Biology subjects

Bosse, K. R.

Publications and source records attributed to Bosse, K. R..

5 recordsLinked to original sources

bioRxiv2024

Blocking MIF secretion enhances CAR T-cell efficacy against neuroblastoma

While chimeric antigen receptor (CAR) T-cell therapies are showing highly promising first results in neuroblastoma, immunosuppressive tumor microenvironments (TME) limit T cell persistence and durable clinical efficacy. To improve CAR T-cell efficacy further, we applied a multi-omics approach including single-cell RNA sequencing and proteomics, which identified 13 targetable immunosuppressive factors in neuroblastoma. Of these, macrophage migration inhibitory factor (MIF) and midkine (MDK) were validated across multiple published RNA datasets. Moreover, they were secreted in high abundance by neuroblastoma tumoroids. Functional validation experiments revealed MIF as a potent inhibitor of CAR T-cells, in vitro and in vivo. Degradation of MIF by PROTAC technology significantly enhanced CAR T-cell activation targeting GPC2 and B7-H3, providing a potential intervention against MIF. By defining the immunosuppressive effects of neuroblastomas TME on CAR T-cell efficacy, particularly the pivotal role of MIF, we provide a therapeutic strategy for improving adoptive cell therapies for this pediatric malignancy.

cancer biology↗

bioRxiv2023

A proteogenomic surfaceome study identifies DLK1 as an immunotherapeutic target in neuroblastoma

Cancer immunotherapies have produced remarkable results in B-cell malignancies; however, optimal cell surface targets for many solid cancers remain elusive. Here, we present an integrative proteomic, transcriptomic, and epigenomic analysis of tumor specimens along with normal tissues to identify biologically relevant cell surface proteins that can serve as immunotherapeutic targets for neuroblastoma, an often-fatal childhood cancer of the developing nervous system. We apply this approach to human-derived cell lines (N=9) and cell/patient-derived xenograft (N=12) models of neuroblastoma. Plasma membrane-enriched mass spectrometry identified 1,461 cell surface proteins in cell lines and 1,401 in xenograft models, respectively. Additional proteogenomic analyses revealed 60 high-confidence candidate immunotherapeutic targets and we prioritized Delta-like canonical notch ligand 1 (DLK1) for further study. High expression of DLK1 directly correlated with the presence of a super-enhancer spanning the DLK1 locus. Robust cell surface expression of DLK1 was validated by immunofluorescence, flow cytometry, and immunohistochemistry. Short hairpin RNA mediated silencing of DLK1 in neuroblastoma cells resulted in increased cellular differentiation. ADCT-701, a DLK1-targeting antibody-drug conjugate (ADC), showed potent and specific cytotoxicity in DLK1-expressing neuroblastoma xenograft models. Moreover, DLK1 is highly expressed in several adult cancer types, including adrenocortical carcinoma (ACC), pheochromocytoma/paraganglioma (PCPG), hepatoblastoma, and small cell lung cancer (SCLC), suggesting potential clinical benefit beyond neuroblastoma. Taken together, our study demonstrates the utility of comprehensive cancer surfaceome characterization and credentials DLK1 as an immunotherapeutic target. HighlightsO_LIPlasma membrane enriched proteomics defines surfaceome of neuroblastoma C_LIO_LIMulti-omic data integration prioritizes DLK1 as a candidate immunotherapeutic target in neuroblastoma and other cancers C_LIO_LIDLK1 expression is driven by a super-enhancer C_LIO_LIDLK1 silencing in neuroblastoma cells results in cellular differentiation C_LIO_LIADCT-701, a DLK1-targeting antibody-drug conjugate, shows potent and specific cytotoxicity in DLK1-expressing neuroblastoma preclinical models C_LI

cancer biology↗

bioRxiv2023

BARD1 germline variants induce haploinsufficiency and DNA repair defects in neuroblastoma

ImportanceHigh-risk neuroblastoma is a complex genetic disease that is lethal in 50% of patients despite intense multimodal therapy. Our genome-wide association study (GWAS) identified single-nucleotide polymorphisms (SNPs) within the BARD1 gene showing the most significant enrichment in neuroblastoma patients, and also discovered pathogenic (P) or likely pathogenic (LP) rare germline loss-of-function variants in this gene. The functional implications of these findings remain poorly understood. ObjectiveTo define the functional relevance of BARD1 germline variation in children with neuroblastoma. DesignWe correlated BARD1 genotype with BARD1 expression in normal and tumor cells and the cellular burden of DNA damage in tumors. To validate the functional consequences of rare germline P-LP BARD1 variants, we generated isogenic cellular models harboring heterozygous BARD1 loss-of-function (LOF) variants and conducted multiple complementary assays to measure the efficiency of DNA repair. Setting(N/A) Participants(N/A) Interventions/Exposures(N/A) Main Outcomes and MeasuresBARD1 expression, efficiency of DNA repair, and genome-wide burden of DNA damage in neuroblastoma tumors and cellular models harboring disease-associated BARD1 germline variants. ResultsBoth common and rare neuroblastoma associated BARD1 germline variants were significantly associated with lower levels of BARD1 mRNA and an increased burden of DNA damage. Using neuroblastoma cellular models engineered to harbor disease-associated heterozygous BARD1 LOF variants, we functionally validated this association with inefficient DNA repair. These BARD1 LOF variant isogenic models exhibited reduced efficiency in repairing Cas9-induced DNA damage, ineffective RAD51 focus formation at DNA doublestrand break sites, and enhanced sensitivity to cisplatin and poly-ADP ribose polymerase (PARP) inhibition. Conclusions and RelevanceConsidering that at least 1 in 10 children diagnosed with cancer carry a predicted pathogenic mutation in a cancer predisposition gene, it is critically important to understand their functional relevance. Here, we demonstrate that germline BARD1 variants disrupt DNA repair fidelity. This is a fundamental molecular mechanism contributing to neuroblastoma initiation that may have important therapeutic implications, and these findings may also extend to other cancers harboring germline variants in genes essential for DNA damage repair. Key PointsO_ST_ABSQuestionC_ST_ABSHow do neuroblastoma patient BRCA1-associated RING domain 1 (BARD1) germline variants impact DNA repair? FindingsNeuroblastoma-associated germline BARD1 variants disrupt DNA repair fidelity. Common risk variants correlate with decreased BARD1 expression and increased DNA double-strand breaks in neuroblastoma tumors and rare heterozygous loss-of-function variants induce BARD1 haploinsufficiency, resulting in defective DNA repair and genomic instability in neuroblastoma cellular models. MeaningGermline variation in BARD1 contributes to neuroblastoma pathogenesis via dysregulation of critical cellular DNA repair functions, with implications for neuroblastoma treatment, risk stratification, and cancer predisposition.

genetics↗

bioRxiv2021

Development of GPC2-directed chimeric antigen receptors using mRNA for pediatric brain tumors

Pediatric brain tumors are the leading cause of cancer death in children with an urgent need for innovative therapies. Here we show that the cell surface oncoprotein glypican 2 (GPC2) is highly expressed on multiple lethal pediatric brain tumors, including medulloblastomas, embryonal tumors with multi-layered rosettes, other CNS embryonal tumors, as well as definable subsets of highly malignant gliomas. To target GPC2 on these pediatric brain tumors with adoptive cellular therapies and mitigate potential inflammatory neurotoxicity, we developed four mRNA chimeric antigen receptor (CAR) T cell constructs using the highly GPC2-specific fully human D3 single chain variable fragment. First, we validated and prioritized these CARs using in vitro cytotoxicity and T cell degranulation assays with GPC2-expressing neuroblastoma cells. Next, we expanded the testing of the two most potent GPC2-directed CAR constructs prioritized from these studies to GPC2-expressing medulloblastoma and high-grade glioma cell lines, showing significant GPC2-specific cell death in multiple models. Finally, locoregional delivery bi-weekly of two to four million GPC2-directed mRNA CAR T cells induced significant and sustained tumor regression in two orthotopic medulloblastoma models, and significantly prolonged survival in an aggressive orthotopic thalamic diffuse midline glioma model. No GPC2-directed CAR T cell related neurologic or systemic toxicity was observed. Taken together, these data show that GPC2 is a highly differentially expressed cell surface protein on multiple malignant pediatric brain tumors that can be targeted safely with local delivery of mRNA CAR T cells. One Sentence SummaryGlypican 2 is expressed on the surface of multiple pediatric brain tumors and can be successfully targeted with mRNA chimeric antigen receptor T cells.

cancer biology↗

bioRxiv2019

LIN28B-PDZ Binding Kinase Signaling Promotes Neuroblastoma Metastasis

Neuroblastoma is an aggressive pediatric malignancy of the neural crest with suboptimal cure rates and a striking predilection for widespread metastases, underscoring the need to identify novel therapeutic vulnerabilities. We recently identified the RNA binding protein LIN28B as a driver in high-risk neuroblastoma and demonstrated it promotes oncogenic cell proliferation by coordinating a RAN-Aurora kinase A network. Here, we demonstrate that LIN28B influences another key hallmark of cancer, metastatic dissemination. Using a murine xenograft model of neuroblastoma dissemination, we show that LIN28B promotes metastasis. We demonstrate that this is in part due to the effects of LIN28B on self-renewal and migration, providing an understanding of how LIN28B shapes the metastatic phenotype. Our studies reveal that the let-7 family, which LIN28B inhibits, opposes the effects of LIN28B. Next, we identify PDZ Binding Kinase (PBK) as a novel LIN28B target. PBK is a serine/threonine kinase that promotes the proliferation and self-renewal of neural stem cells and serves as an oncogenic driver in multiple aggressive malignancies. We demonstrate that PBK is both a novel direct target of let-7 and that MYCN regulates PBK expression, thus elucidating two oncogenic drivers that converge on PBK. Functionally, PBK promotes self-renewal and migration, phenocopying LIN28B. Taken together, our findings define a role for LIN28B in neuroblastoma metastasis and define the targetable kinase PBK as a potential novel vulnerability in metastatic neuroblastoma.

cancer biology↗