37%), which encodes for a cell surface receptor involved in lymphocyte activation and homing, and was found expressed at high levels in all tested cases, independent of genetic alterations. These findings have implications for the understanding of the pathogenesis of this disease and the development of personalized medicine approaches.
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http://dx.doi.org/10.1158/2643-3249.bcd-20-0051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7679070PMC
July 2020

A Random Matrix Theory Approach to Denoise Single-Cell Data.

Patterns (N Y) 2020 Jun 4;1(3):100035. Epub 2020 May 4.

Department of Systems Biology, Columbia University, New York NY 10032, USA.

Single-cell technologies provide the opportunity to identify new cellular states. However, a major obstacle to the identification of biological signals is noise in single-cell data. In addition, single-cell data are very sparse. We propose a new method based on random matrix theory to analyze and denoise single-cell sequencing data. The method uses the universal distributions predicted by random matrix theory for the eigenvalues and eigenvectors of random covariance/Wishart matrices to distinguish noise from signal. In addition, we explain how sparsity can cause spurious eigenvector localization, falsely identifying meaningful directions in the data. We show that roughly 95% of the information in single-cell data is compatible with the predictions of random matrix theory, about 3% is spurious signal induced by sparsity, and only the last 2% reflects true biological signal. We demonstrate the effectiveness of our approach by comparing with alternative techniques in a variety of examples with marked cell populations.
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http://dx.doi.org/10.1016/j.patter.2020.100035DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7660363PMC
June 2020

High tumor mutational burden and T-cell activation are associated with long-term response to anti-PD1 therapy in Lynch syndrome recurrent glioblastoma patient.

Cancer Immunol Immunother 2021 Mar 3;70(3):831-842. Epub 2020 Nov 3.

Unit of Molecular Neuro-Oncology, Fondazione IRCCS Istituto Neurologico Carlo Besta, Via Celoria 11, 20133, Milan, Italy.

Background: Glioblastomas (GBMs) in patients harboring somatic or germinal mutations of mismatch-repair (MMR) genes exhibit a hypermutable phenotype. Here, we describe a GBM patient with increased tumor mutational burden and germline MMR mutations, treated using anti-PD1 therapy.

Methods: A woman with newly diagnosed GBM (nGBM) was treated by surgery, radiotherapy, and temozolomide. The tumor recurred after 13 months leading to a second surgery and treatment with nivolumab. Whole-exome sequencing was performed on the nGBM, recurrent GBM (rGBM), and blood. Immune infiltration was investigated by immunohistochemistry and the immune response in the blood during treatment was analyzed by flow cytometry.

Results: High density of infiltrating CD163 + cells was found in both GBM specimens. Large numbers of CD3 + and CD8 + T cells were homogeneously distributed in the nGBM. The infiltration of CD4 + T cells and a different CD8 + T cell density were observed in the rGBM. Both GBM shared 12,431 somatic mutations, with 113 substitutions specific to the nGBM and 1,683 specific to the rGBM. Germline variants included pathogenic mutation in the MSH2 (R359S) gene, suggesting the diagnosis of Lynch syndrome. Systemic immunophenotyping revealed the generation of CD8 + T memory cells and persistent activation of CD4 + T cells. The patient is still receiving nivolumab 68 months after the second surgery.

Conclusions: Our observations indicate that the hypermutator phenotype associated with germinal mutations of MMR genes and abundant T-cell infiltration contributes to a durable clinical benefit sustained by a persistent and robust immune response during anti-PD1 therapy.
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http://dx.doi.org/10.1007/s00262-020-02769-4DOI Listing
March 2021

Midkine rewires the melanoma microenvironment toward a tolerogenic and immune-resistant state.

Nat Med 2020 12 19;26(12):1865-1877. Epub 2020 Oct 19.

Melanoma Laboratory, Molecular Oncology Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.

An open question in aggressive cancers such as melanoma is how malignant cells can shift the immune system to pro-tumorigenic functions. Here we identify midkine (MDK) as a melanoma-secreted driver of an inflamed, but immune evasive, microenvironment that defines poor patient prognosis and resistance to immune checkpoint blockade. Mechanistically, MDK was found to control the transcriptome of melanoma cells, allowing for coordinated activation of nuclear factor-κB and downregulation of interferon-associated pathways. The resulting MDK-modulated secretome educated macrophages towards tolerant phenotypes that promoted CD8 T cell dysfunction. In contrast, genetic targeting of MDK sensitized melanoma cells to anti-PD-1/anti-PD-L1 treatment. Emphasizing the translational relevance of these findings, the expression profile of MDK-depleted tumors was enriched in key indicators of a good response to immune checkpoint blockers in independent patient cohorts. Together, these data reveal that MDK acts as an internal modulator of autocrine and paracrine signals that maintain immune suppression in aggressive melanomas.
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http://dx.doi.org/10.1038/s41591-020-1073-3DOI Listing
December 2020

Mutant SF3B1 promotes AKT- and NF-κB-driven mammary tumorigenesis.

J Clin Invest 2021 01;131(1)

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

Mutations in the core RNA splicing factor SF3B1 are prevalent in leukemias and uveal melanoma, but hotspot SF3B1 mutations are also seen in epithelial malignancies such as breast cancer. Although hotspot mutations in SF3B1 alter hematopoietic differentiation, whether SF3B1 mutations contribute to epithelial cancer development and progression is unknown. Here, we identify that SF3B1 mutations in mammary epithelial and breast cancer cells induce a recurrent pattern of aberrant splicing leading to activation of AKT and NF-κB, enhanced cell migration, and accelerated tumorigenesis. Transcriptomic analysis of human cancer specimens, MMTV-cre Sf3b1K700E/WT mice, and isogenic mutant cell lines identified hundreds of aberrant 3' splice sites (3'ss) induced by mutant SF3B1. Consistently between mouse and human tumors, mutant SF3B1 promoted aberrant splicing (dependent on aberrant branchpoints as well as pyrimidines downstream of the cryptic 3'ss) and consequent suppression of PPP2R5A and MAP3K7, critical negative regulators of AKT and NF-κB. Coordinate activation of NF-κB and AKT signaling was observed in the knockin models, leading to accelerated cell migration and tumor development in combination with mutant PIK3CA but also hypersensitizing cells to AKT kinase inhibitors. These data identify hotspot mutations in SF3B1 as an important contributor to breast tumorigenesis and reveal unique vulnerabilities in cancers harboring them.
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http://dx.doi.org/10.1172/JCI138315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7773370PMC
January 2021

CtIP-mediated DNA resection is dispensable for IgH class switch recombination by alternative end-joining.

Proc Natl Acad Sci U S A 2020 10 28;117(41):25700-25711. Epub 2020 Sep 28.

Institute for Cancer Genetics, Vagelos College for Physicians and Surgeons, Columbia University, New York, NY 10032;

To generate antibodies with different effector functions, B cells undergo Immunoglobulin Heavy Chain (IgH) class switch recombination (CSR). The ligation step of CSR is usually mediated by the classical nonhomologous end-joining (cNHEJ) pathway. In cNHEJ-deficient cells, a remarkable ∼25% of CSR can be achieved by the alternative end-joining (Alt-EJ) pathway that preferentially uses microhomology (MH) at the junctions. While A-EJ-mediated repair of endonuclease-generated breaks requires DNA end resection, we show that CtIP-mediated DNA end resection is dispensable for A-EJ-mediated CSR using cNHEJ-deficient B cells. High-throughput sequencing analyses revealed that loss of ATM/ATR phosphorylation of CtIP at T855 or ATM kinase inhibition suppresses resection without altering the MH pattern of the A-EJ-mediated switch junctions. Moreover, we found that ATM kinase promotes Alt-EJ-mediated CSR by suppressing interchromosomal translocations independent of end resection. Finally, temporal analyses reveal that MHs are enriched in early internal deletions even in cNHEJ-proficient B cells. Thus, we propose that repetitive IgH switch regions represent favored substrates for MH-mediated end-joining contributing to the robustness and resection independence of A-EJ-mediated CSR.
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http://dx.doi.org/10.1073/pnas.2010972117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7568320PMC
October 2020

A single-cell atlas of the mouse and human prostate reveals heterogeneity and conservation of epithelial progenitors.

Elife 2020 09 11;9. Epub 2020 Sep 11.

Department of Medicine, Columbia University Irving Medical Center, New York, United States.

Understanding the cellular constituents of the prostate is essential for identifying the cell of origin for prostate adenocarcinoma. Here, we describe a comprehensive single-cell atlas of the adult mouse prostate epithelium, which displays extensive heterogeneity. We observe distal lobe-specific luminal epithelial populations (LumA, LumD, LumL, and LumV), a proximally enriched luminal population (LumP) that is not lobe-specific, and a periurethral population (PrU) that shares both basal and luminal features. Functional analyses suggest that LumP and PrU cells have multipotent progenitor activity in organoid formation and tissue reconstitution assays. Furthermore, we show that mouse distal and proximal luminal cells are most similar to human acinar and ductal populations, that a PrU-like population is conserved between species, and that the mouse lateral prostate is most similar to the human peripheral zone. Our findings elucidate new prostate epithelial progenitors, and help resolve long-standing questions about anatomical relationships between the mouse and human prostate.
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http://dx.doi.org/10.7554/eLife.59465DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7529463PMC
September 2020

MGMT genomic rearrangements contribute to chemotherapy resistance in gliomas.

Nat Commun 2020 08 4;11(1):3883. Epub 2020 Aug 4.

Seve Ballesteros Foundation Brain Tumor Group, Molecular Oncology Programme, Spanish National Cancer Research Center, CNIO, 28029, Madrid, Spain.

Temozolomide (TMZ) is an oral alkylating agent used for the treatment of glioblastoma and is now becoming a chemotherapeutic option in patients diagnosed with high-risk low-grade gliomas. The O-6-methylguanine-DNA methyltransferase (MGMT) is responsible for the direct repair of the main TMZ-induced toxic DNA adduct, the O6-Methylguanine lesion. MGMT promoter hypermethylation is currently the only known biomarker for TMZ response in glioblastoma patients. Here we show that a subset of recurrent gliomas carries MGMT genomic rearrangements that lead to MGMT overexpression, independently from changes in its promoter methylation. By leveraging the CRISPR/Cas9 technology we generated some of these MGMT rearrangements in glioma cells and demonstrated that the MGMT genomic rearrangements contribute to TMZ resistance both in vitro and in vivo. Lastly, we showed that such fusions can be detected in tumor-derived exosomes and could potentially represent an early detection marker of tumor recurrence in a subset of patients treated with TMZ.
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http://dx.doi.org/10.1038/s41467-020-17717-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7403430PMC
August 2020

Identification of relevant genetic alterations in cancer using topological data analysis.

Nat Commun 2020 07 30;11(1):3808. Epub 2020 Jul 30.

Department of Genetics and Institute for Biomedical Informatics, Perelman School of Medicine, University of Pennsylvania, 3400 Civic Center Blvd., Philadelphia, PA, 19104, USA.

Large-scale cancer genomic studies enable the systematic identification of mutations that lead to the genesis and progression of tumors, uncovering the underlying molecular mechanisms and potential therapies. While some such mutations are recurrently found in many tumors, many others exist solely within a few samples, precluding detection by conventional recurrence-based statistical approaches. Integrated analysis of somatic mutations and RNA expression data across 12 tumor types reveals that mutations of cancer genes are usually accompanied by substantial changes in expression. We use topological data analysis to leverage this observation and uncover 38 elusive candidate cancer-associated genes, including inactivating mutations of the metalloproteinase ADAMTS12 in lung adenocarcinoma. We show that ADAMTS12 mice have a five-fold increase in the susceptibility to develop lung tumors, confirming the role of ADAMTS12 as a tumor suppressor gene. Our results demonstrate that data integration through topological techniques can increase our ability to identify previously unreported cancer-related alterations.
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http://dx.doi.org/10.1038/s41467-020-17659-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7393176PMC
July 2020
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