Biology subjects

Sarhan, D.

Publications and source records attributed to Sarhan, D..

8 recordsLinked to original sources

bioRxiv2026

Principles of subclonal gene dosage across human cancer

Intratumor heterogeneity drives disease progression, but even though subclonal copy number variation (CNV) is a major contributor to this heterogeneity1, its impact on cell phenotype is not fully understood. Here, by applying high-quality joint whole genome sequencing and mRNA profiling in single cells (DNTR-seq2) to solid tumors and leukemias from 57 patients, we have analyzed the in vivo transcriptional effect of subclonal CNVs within a tumor. We found that gene dosage is generally additive in low and moderate copy states, but that cancer-type-specific compensation is common, and core promoter elements are associated with reduced additivity. We find that different classes of subclonal CNV impose varying degrees of transcriptional constraints on a cell, with highly amplified megabase-size regions associated with a strong effect both in cis and trans, while arm level CNVs generally exert a milder effect. We also describe a previously unappreciated class of tumors with transient clonality, where every cell is genetically highly distinct. We find that transient clonality is common in ovarian cancer and soft tissue sarcoma, that it is preceded by a whole genome duplication event, and that gene dosage in these tumors affects transcript abundance to a similar degree as in cancer with stable subclones.

cancer biology↗

bioRxiv2025

Recognition of Ovarian Tumor-Derived Non-canonical Peptide Induces Memory-Like Features in Natural Killer Cells

Immune cell-based immunotherapy has emerged as a promising strategy for both hematologic malignancies and solid tumors. The adaptive properties of T and B cells, namely, antigen specificity and long-term immune memory, form the foundation of approaches such as chimeric antigen receptor (CAR) T-cell therapy and tumor vaccines. In contrast, natural killer (NK) cells, traditionally classified as innate lymphocytes, have been appreciated primarily for their immediate cytotoxicity against tumor cells, but not for long-term memory-like responses. Recent evidence has revealed a subset of NK cells defined as adaptive NK cells (aNK) capable of developing adaptive features, particularly in response to cytomegalovirus (CMV) primarily, and more recently, to ovarian tumor-derived antigens. However, whether tumor-derived peptides can specifically induce NK cell memory, and the corresponding interaction patterns, remains largely unexplored. In this study, we employed a proteogenomic approach combining RNA sequencing (RNA-seq) with HLA-E immunoprecipitation to identify both canonical and non-canonical peptides presented by primary ovarian tumor cells. Among four tumor-derived neo-antigenic peptides, the 9-mer peptide APAPAPAPL demonstrated the strongest binding affinity to HLA-E and engagement with the NK receptors NKG2C/A. Functional in vitro assays confirmed that this peptide could induce memory-like NK cell responses, including antigen-specific recall activity and enhanced tumor cytotoxicity. Furthermore, structural modeling using AutoDock Vina and Rosetta Dock illustrated that peptides with similar binding capacity shared conserved interaction patterns and docking orientations. Together, this systemic study highlights a novel mechanism for inducing NK cell memory through tumor-derived neoantigens. It also paves the way for the development of NK cell-targeted cancer vaccines, representing a new direction in tumor immunotherapy beyond conventional T cell-centered strategies.

immunology↗

bioRxiv2025

Single-Cell Integration of Chromatin Accessibility and Transcriptomics Reveals Regulatory Networks in Ovarian Tumor-Infiltrating Adaptive NK Cells

Natural killer (NK) cells are traditionally recognized for their rapid, non-specific responses against virus-infected or malignantly transformed cells, functioning as key effectors of innate immunity. However, a distinct subset known as adaptive NK (aNK) cells has been shown to acquire memory-like properties following viral infections, indicating their capacity for antigen-specific immune recall. Intriguingly, aNK cells have also been identified within the tumor microenvironment, where they can mediate tumor-specific recall responses. Yet, the regulatory mechanisms governing their function in tumor-infiltrating aNK cells remain largely undefined. In this study, we integrated publicly available multiomics datasets from ovarian cancer, including single-cell chromatin accessibility (scATAC-seq) and single-cell RNA sequencing (scRNA-seq), to identify chromatin-accessible regions and construct transcription factors (TF)-gene regulatory networks. To validate and extend these findings, we performed Smart-seq3 on NK cells isolated from ovarian tumors and applied SCENIC analysis to identify TF-driven gene regulation. By integrating results from both analyses, we identified PRDM1 and STAT2 as key TFs, along with their downstream targets CRCP and MTFP1, specifically enriched in tumor-infiltrating aNK cells. The expression levels of CRCP and MTFP1 positively correlated with NK cell infiltration in ovarian cancer tissues, suggesting their potential functions in supporting tumor-specific NK cell memory responses. In addition, external validation using data from the PROMIX clinical trial demonstrated that PRDM1 and STAT2 expression levels are positively associated with both overall survival and aNK cell-associated transcriptional features. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/676040v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@6df026org.highwire.dtl.DTLVardef@1b16dc8org.highwire.dtl.DTLVardef@19e916borg.highwire.dtl.DTLVardef@15d431a_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2024

Genderized Gut and Oral Microbiome Shifts: Uncovering Sex-Specific Dysbiosis in Pancreatic Cancer

BackgroundPancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers, responsible for approximately 466,000 deaths globally in 2020. Its incidence increases by about 1% annually, with a higher occurrence in males than females. While differences in immune responses and tumor biology between sexes have been explored, the role of the microbiome in gender-specific PDAC progression is still unclear. Investigating these differences could offer crucial insights for personalized treatment strategies for males and females. MethodsThis study reanalyzed oral and gut microbiome data from BioProject: PRJNA832909, comprising 191 samples from PDAC patients and healthy controls. Using shotgun metagenomic sequencing, we examined gender-specific bacterial signatures. Alpha diversity (richness) and beta diversity (community composition) were analyzed. Differentially abundant bacterial taxa were identified via LEfSe, and gender-specific bacterial panels were validated using CombiROC. ResultsAlpha diversity analysis revealed significant differences in microbial richness, particularly between male and female PDAC patients and their healthy controls. Beta diversity demonstrated distinct microbial shifts between the PDAC and control groups across genders. LEfSe identified several pathogenic bacteria contributing to gender-specific dysbiosis, including Streptococcus, Fusobacterium, and Prevotella. Shared and sex-specific bacterial species in PDAC were highlighted through Venn diagram analysis. CombiROC validated the predictive ability of these bacterial markers, with AUC values exceeding 0.90 for both sexes. ConclusionThis study uncovered gender-specific microbial patterns in PDAC patients, potentially influenced by sex-specific immune responses. These findings provide important insights into the progression of PDAC and support sex-targeted diagnostic and therapeutic interventions. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/616338v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@4b1d97org.highwire.dtl.DTLVardef@183c267org.highwire.dtl.DTLVardef@1500547org.highwire.dtl.DTLVardef@8a854c_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2024

Advancing Adoptive Cell Therapy: Optimized Expansion of Adaptive NK Cells for Solid Tumors

BackgroundImmune therapies are emerging as a critical component of cancer treatment, capable of delivering durable and potentially curative responses. While CAR-T cell therapy has proven effective for hematological malignancies, it faces challenges in treating solid tumors due to tumor antigen heterogeneity, an immunosuppressive tumor microenvironment, and physical barriers hindering CAR-T cell infiltration. NK cells, particularly adaptive NK (aNK) cells, offer a promising alternative due to their ability to recognize and kill tumor cells without prior sensitization and their resistance to immunosuppressive environments. PurposeThe study investigates the role of cytokines, specifically IL-21 and IL-15, in enhancing aNK cell expansion and activation using peripheral blood mononuclear cells (PBMCs) from healthy donors and tumor-infiltrating lymphocytes (TILs) from glioblastoma (GBM) patients. MethodsBuffy coats and GBM TILs were collected from Karolinska Hospital. NK cells were isolated and expanded in vitro with IL-15 and IL-21 cytokines and feeder cells (K562 and K562E). Furthermore, tumor lysate was added in the cultures to boost memory responses in aNK cells. NK cell functionality, cytotoxicity, and phenotyping was assessed using flow cytometry and statistical analysis (t-test and two-way ANOVA) used to validate the results. Further animal model was used to validate the cytotoxicity capacities of these cells against GBM tumors using a zebrafish model. ResultsIL-21 drives the expansion of aNK better than IL-15 controls, data shown in PBMCs and TILs derived from GBM patients and IPLA OVCA patients. Additionally, the use of tumor lysate as a booster for restimulation further amplifies the cytotoxic capacity of aNK cells against autologous tumors. The zebrafish model validates this method, by decreasing the tumor size in zebrafish animals after 3 days of injection. ConclusionThe results demonstrate that IL-21 is essential for the specific expansion of aNK cells, enhancing their aggressiveness towards tumor cells. Additionally, tumor lysate significantly increases the cytotoxic efficacy of aNK cells upon restimulation with the same tumor cells. These findings suggest that IL-21 plays a crucial role in the specific expansion and activation of aNK cells, enhancing their aggressiveness towards tumor cells. By optimizing the expansion protocol, this method aims to advance the clinical application of aNK cells in immunotherapies for solid tumors, offering a potential solution to the limitations faced by current CAR-T therapies.

immunology↗

bioRxiv2024

Enhanced NK cell proficiency in solid tumors through antigen-specific memory via NKG2C/A-HLA-E/ABC recruited to tumors by CXCR2

Natural killer (NK) cells have emerged as promising effectors in cancer immunotherapy due to their ability to recognize and eliminate tumor cells. To investigate the immunological memory and tumor reactivity of adaptive (a)NK cells in the context of desmoplastic tumors, we used human ovarian cancer as a model. Through in vitro culture systems resembling dendritic cell (DC)-mediated T cell activation, we demonstrated that aNK cells exhibit antigen-specific cytotoxic responses and memory generation towards ovarian tumor antigens. Furthermore, mature DCs presenting tumor-associated antigens induced the expansion of aNK cells, suggesting antigen-specific proliferation. Single-cell transcriptomics revealed a distinct genetic signature of aNK cells in tumor samples, characterized by a cytotoxic phenotype and interactions with myeloid cells, particularly DCs. The spatial analysis confirmed the intratumoral presence of aNK cells, with higher abundance in the tumor nest compared to conventional (c)NK cells. Functional assays demonstrated the cytotoxicity of expanded aNK cells against autologous ovarian tumors, accompanied by an activated receptor profile. Importantly, aNK cells displayed antigen-specific memory responses towards primary tumors, maintaining specificity over time. Blockade of NKG2C and HLA-E influenced aNK cell recall responses, indicating their roles in the adaptive NK cell immune memory. Additionally, CXCR2 was essential for efficient aNK cell migration toward tumors. These findings shed light on the therapeutic potential of aNK cells in ovarian cancer immunotherapy, highlighting their ability to develop immunological memory and effectively eradicate tumor cells.

immunology↗

bioRxiv2024

IMMUNOPHENOTYPING AND FUNCTIONAL ANALYSIS OF NK CELL SUBSETS IN Mycobacterium tuberculosis-INFECTED INDIVIDUALS

Infection with Mycobacterium tuberculosis remains a global health challenge, with diverse clinical outcomes ranging from latent TB (LTB) infection to active TB disease (ATB). We conducted a comprehensive analysis of NK cell subsets and function in individuals with LTB, ATB, and healthy controls to elucidate their potential association with TB pathogenesis. Peripheral blood mononuclear cells (PBMCs) were isolated from Mtb-infected individuals and analyzed by mass cytometry and flow cytometry. We identified distinct NK cell subsets and evaluated their functional responses to stimulation. Our findings revealed comparable frequencies of total and NK cell subsets across LTB, ATB, and controls. Functional assays demonstrated similar degranulation, cytokine production, and proliferation capabilities among NK cell subsets across the three groups. This study provides insights into the heterogeneity of NK cell responses in TB and highlights the need for standardized methodologies with well-characterized cohorts controlling for donor background. Further investigations are warranted to delineate the specific roles of NK cells in TB immunity and pathogenesis.

immunology↗

bioRxiv2021

Regulatory FOXP3+ T cells in uterine sarcomas are associated with favorable prognosis, low extracellular matrix expression and reduced YAP activation

PurposeUterine sarcomas are rare but deadly malignancies without effective treatment. The goal of this study was to characterize and identify potential mechanisms underlying observed variations in the immune microenvironment of different sarcoma subtypes, using integrated clinicopathological and molecular methods. Experimental designFifty-eight cases of uterine sarcoma with full clinicopathological annotation were analyzed for their immune landscape in the tumor microenvironment, gene, and protein expression. Cases included leiomyosarcoma (LMS; n=13), low-grade endometrial stromal sarcoma (ESS; n=16), undifferentiated uterine sarcoma (UUS; n=26), and YWHAE-FAM22 translocation-bearing ESS (YFAM; n=3). Image analysis was used to quantify immune cells and immune regulatory proteins. Gene ontology and network enrichment analysis of matching transcriptomic data was used to relate over- and under expressed genes to pathways and further to the immune phenotype and clinicopathological findings. ResultsImmune cell characterization revealed overall prevalence of regulatory T cells and the pro-tumor M2-like macrophages. Cytotoxic T cells were only found in ESS and UUS tumors. Expression of immune regulatory proteins was heterogeneous, with PD-L1 being undetectable. Hierarchical clustering of patients showed four immune signatures independent of tumor type, where infiltration of non-exhausted FOXP3+ cells and M1-like macrophages were associated with greater overall survival. High CD8+/FOXP3+ ratio in UUS and ESS was associated with poor survival and upregulation of extracellular matrix (ECM)-related genes and proteins and YAP nuclear localization. ConclusionsUterine sarcomas present distinct immune signatures with prognostic value, independent of tumor type. This study suggests that the ECM is a potential regulator of the immune microenvironment in uterine sarcomas.

cancer biology↗