Biology subjects

Bhinder, B.

Publications and source records attributed to Bhinder, B..

14 recordsLinked to original sources

bioRxiv2025

APOBEC3A-Induced DNA Damage Drives Polymerase Theta Dependency and Synthetic Lethality in Cancer

APOBEC3 cytidine deaminases drive cancer evolution. There is an unmet need to target cancer cells with APOBEC3 activity. Here, we identify error-prone theta-mediated end joining (TMEJ) as the main pathway for repairing APOBEC3-induced double-strand breaks (DSBs). Using fluorescent DSB repair reporters and a novel biochemical assay, we demonstrate that APOBEC3A competes with replication protein A (RPA) for single-stranded DNA overhangs, exposing microhomologous sequences to shift DSB repair towards error-prone TMEJ. Genomic analysis of clinical tumor samples confirmed the co-occurrence and proximity between APOBEC3-induced mutational footprints, microhomology-mediated deletions (MMDs), and TMEJ-associated chromosomal instability signatures. Crucially, inhibition of DNA polymerase theta (Pol{theta}) synergizes with APOBEC3A-induced DSBs to induce synthetic lethality in vitro and in vivo. Collectively, our findings identify TMEJ as the preferred mechanism for repairing APOBEC3A-induced DSBs and establish Pol{theta} inhibition as a novel promising strategy to eliminate cancer cells with APOBEC3A activity.

cancer biology↗

bioRxiv2025

Transcriptome sequencing of Hodgkin lymphoma Hodgkin and Reed-Stenberg cells reveals escape from NK cell recognition and an unfolded protein response

The mutational profile of classic Hodgkin lymphoma (cHL) overlaps with that of related B cell lymphomas, including primary mediastinal B cell lymphoma (PMBL), and yet these are different histologically and clinically. To discover the molecular features that distinguish cHL, we deployed flow cytometric cell sorting and low-input RNA sequencing to generate full transcriptome data from viable, isolated Hodgkin and Red-Sternberg (HRS) cells from eighteen primary tumors, alongside matched intra-tumoral non-neoplastic B cells and four cell lines. Comparison of HRS cells to normal cellular subsets revealed evidence of abortive plasma cell differentiation, with an unfolded protein response signature, shared with plasma cell neoplasms, but not other B-cell lymphoma types. Comparison of cHL to PMBL revealed similarities but also key differences in B cell differentiation programs accompanied by upregulation of genes involved in microtubule cytoskeleton organization in cHL, which may be related to the unique multinucleated nature of HRS cells. In HRS cells, we also observed a downregulation of SLAM family receptors, which are crucial for NK cell activation, providing a potential mechanism for immune evasion from NK-mediated killing. STATEMENT OF SIGNIFICANCEThis study defines a unique transcriptional program in classic Hodgkin Lymphoma (cHL) marked by oncogenic signaling, chromatin integrity, DNA repair, and immune escape including loss of NK-activating receptors. HRS cells resemble plasma cells and have an unfolded protein response signature, which distinguishes them from diffuse large and primary mediastinal B cell lymphomas.

cancer biology↗

bioRxiv2025

Pan-Cancer PDOs Preserve Tumor Heterogeneity and Uncover Therapeutic Vulnerabilities

We developed a tumor-matched, pan-cancer patient-derived organoid (PDO) platform comprising 220 PDOs from 190 patients across 15 cancer types to advance functional precision oncology. Our comprehensively characterized PDOs showed 93% histopathology concordance, 80% median genomic concordance for driver mutations, and a 0.85 median gene expression correlation with parent tumors. Gene expression in PDOs remained stable across [≥] 10 passages, supporting reproducibility for long-term drug screening. Even PDOs with low genomic concordance retained oncogenic drivers, supporting their use as disease models. Clonality analysis revealed that 85% of PDOs preserved dominant tumor clones. Higher genomic concordance was associated with greater clonal similarity, while lower genomic concordance was associated with clonal divergence. Functional assays showed that 58% of PDOs from a subset of patients ineligible for FDA-approved PARP inhibitors responded to Talazoparib, with sensitivity linked to alterations in DNA damage repair. Combination screens revealed drugs that effectively overcame resistance, especially in TP53-mutant PDOs. In summary, our platform supports investigation of targeted therapies, identification of molecular features linked to drug sensitivity, and translational discovery, offering insights into personalized cancer treatment beyond current biomarker guidelines.

cancer biology↗

bioRxiv2025

Simultaneous immunomodulation and epithelial-to-mesenchymal transition drives lung adenocarcinoma progression

Lung cancer remains the deadliest cancer in the United States, with lung adenocarcinoma (LUAD) as its most prevalent subtype. While computed tomography (CT)-based screening has improved early detection and enabled curative surgeries, the molecular and cellular dynamics driving early-stage LUAD progression remain poorly understood, limiting non-surgical treatment options. To address this gap, we profiled 2.24 million cells from 122 early-stage LUAD patients using multiplexed imaging mass cytometry (IMC). This analysis revealed the molecular, spatial, and temporal dynamics of LUAD development. Our findings uncover a binary progression model. LUAD advances through either inflammation, driven by a balance of cytotoxic and regulatory immune activity, or fibrosis, characterized by stromal activation. Surprisingly, tumor cell populations did not increase significantly. Instead, they displayed a mixed phenotypic profile consistent with epithelial-to-mesenchymal transition (EMT), effectively masking the expansion of malignant cells. Furthermore, we addressed discrepancies between CT-based and histology-based subtyping. CT scans, while non-invasive, often mischaracterize invasive fibrotic tumors--which account for 20.5% of LUAD cases--as mild, non-solid ground glass opacities (GGOs). Using high-content IMC imaging, we demonstrate that these tumors harbor significant risks and advocate for improved diagnostic strategies. These strategies should integrate molecular profiling to refine patient stratification and therapeutic decision-making. Altogether, our study provides a high-resolution, systems-level view of the tumor microenvironment in early-stage LUAD. We characterize key transitions in oncogenesis and propose a precision-driven framework to enhance the detection and management of aggressive disease subtypes.

cancer biology↗

bioRxiv2025

Acquisition of discrete immune suppressive barriers contributes to the initiation and progression of preinvasive to invasive human lung cancer.

Computerized chest tomography (CT)-guided screening in populations at risk for lung cancer has increased the detection of preinvasive subsolid nodules, which progress to solid invasive adenocarcinoma. Despite the clinical significance, there is a lack of effective therapies for intercepting the progression of preinvasive to invasive adenocarcinoma. To uncover determinants of early disease emergence and progression, we used integrated single-cell approaches, including scRNA-seq, multiplexed imaging mass cytometry and spatial transcriptomics, to construct the first high-resolution map of the composition, lineage/functional states, developmental trajectories and multicellular crosstalk networks from microdissected non-solid (preinvasive) and solid compartments (invasive) of individual part-solid nodules. We found that early disease initiation and subsequent progression are associated with the evolution of immune-suppressive cellular phenotypes characterized by decreased cytotoxic CD8 T and NK cells, increased T cell exhaustion and accumulation of immunosuppressive regulatory T cells (Tregs) and M2-like macrophages expressing TREM2. Within Tregs, we identified a unique population of 4-1BB+ Treg subset enriched for the IL2-STAT5 suppressive pathway with transcription profiles supporting discrete metabolic alterations. Spatial analysis showed increased density of suppressive immune cells around tumor cells, increased exhaustion phenotype of both CD4 and CD8 T cells expressing chemokine CXCL13, and spatial micro-complex of endothelial and lymphocyte interactions within tertiary lymphoid structures. The single-cell architecture identifies determinants of early disease emergence and progression, which may be developed not only as diagnostic/prognostic biomarkers but also as targets for disease interception. Additionally, our dataset constitutes a valuable resource for the preinvasive lung cancer research community.

cancer biology↗

bioRxiv2024

The Orphan G Protein-Coupled Receptor GPR52 is a Novel Regulator of Breast Cancer Multicellular Organization

Statement of SignificanceWe showed that loss of the orphan G protein-coupled receptor GPR52 in human breast cell lines leads to increased cell clustering, hybrid/partial EMT, and increased tumor burden in zebrafish. BackgroundG protein-coupled receptors (GPCRs) are the largest class of membrane-bound receptors that transmit critical signals from extracellular to intracellular spaces. Transcriptomic data of resected breast tumors show that low mRNA expression of orphan GPCR GPR52 correlates with reduced overall survival in patients with breast cancer, leading to the hypothesis that loss of GPR52 supports breast cancer progression. MethodsCRISPR-Cas9 was used to knockout GPR52 in the human triple-negative breast cancer (TNBC) cell lines MDA-MB-468 and MDA-MB-231, and in the non-cancerous breast epithelial cell line MCF10A. 2D and 3D in vitro studies, electron microscopy, Matrigel culture, and a zebrafish xenograft model were used to assess the morphology and behavior of GPR52 KO cells. RNA-sequencing and proteomic analyses were also conducted on these cell lines, and transcriptomic data from The Cancer Genome Atlas (TCGA) database were used to compare GPR52-null and wild-type (WT) signatures in breast cancer. ResultsLoss of GPR52 was found to be associated with increased cell-cell interaction in 2D cultures, altered 3D spheroid morphology, and increased propensity to organize and invade collectively in Matrigel. Furthermore, GPR52 loss was associated with features of EMT in MDA-MB-468 cells, and zebrafish injected with GPR52 KO cells developed a greater total cancer area than those injected with control cells. RNA sequencing and proteomic analyses of GPR52-null breast cancer cells revealed an increased cAMP signaling signature. Consistently, we found that treatment of wild-type (WT) cells with forskolin, which stimulates the production of cAMP, induces phenotypic changes associated with GPR52 loss, and inhibition of cAMP production rescued some GPR52 KO phenotypes. ConclusionGPR52 is an orphan GPCR and its role in cancer progression has not been previously characterized. We found that GPR52 loss in breast cancer cells can lead to increased cell clustering, collective invasion, and EMT in vitro. These are features of increased cancer aggression. Our results reveal that GPR52 loss is a potential mechanism by which breast cancer progression may occur and support the investigation of GPR52 agonism as a therapeutic option for breast cancer. O_FIG O_LINKSMALLFIG WIDTH=169 HEIGHT=200 SRC="FIGDIR/small/604482v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@8b0f55org.highwire.dtl.DTLVardef@19bd139org.highwire.dtl.DTLVardef@c52117org.highwire.dtl.DTLVardef@1ffa2bd_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO C_FIG

cancer biology↗

bioRxiv2023

Novel co-culture strategies of tumor organoids with autologous T-cells reveal clinically relevant combinations of immune-checkpoint and targeted therapies

Patient derived tumor organoids (PDTOs) have become relevant pre-clinical models for therapeutic modelling since they highly recapitulate patients response to treatment. Nevertheless, their value for immunotherapy modelling has not been fully explored. We developed a tumor processing protocol that enable the establishment of PDTOs and tumor infiltrating lymphocytes (TILs) isolation. By the optimization of functional assays, we compared the T-cells effector functions of matching PBMCs and TILs, demonstrating that PBMCs after co-culture and TILs after initial expansion display similar responses. In addition, the evaluation of cytokine production by fluorospot in combination with an image-based killing assay enable the screening of different immune-checkpoint inhibitors as well as its combination with target inhibitors. Our proof-of-concept functional assays showed the potential and versatility of PDTOs and T-cells co-culture systems for immunotherapy screening. The optimization of scalable functional assays downstream co-culture represents a significant step forward to increase the value of PDTOs as pre-clinical models for immunotherapeutic screens.

immunology↗

bioRxiv2022

The cytidine deaminase APOBEC3G drives cancer mutagenesis and clonal evolution in bladder cancer

Mutagenic processes leave distinct signatures in cancer genomes. The mutational signatures attributed to APOBEC3 cytidine deaminases are pervasive in human cancers. However, data linking individual APOBEC3 proteins to cancer mutagenesis in vivo are limited. Here, we show that transgenic expression of human APOBEC3G promotes mutagenesis, genomic instability, and kataegis, leading to shorter survival in a murine bladder cancer model. Acting as mutagenic fuel, APOBEC3G increases the clonal diversity of bladder cancers, driving divergent cancer evolution. We characterize the single base substitution signature induced by APOBEC3G in vivo, showing the induction of a mutational signature different from that caused by APOBEC3A and APOBEC3B. Analysis of thousands of human cancers reveals the contribution of APOBEC3G to the mutational profiles of multiple cancer types, including bladder cancer. Our findings define the role of APOBEC3G in cancer mutagenesis and clonal heterogeneity. These results potentially inform future therapeutic efforts that restrict tumor evolution.

genomics↗

bioRxiv2022

An RNAi screen reveals common host-virus gene signatures: implication for anti-viral drug discovery

Dengue is the most common mosquito-borne viral disease that in recent years has become a major international public health concern. Dengue is a tropical neglected disease with increasing global incidences, affecting millions of people worldwide, and without the availability of specific treatments to combat it. The identification of host-target genes essential for the virus life cycle, for which effective modulators may already exist, would provide an alternative path to a rapid drug development of the much needed anti-Dengue agents. For this purpose, we performed the first genome-wide RNAi screen, combining two high content readouts for DENV infection (FLUO) and host cell toxicity (NUCL), against an arrayed lentiviral based shRNA library covering 16,000 genes with a redundancy of at least 5 hairpins per gene. The screen identified 1,924 gene candidates in total; of which, 1,730 gene candidates abrogated Dengue infection, while 194 gene candidates were found to enhance its infectivity in HEK293 cells. A first pass clustering analysis of hits revealed a well orchestrated gene-network dependency on host cell homeostasis and physiology triggering distinct cellular pathways for infectivity, replication, trafficking and egress; a second analysis revealed a comprehensive gene signature of 331 genes common to hits identified in 28 published RNAi host-viral interactions screens. Taken together, our findings provide novel antiviral molecular targets with the potential for drug discovery and development.

systems biology↗

bioRxiv2021

The evolution of genomic, transcriptomic, and single-cell protein markers of metastatic upper tract urothelial carcinoma

The molecular characteristics of metastatic upper tract urothelial carcinoma (UTUC) are unknown. The genomic and transcriptomic differences between primary and metastatic UTUC is not well described either. We combined whole-exome sequencing, RNA-sequencing, and Imaging Mass Cytometry (IMC) of 44 tumor samples from 28 patients with high-grade primary and metastatic UTUC. IMC enables spatially resolved single-cell analyses to examine the evolution of cancer cell, immune cell, and stromal cell markers using mass cytometry with lanthanide metal-conjugated antibodies. We discovered that actionable genomic alterations are frequently discordant between primary and metastatic UTUC tumors in the same patient. In contrast, molecular subtype membership and immune depletion signature were stable across primary and matched metastatic UTUC. Molecular and immune subtypes were consistent between bulk RNA-sequencing and mass cytometry of protein markers from 340,798 single-cells. Molecular subtyping at the single cell level was highly conserved between primary and metastatic UTUC tumors within the same patient.

cancer biology↗

bioRxiv2021

Molecular evolution of classic Hodgkin lymphoma revealed through whole genome sequencing of Hodgkin and Reed Sternberg cells

The rarity of malignant Hodgkin and Reed Sternberg (HRS) cells within a classic Hodgkin lymphoma (cHL) biopsy limits the ability to study the genomics of cHL. To circumvent this, our group has previously optimized fluorescence-activated cell sorting to purify HRS cells. Here we leveraged this method to report the first whole genome sequencing landscape of HRS cells and reconstruct the chronology and likely etiology of pathogenic events prior to the clinical diagnosis of cHL. We identified alterations in driver genes not previously described in cHL, a high activity of the APOBEC mutational signature, and the presence complex structural variants including chromothripsis. We found that the high ploidy observed in cHL is often acquired through multiple, independent large chromosomal gain events including whole genome duplication. The first of these likely occurs several years prior to the diagnosis of cHL, and the last gains typically occur very close to the time of diagnosis. Evolutionary timing analyses revealed that driver mutations in B2M, BCL7A, GNA13, and PTPN1, and the onset of AID driven mutagenesis usually preceded large chromosomal gains. The study provides the first temporal reconstruction of cHL pathogenesis and suggests a relatively long time course between the first pathogenic event and the clinical diagnosis.

cancer biology↗

bioRxiv2021

Cysteine induces mitochondrial reductive stress in glioblastoma through hydrogen peroxide production

Glucose and amino acid metabolism are critical for glioblastoma (GBM) growth, but little is known about the specific metabolic alterations in GBM that are targetable with FDA-approved compounds. To investigate tumor metabolism signatures unique to GBM, we interrogated The Cancer Genome Atlas for alterations in glucose and amino acid signatures in GBM relative to other human cancers and found that GBM exhibits the highest levels of cysteine and methionine pathway gene expression of 32 human cancers. Treatment of patient-derived GBM cells with the FDA-approved cysteine compound N-acetylcysteine (NAC) reduce GBM cell growth and mitochondrial oxygen consumption, which was worsened by glucose starvation. Mechanistic experiments revealed that cysteine compounds induce rapid mitochondrial H2O2 production and reductive stress in GBM cells, an effect blocked by oxidized glutathione, thioredoxin, and redox enzyme overexpression. These findings indicate that GBM is uniquely susceptible to NAC-driven reductive stress and could synergize with glucose-lowering treatments for GBM.

cancer biology↗

bioRxiv2019

Age, Sex, and Genetics Influence the Abundance of Infiltrating Immune Cells in Human Tissues

Despite infiltrating immune cells playing an essential role in human disease and the patient response to treatment, the central mechanisms influencing variability in infiltration patterns are unclear. Using bulk RNA-seq data from 53 GTEx tissues, we applied cell-type deconvolution algorithms to evaluate the immune landscape across the healthy human body. We first performed a differential expression analysis of inflamed versus non-inflamed samples to identify essential pathways and regulators of infiltration. Next, we found 21 of 73 infiltration-related phenotypes to be associated with either age or sex (FDR < 0.1). Through our genetic analysis, we discovered 13 infiltration-related phenotypes have genome-wide significant associations (iQTLs) (P < 5.0 x 10-8), with a significant enrichment of tissue-specific expression quantitative trait loci in suggested iQTLs (P < 10-5). We highlight an association between neutrophil content in lung tissue and a variant near the CUX1 transcription factor gene (P = 9.7 x 10-11), which has been previously linked to neutrophil infiltration, inflammatory mechanisms, and the regulation of several immune response genes. Together, our results identify key factors influencing inter-individual variability of specific tissue infiltration patterns, which could provide insights on therapeutic targets for shifting infiltration profiles to a more favorable one.

genetics↗

bioRxiv2018

Predicting peptide presentation by major histocompatibility complex class I using one million peptides

Improved computational tools are needed to prioritize putative neoantigens within immunotherapy pipelines for cancer treatment. Herein, we assemble a database of over one million human peptides presented by major histocompatibility complex class I (MHC-I), the largest known database of its type. We use these data to train a random forest classifier (ForestMHC) to predict likelihood of MHC-I presentation. The information content of features mirrors the canonical importance of positions two and nine in determining likelihood of binding. Our random forest-based method outperforms NetMHC and NetMHCpan on test sets, and it outperforms both these methods and MixMHCpred on new mass spectrometry data from an ovarian carcinoma sample. Furthermore, the random forest scores correlate monotonically with peptide binding affinities, when known. Finally, we examine the effect size of gene expression on peptide presentation and find a moderately strong relationship. The ForestMHC method is a promising modality to prioritize neoantigens for experimental testing in immunotherapy.

bioinformatics↗