Biology subjects

Testa, J. R.

Publications and source records attributed to Testa, J. R..

4 recordsLinked to original sources

bioRxiv2023

Spatial landscape of malignant pleural and peritoneal mesothelioma tumor immune microenvironment

Immunotherapies have shown modest clinical benefit thus far for malignant mesothelioma (MM). A deeper understanding of immune cell spatial distribution within the tumor immune microenvironment (TIME) is needed to identify interactions between tumor and different immune cell types that might impact the effectiveness of potential immunotherapies. We performed multiplex immunofluorescence (mIF) using tissue microarrays (TMAs, n=3) of samples from patients with malignant peritoneal (n=25) and pleural (n=88) mesothelioma (MPeM and MPM, respectively) to elucidate the spatial distributions of major immune cell populations and their association with LAG3, BAP1, NF2, and MTAP expression, the latter as a proxy for CDKN2A/B. We also analyzed the relationship between the spatial distribution of major immune cell types with MM patient prognosis and clinical features. The distribution of immune cells within the TIME is similar between MPM and MPeM. However, there is a higher level of interaction between immune cells and tumor cells in MPM than MPeM. Within MPM tumors, there is increased amount of interaction between tumor cells and CD8+ T cells in BAP1-low than in BAP1-high expressing tumors. The cell-cell interactions identified in this investigation have potential implications for the immune response against MM tumors and could be a factor in the different behaviors of MPM and MPeM. Our findings provide a valuable resource for the MM cancer research community and exemplifies the utility of spatial resolution within single-cell analyses. Our mesothelioma spatial atlas mIF dataset is available at https://mesotheliomaspatialatlas.streamlit.app/.

bioinformatics↗

bioRxiv2022

Isolated BAP1 loss in malignant pleural mesothelioma predicts immunogenicity with implications for immunotherapeutic response

Malignant pleural mesothelioma (MPM), an aggressive cancer of the mesothelial cells lining the pleural cavity, lacks effective treatments. Multiple somatic mutations and copy number losses in tumor suppressor genes (TSGs) BAP1, CDKN2A/B, and NF2 are associated with MPM. The impact of single versus multiple losses of TSG on MPM biology, the immune tumor microenvironment, clinical outcomes, and treatment responses are unknown. Tumors with alterations in BAP1 alone were associated with a longer overall patient survival rate compared to tumors with CDKN2A/B and/or NF2 alterations with or without BAP1 and formed a distinct immunogenic subtype with altered transcription factor and pathway activity patterns. CDKN2A/B loss consistently contributed to an adverse clinical outcome. Since the loss of only BAP1 was associated with the PD-1 therapy response signature and higher LAG3 and VISTA gene expression, it is a candidate for immune checkpoint blockade therapy. Our results on the impact of TSG genotypes on MPM and the correlations between TSG alterations and molecular pathways provide a foundation for developing MPM therapies.

bioinformatics↗

bioRxiv2020

Inactivation of p21-Activated Kinase 2 (Pak2) Inhibits the Development of Nf2-Deficient Malignant Mesothelioma

Malignant mesotheliomas (MM) show frequent somatic loss of the NF2 tumor suppressor gene. The NF2 product, Merlin, is implicated in several tumor-related pathways, including p21-activated kinase (PAK) signaling. Merlin is both a phosphorylation target for PAK and a negative regulator of this oncogenic kinase. Merlin loss results in PAK activation, and PAK inhibitors hold promise for the treatment of NF2-deficient tumors. To test this possibility in an in vivo genetic system, Nf2f/f;Cdkn2af/f mice were crossed to mice with conditional knockout of Pak2, a highly expressed group I Pak member. Cohorts of these animals were injected in either the thoracic or peritoneal cavities with adeno-Cre virus to delete floxed alleles in the mesothelial lining. Loss of Pak2 resulted in a markedly decreased incidence and delayed onset and progression of pleural and peritoneal MMs in Nf2;Cdkn2a-deficient (NC) mice, as documented by Kaplan-Meier survival curves and in vivo bioluminescent imaging. RNA-seq revealed that MMs from NC;Pak2-/- mice showed downregulated expression of genes involved in several oncogenic pathways (Wnt, Akt) when compared to MMs from mice retaining Pak2. Kinome profiling showed that, as compared to NC MM cells, NC;Pak2-/- MM cells had multiple kinase changes indicative of an epithelial to mesenchymal transition. Collectively, these findings suggest that NC;Pak2-/- MMs adapt by reprogramming their kinome and gene signature profiles to bypass the need for PAK activity via the activation of other compensatory oncogenic kinase pathways. The identification of such secondary pathways offers opportunities for rational combination therapies to circumvent resistance to anti-PAK drugs.Competing Interest StatementThe authors have declared no competing interest.View Full Text

cancer biology↗

bioRxiv2020

Kinetics characterization of ASXL1/2-mediated allosteric regulation of BAP1 deubiquitinase

BAP1 is a ubiquitin hydrolase whose deubiquitinase activity is mediated by polycomb group-like protein ASXL2. Cancer-related mutations/deletions of BAP1 lead to loss-of-function either by directly targeting the catalytic (UCH) or ULD domains of BAP1, the latter disrupts binding to ASXL2, an obligate partner for BAP1 enzymatic activity. However, the biochemical and biophysical properties of the domains involved in forming the enzymatically active complex are unknown. Here we investigate the molecular dynamics, kinetics and stoichiometry of these interactions. We demonstrate that the BAP1 and ASXL2 domain/proteins or protein complexes produced in either bacteria or baculovirus are structurally and functionally active. The interaction between BAP1 and ASXL2 is direct, specific, and stable to in vitro biochemical and biophysical manipulations as detected by isothermal titration calorimetry, GST association, and optical biosensor assays. Association of the ASXL2-AB box greatly stimulates BAP1 deubiquitinase activity. A stable ternary complex can be formed comprised of the BAP1-UCH, BAP1-ULD, and ASXL2-AB domains. Binding of the BAP1-ULD domain to the ASXL2-AB box is rapid, with fast association and slow dissociation rates. Stoichiometric analysis revealed that one molecule of the ULD domain directly interacts with one molecule of the AB Box. Real-time kinetics analysis of ULD/AB protein complex to the UCH domain of BAP1, based on SPR, indicated that formation of the ULD/AB complex with the UCH domain is a single-step event with fast association and slow dissociation rates. These structural and dynamic parameters implicate the possibility for future small-molecule approaches to reactivate latent wild-type UCH activity in BAP-mutant malignancies.

cancer biology↗