Biology subjects

Costello, J. F.

Publications and source records attributed to Costello, J. F..

7 recordsLinked to original sources

bioRxiv2023

Challenges in the discovery of tumor-specific alternative splicing-derived cell-surface antigens in glioma

BackgroundDespite advancements in cancer immunotherapy, solid tumors remain formidable challenges. In glioma, profound inter-and intra-tumoral heterogeneity of antigen landscape hampers therapeutic development. Therefore, it is critical to consider alternative sources to expand the repertoire of targetable (neo-)antigens and improve therapeutic outcomes. Accumulating evidence suggests that tumor-specific alternative splicing (AS) could be an untapped reservoir of neoantigens. ResultsIn this study, we investigated tumor-specific AS events in glioma, focusing on those predicted to generate major histocompatibility complex (MHC)-presentation-independent, cell-surface neoantigens that could be targeted by antibodies and chimeric antigen receptor (CAR)-T cells. We systematically analyzed bulk RNA-sequencing datasets comparing 429 tumor samples (from The Cancer Genome Atlas [TCGA]) and 9,166 normal tissue samples (from the Genotype-Tissue Expression project [GTEx]), and identified 13 AS events in 7 genes predicted to be expressed in more than 10% of the patients, including PTPRZ1 and BCAN, which were corroborated by an external RNA-sequencing dataset. Subsequently, we validated our predictions and elucidated the complexity of the isoforms using full-length transcript amplicon sequencing on patient-derived glioblastoma cells. However, analyses of the RNA-sequencing datasets of spatially mapped and longitudinally collected clinical tumor samples unveiled remarkable spatiotemporal heterogeneity of the candidate AS events. Furthermore, proteomics analysis did not reveal any peptide spectra matching the putative neoantigens. ConclusionsOur investigation illustrated the diverse characteristics of the tumor-specific AS events and the challenges of antigen exploration due to their notable spatiotemporal heterogeneity and elusive nature at the protein levels. Redirecting future efforts toward intracellular, MHC-presented antigens could offer a more viable avenue.

cancer biology↗

bioRxiv2023

Tumor-wide RNA splicing aberrations generate immunogenic public neoantigens

T-cell-mediated immunotherapies are limited by the extent to which cancer-specific antigens are homogenously expressed throughout a tumor. We reasoned that recurrent splicing aberrations in cancer represent a potential source of tumor-wide and public neoantigens, and to test this possibility, we developed a novel pipeline for identifying neojunctions expressed uniformly within a tumor across diverse cancer types. Our analyses revealed multiple neojunctions that recur across patients and either exhibited intratumor heterogeneity or, in some cases, were tumor-wide. We identified CD8+ T-cell clones specific for neoantigens derived from tumor-wide and conserved neojunctions in GNAS and RPL22, respectively. TCR-engineered CD8+ T-cells targeting these mutations conferred neoantigen-specific tumor cell eradication. Furthermore, we revealed that cancer-specific dysregulation in splicing factor expression leads to recurrent neojunction expression. Together, these data reveal that a subset of neojunctions are both intratumorally conserved and public, providing the molecular basis for novel T-cell-based immunotherapies that address intratumoral heterogeneity.

cancer biology↗

bioRxiv2023

Clarifying the Transcriptional Profiles of Malignant Clones through Multiscale and Multiomic Analysis of Individual Tumors

Tumors may contain billions of cells including distinct malignant clones and nonmalignant cell types. Clarifying the evolutionary histories, prevalence, and defining molecular features of these cells is essential for improving clinical outcomes, since intratumoral heterogeneity provides fuel for acquired resistance to targeted therapies. Here we present a statistically motivated strategy for deconstructing intratumoral heterogeneity through multiomic and multiscale analysis of serial tumor sections (MOMA). By combining deep sampling of IDH-mutant astrocytomas with integrative analysis of single-nucleotide variants, copy-number variants, and gene expression, we reconstruct and validate the phylogenies, spatial distributions, and transcriptional profiles of distinct malignant clones. By genotyping nuclei analyzed by single-nucleus RNA-seq for truncal mutations, we further show that commonly used algorithms for identifying cancer cells from single-cell transcriptomes may be inaccurate. We also demonstrate that correlating gene expression with tumor purity in bulk samples can reveal optimal markers of malignant cells and use this approach to identify a core set of genes that is consistently expressed by astrocytoma truncal clones, including AKR1C3, whose expression is associated with poor outcomes in several types of cancer. In summary, MOMA provides a robust and flexible strategy for precisely deconstructing intratumoral heterogeneity and clarifying the core molecular properties of distinct cellular populations in solid tumors.

cancer biology↗

bioRxiv2021

Identification of Cancer-Associated Fibroblasts in Glioblastoma and Defining Their Pro-tumoral Effects

Despite their identification in some cancers, pro-tumoral cancer-associated fibroblasts (CAFs) were presumed absent in glioblastoma given the lack of brain fibroblasts. Serial trypsinization of primary glioblastoma cultures yielded cells with CAF morphology, CAF transcriptomic profile, and mesenchymal lineage in single-cell RNA-seq. Glioblastoma CAFs were attracted to glioblastoma stem cells (GSCs) and CAFs enriched GSCs. We created a resource of inferred crosstalk by mapping expression of receptors to their cognate ligands, identifying PDGF-{beta} and TGF-{beta} as mediators of GSC effects on CAFs, and osteopontin and hepatocyte growth factor as mediators of CAF-induced GSC enrichment. Glioblastoma CAFs also induced M2 macrophage polarization by producing the EDA fibronectin variant. Glioblastoma CAFs were enriched in the subventricular zone which houses neural stem cells that produce GSCs. Including CAFs in GSC-derived xenografts induced in vivo growth. These findings are among the first to identify glioblastoma CAFs and their GSC interactions, making them an intriguing target.

cancer biology↗

bioRxiv2020

Cancer-specific loss of TERT activation sensitizes glioblastoma to DNA damage

Most glioblastomas (GBMs) achieve cellular immortality by acquiring a mutation in the telomerase reverse transcriptase (TERT) promoter. TERT promoter mutations create a binding site for a GA binding protein (GABP) transcription factor complex, whose assembly at the promoter is associated with TERT reactivation and telomere maintenance. Here, we demonstrate increased binding of a specific GABPB1L isoform-containing complex to the mutant TERT promoter. Furthermore, we find that TERT promoter mutant GBM cells, unlike wild-type cells, exhibit a critical near-term dependence on GABPB1L for proliferation, both in cell culture and post-tumor establishment in vivo. Upregulation of the protein paralogue GABPB2, which is normally expressed at very low levels, can rescue this dependence. More importantly, when combined with frontline temozolomide (TMZ) chemotherapy, inducible GABPB1L knockdown and the associated TERT reduction led to an impaired DNA damage response that resulted in profoundly reduced growth of intracranial GBM tumors. Together, these findings provide new insights into the mechanism of cancer-specific TERT regulation, uncover rapid effects of GABPB1L-mediated TERT suppression in GBM maintenance, and establish GABPB1L inhibition in combination with chemotherapy as a novel therapeutic strategy for TERT promoter mutant GBM.

cancer biology↗

bioRxiv2020

DNA methylation classification in diffuse glioma shows little spatial heterogeneity after adjusting for tumor purity

Intratumoral heterogeneity is a hallmark of diffuse gliomas. We used neuronavigation to acquire 133 image-guided and spatially-separated stereotactic biopsy samples from 16 adult patients with a diffuse glioma, which we characterized using DNA methylation arrays. Samples were obtained from regions with and without imaging abnormalities. Methylation profiles were analyzed to devise a three-dimensional reconstruction of genetic and epigenetic heterogeneity. Molecular aberrations indicated that tumor was found outside imaging abnormalities, underlining the infiltrative nature of this tumor and the limitations of current routine imaging modalities. We demonstrate that tumor purity is highly variable between samples and largely explains apparent epigenetic spatial heterogeneity. Indeed, we observed that DNA methylation subtypes are highly conserved in space after adjusting for tumor purity. Genome-wide heterogeneity analysis showed equal or increased heterogeneity among normal tissue when compared to tumor. These findings were validated in a separate cohort of 61 multi-sector tumor and 64 normal samples. Our findings underscore the infiltrative nature of diffuse gliomas and suggest that heterogeneity in DNA methylation is innate to somatic cells and not a characteristic feature of this tumor type.

cancer biology↗

bioRxiv2018

Saturation mutagenesis of disease-associated regulatory elements

The majority of common variants associated with common diseases, as well as an unknown proportion of causal mutations for rare diseases, fall in noncoding regions of the genome. Although catalogs of noncoding regulatory elements are steadily improving, we have a limited understanding of the functional effects of mutations within them. Here, we performed saturation mutagenesis in conjunction with massively parallel reporter assays on 20 disease-associated gene promoters and enhancers, generating functional measurements for over 30,000 single nucleotide substitution and deletion mutations. We find that the density of putative transcription factor binding sites varies widely between regulatory elements, as does the extent to which evolutionary conservation or various integrative scores predict functional effects. These data provide a powerful resource for interpreting the pathogenicity of clinically observed mutations in these disease-associated regulatory elements, and also comprise a gold-standard dataset for the further development of algorithms that aim to predict the regulatory effects of noncoding mutations.

genetics↗