Biology subjects

Naqvi, A. S.

Publications and source records attributed to Naqvi, A. S..

7 recordsLinked to original sources

bioRxiv2025

Neuronal cell adhesion molecule (NRCAM) variant defined by microexon skipping is an essential, antigenically distinct, and targetable surface proteoform in high-grade glioma

To overcome the paucity of known tumor-specific surface antigens in pediatric high-grade glioma (pHGG), we contrasted splicing patterns in pHGGs and normal brain samples. Among alternative splicing events affecting extracellular protein domains, the most pervasive alteration was the skipping of [≤]30 nucleotide-long exons. Several of these skipped microexons mapped to L1-IgCAM family members, such as NRCAM. Bulk and single-nuclei short- and long-read RNA-seq revealed uniform skipping of NRCAM microexons 5 and 19 in virtually every pHGG sample. Importantly, the {Delta}ex5{Delta}ex19 (but not the full-length) NRCAM proteoform was essential for pHGG cell migration and invasion in vitro and tumor growth in vivo. We developed a monoclonal antibody selective for {Delta}ex5{Delta}ex19 NRCAM and demonstrated that "painting" of pHGG cells with this antibody enables killing by T cells armed with an FcRI-based universal immune receptor. Thus, pHGG-specific NRCAM and possibly other L1-IgCAM proteoforms are promising and highly selective targets for adoptive immunotherapies.

cancer biology↗

bioRxiv2024

The splicing modulator CLK1 is a candidate oncogenic dependency in pediatric high-grade gliomas

BackgroundPediatric brain tumors are the leading cause of disease-related mortality in children, yet many aggressive tumors lack effective therapies. RNA splicing is a hallmark of cancer, but it has not yet been systematically studied in pediatric brain tumors. MethodsWe analyzed 729 pediatric brain tumors spanning histologies and molecular subtypes to quantify differential tumor splicing. We developed the Splicing Burden Index (SBI) to enable cross-sample comparisons and performed hierarchical clustering of highly variable splice events to define splicing-informed tumor groups. These were integrated with clinical outcomes, pathway activity, and proteogenomic data. Recurrent splice events were prioritized for predicted functional impact, and in vitro perturbation studies were performed targeting the splicing kinase CDC-like kinase 1 (CLK1). ResultsSBI revealed substantial inter- and intra-histology heterogeneity. Clusters were enriched for histologies and molecular subtypes, several of which were independently associated with survival beyond histology and clinical covariates. Spliceosome pathway activity varied across clusters and was associated with worse survival, yet was not correlated with SBI, indicating distinct dimensions of splicing dysregulation. Functional prioritization identified a recurrent in CLK1 exon 4, required for canonical kinase activity. CLK1 exon 4 inclusion followed an oncofetal pattern and showed context-dependent associations with outcome distinct from total CLK1 expression. Pharmacologic inhibition and exon 4-specific perturbation of CLK1 reduced tumor cell viability and disrupted cancer-relevant splicing and transcriptional programs. ConclusionsThis study systematically characterizes splicing in pediatric brain tumors, identifies splicing-informed subgroups, and prioritizes CLK1 exon 4 as an oncofetal tumor-specific event, motivating further preclinical exploration. Key Points[bullet] Splicing analysis of 729 pediatric CNS tumors identifies splicing-defined clusters. [bullet]CLK1 exon 4 inclusion is widespread and developmentally regulated. [bullet]Exon-level CLK1 regulation shows context-dependent links to prognosis in aggressive CNS tumors.

genomics↗

bioRxiv2024

The Open Pediatric Cancer Project

BackgroundIn 2019, the Open Pediatric Brain Tumor Atlas (OpenPBTA) was created as a global, collaborative open-science initiative to genomically characterize 1,074 pediatric brain tumors and 22 patient-derived cell lines. Here, we present an extension of the OpenPBTA called the Open Pediatric Cancer (OpenPedCan) Project, a harmonized open-source multi-omic dataset from 6,112 pediatric cancer patients with 7,096 tumor events across more than 100 histologies. Combined with RNA-Seq from the Genotype-Tissue Expression (GTEx) and The Cancer Genome Atlas (TCGA), OpenPedCan contains nearly 48,000 total biospecimens (24,002 tumor and 23,893 normal specimens). FindingsWe utilized Gabriella Miller Kids First (GMKF) workflows to harmonize WGS, WXS, RNA-seq, and Targeted Sequencing datasets to include somatic SNVs, InDels, CNVs, SVs, RNA expression, fusions, and splice variants. We integrated summarized CPTAC whole cell proteomics and phospho-proteomics data, miRNA-Seq data, and have developed a methylation array harmonization workflow to include m-values, beta-vales, and copy number calls. OpenPedCan contains reproducible, dockerized workflows in GitHub, CAVATICA, and Amazon Web Services (AWS) to deliver harmonized and processed data from over 60 scalable modules which can be leveraged both locally and on AWS. The processed data are released in a versioned manner and accessible through CAVATICA or AWS S3 download (from GitHub), and queryable through PedcBioPortal and the NCIs pediatric Molecular Targets Platform. Notably, we have expanded PBTA molecular subtyping to include methylation information to align with the WHO 2021 Central Nervous System Tumor classifications, allowing us to create research-grade integrated diagnoses for these tumors. ConclusionsOpenPedCan data and its reproducible analysis module framework are openly available and can be utilized and/or adapted by researchers to accelerate discovery, validation, and clinical translation.

genomics↗

bioRxiv2023

AutoGVP: a dockerized workflow integrating ClinVar and InterVar germline sequence variant classification

SummaryWith the increasing rates of exome and whole genome sequencing, the ability to classify large sets of germline sequencing variants using up-to-date American College of Medical Genetics - Association for Molecular Pathology (ACMG-AMP) criteria is crucial. Here, we present Automated Germline Variant Pathogenicity (AutoGVP), a tool that integrates germline variant pathogenicity annotations from ClinVar and sequence variant classifications from a modified version of InterVar (PVS1 strength adjustments, removal of PP5/BP6). This tool facilitates large-scale, clinically-focused classification of germline sequence variants in a research setting. Availability and ImplementationAutoGVP is an open-source dockerized workflow implemented in R and freely available on GitHub at https://github.com/diskin-lab-chop/AutoGVP.

bioinformatics↗

bioRxiv2022

FBXW7β isoform drives transcriptional activation of a proinflammatory TNF cluster in normal and malignant pro-B cells

Non-canonical exon usage plays many important roles in cellular phenotypes, but its contribution to human B-cell development remains sketchily understood. To fill this gap, we collected various B-cell fractions from bone marrow and tonsil donors, performed RNA-seq, and examined transcript variants. We identified 150 genes that harbor local splicing variations in all pairwise comparisons. One of them encodes FBXW7, an E3 ubiquitin ligase implicated as a cancer driver in several blood cancers. Surprisingly, we discovered that in normal human pro-B cells, the predominant transcript utilized an alternative first exon to produce the poorly characterized FBXW7{beta} isoform, previously thought to be restricted to neural tissues. The FBXW7{beta} transcript was also abundant in cell lines and primary samples of pediatric B-cell acute lymphoblastic leukemia (B-ALL), which originates in the bone marrow. When overexpressed in a heterologous cell system, this transcript yielded the expected protein product, as judged by anti-FLAG immunoblotting and mass spectrometry. Furthermore, in REH B-ALL cells, FBXW7{beta} mRNA was the only FBXW7 isoform enriched in the polyribosome fraction. To shed light on possible functions of FBXW7{beta}, we utilized gain- and loss-of-function approaches and identified an FBXW7{beta}-dependent inflammatory gene signature, apparent in a subset of B-ALL with high FBXW7{beta} expression. This signature contained several members of the TNF superfamily, including those comprising the HLA Class III cluster (LTB, LST1, NCR3, LTA, and NFKBIL1). Our findings suggest that FBXW7{beta} expression drives proinflammatory responses, which could contribute to normal B-cell development, leukemogenesis and responses to anti-cancer therapies. Key pointsO_LIPreviously thought to be restricted to neural tissues, FBXW7{beta} is the predominant FBXW7 isoform in normal and malignant human pro-B cells. C_LIO_LIFBXW7{beta} promotes transcriptional activation of a proinflammatory gene cluster that contains TNF superfamily members. C_LI

cell biology↗

bioRxiv2019

The role of SRSF3 splicing factor in generating circular RNAs

Circular RNAs (circRNAs) represent a novel class of non-coding RNAs that are emerging as potentially important regulators of gene expression. circRNAs are typically generated from host gene transcripts through a non-canonical back-splicing mechanism, whose regulation is still not well understood. To explore regulation of circRNAs in cancer, we generated sequence data from RNase R-resistant transcripts in human p493-6 B-lymphoid cells and identified thousands of novel as well as previously identified circRNAs. Approximately 40% of expressed genes generated a circRNA, with half of them generating multiple isoforms, suggesting the involvement of alternative back-splicing and regulatory RNA-binding proteins (RBPs). We observed that genes generating circRNAs with back-spliced exonic junctions were enriched for RBP recognition motifs, including multiple splicing factors, most notably SRSF3, a splicing factor known to promote exon inclusion. To test the role of SRSF3 role in circRNA production, we performed traditional RNA-seq in p493-6 B-lymphoid cells with and without SRSF3 knockdown, and identified 926 mRNA transcripts, whose canonical splicing was affected by SRSF3. We found that a subset (205) of these SRSF3 targets served as host transcripts for circRNA, suggesting that SRSF3 may regulate exon circularization. Since this splicing factor is deregulated in hematologic malignancies, we hypothesize that SRSF3-dependent circRNAs, similar to their mRNA counterparts, might contribute to the pathogenesis of lymphomas and leukemias.

molecular biology↗

bioRxiv2017

Aberrant splicing in B-cell acute lymphoblastic leukemia

Aberrant splicing is a hallmark of leukemias with mutations in splicing factor (SF)-encoding genes. Here we investigated its prevalence in pediatric B-cell acute lymphoblastic leukemias (B-ALL), where SFs are not mutated. By comparing them to normal pro-B cells, we found thousands of aberrant local splice variations (LSVs) per sample, with 279 LSVs in 241 genes present in every comparison. These genes were enriched in RNA processing pathways and encoded ~100 SFs, e.g. hnRNPA1. hnRNPA1 3UTR was pervasively misspliced, yielding the transcript subject to nonsense-mediated decay. Thus, we knocked it down in B-lymphoblastoid cells, identified 213 hnRNPA1-dependent splicing events, and defined the hnRNPA1 splicing signature in pediatric leukemias. One of its elements was DICER1, a known tumor suppressor gene; its LSVs were consistent with reduced translation of DICER1 mRNA. Additionally, we searched for LSVs in other leukemia and lymphoma drivers and discovered 81 LSVs in 41 genes. 77 LSVs were confirmed using two large independent B-ALL RNA-seq datasets. In fact, the twenty most common B-ALL drivers showed higher prevalence of aberrant splicing than of somatic mutations. Thus, post-transcriptional deregulation of SF can drive widespread changes in B-ALL splicing and likely contribute to disease pathogenesis.

cancer biology↗