Biology subjects

Viola, J. P. B.

Publications and source records attributed to Viola, J. P. B..

5 recordsLinked to original sources

bioRxiv2025

Resistance to BRAF inhibitors drives melanoma sensitivity to Chk1 inhibition

BRAF inhibitor-resistant melanomas (BRAFiR) acquire (epi)genetic and functional alterations that enable them to evade alternative treatments. Identifying these alterations is critical to advancing treatment strategies. Here, we explored the effect of Chk1 inhibition (Chk1i) on BRAFiR cells, revealing higher sensitivity compared to treatment-naive cells both in vitro and in vivo. Using FUCCI-labeling and time-lapse microscopy, we show that S phase progression is required for Chk1i-induced cytotoxicity in BRAFiR cells, but not in treatment-naive cells. Replication stress markers, including reduced BrdU incorporation and increased phospho-RPA and {gamma}H2AX, were exclusive to BRAFiR cells exposed to Chk1i. Untreated BRAFiR cells exhibited upregulated DNA replication genes, reduced progressing forks and increased origin firing, suggesting intrinsic replication changes. MAPK pathway reactivation in treatment-naive cells mimicked BRAFiR traits, increasing sensitivity to Chk1i. These findings indicate that Chk1i exploits elevated replication stress specifically in BRAFiR, highlighting its therapeutic potential in overcoming MAPK inhibitor resistance in melanoma.

cancer biology↗

bioRxiv2022

Polycomb Repressive Complex 1 subunit Cbx4 positively regulates effector responses in CD8 T cells

CD8 T cell differentiation is controlled by the crosstalk of various transcription factors and epigenetic modulators. Uncovering the different players in regulating this process is fundamental to improving immunotherapy and designing novel therapeutic approaches. Here, we show that Polycomb Repressive Complex (PRC)1 subunit Chromobox (Cbx)4 favors differentiation to effector CD8 T cells. Cbx4 deficiency in CD8 T cells induced transcriptional signature and phenotype of memory cells, increasing the formation of memory population during acute viral infection. It has been previously shown that besides chromodomain-mediated binding to H3K27me3, Cbx4 function as a SUMO E3 ligase in a SUMO interacting motifs (SIM)-dependent way. The overexpression of Cbx4 mutants in distinct domains showed that this protein regulates CTL differentiation primarily in a SIM-dependent way and partially through its chromodomain. Our data revealed a novel role of a Polycomb group protein Cbx4 controlling CD8 T lymphocyte differentiation and indicates the SUMOylation process as a key molecular mechanism connected to chromatin modification in this process. SummaryUnderstanding the epigenetic control of CTL differentiation is critical for the manipulation of these cells in immunotherapy protocols. This article demonstrates a novel role for Cbx4, a Polycomb-group protein, in supporting CD8 T cell commitment to an effector cell phenotype.

immunology↗

bioRxiv2021

Atazanavir is a competitive inhibitor of SARS-CoV-2 Mpro, impairing variants replication in vitro and in vivo

Atazanavir (ATV) has already been considered as a potential repurposing drug to 2019 coronavirus disease (COVID-19), however, there are controversial reports on its mechanism of action and effectiveness as anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Through the pre-clinical chain of experiments: enzymatic, molecular docking, cell-based, and in vivo assays, it is demonstrated here that both SARS-CoV-2 B.1 lineage and variant of concern gamma are susceptible to this antiretroviral. Enzymatic assays and molecular docking calculations showed that SARS-CoV-2 main protease (Mpro) was inhibited by ATV, with Morrisons inhibitory constant (Ki) 1.5-fold higher than boceprevir (GC376, a positive control). ATV was a competitive inhibition, increasing the Mpros Michaelis-Menten (Km) more than 6-fold. Cell-based assays indicated that SARS-CoV-2 gamma is more susceptible to ATV than its predecessor strain B.1. Using oral administration of ATV in mice to reach plasmatic exposure similar to humans, transgenic mice expression in human angiotensin converting enzyme 2 (K18-hACE2) were partially protected against lethal challenge with SARS-CoV-2 gamma. Moreover, less cell death and inflammation were observed in the lung from infected and treated mice. Our studies may contribute to a better comprehension of the Mpro/ATV interaction, which could pave the way to the development of specific inhibitors of this viral protease.

pharmacology and toxicology↗

bioRxiv2021

The transcriptional cofactor IRF2BP2 plays a key role in T cell homeostasis and Treg cell expansion

The levels of the co-transcriptional regulator IRF2BP2 (Interferon Regulatory Factor-2 Binding Protein-2) decrease with T cell activation and, when ectopically expressed, it reduces T cell proliferation. To further characterize the function of IRF2BP2 in T cell responses in vivo, we generated a conditional transgenic knock-in mouse that overexpresses IRF2BP2 in T lymphocytes. Overexpression of IRF2BP2 leads to a reduction in the T cell compartment of naive animals, upregulation of Foxp3 and Ifng; an increase in the frequency of regulatory T cells (Tregs), a preferential Th1 differentiation with increase of IFN-{gamma} production and a reduction of T cell proliferation, suggesting a disruption in T cell homeostasis. Interestingly, knock-in mice displayed reduced clinical and inflammatory signs of Experimental Autoimmune Encephalomyelitis (EAE) when compared to the control mice, with an augmented frequency of Treg cells. Altogether, our findings indicate that IRF2BP2 might help to control exacerbated T cell responses and point to a role for IRF2BP2 in preventing T cell autoimmunity.

immunology↗

bioRxiv2020

SARS-CoV-2 genomic and quasispecies analyses in cancer patients reveal relaxed intrahost virus evolution

Numerous factors have been identified to influence susceptibility to SARS-CoV-2 infection and disease severity. Cancer patients are more prone to clinically evolve to more severe COVID-19 conditions, but the determinants of such a more severe outcome remain largely unknown. We have determined the full-length SARS-CoV-2 genomic sequences of cancer patients and healthcare workers (HCW; non-cancer controls) by deep sequencing and investigated the within-host viral quasispecies of each infection, quantifying intrahost genetic diversity. Naso- and oropharyngeal SARS-CoV-2+ swabs from 57 cancer patients and 14 healthcare workers (HCW) from the Brazilian Cancer Institute were collected in April-May 2020. Complete genome amplification using ARTIC network V3 multiplex primers was performed followed by next-generation sequencing. Assemblies were conducted in Geneious R11, where consensus sequences were extracted and intrahost single nucleotide variants (iSNVs) were identified. Maximum likelihood phylogenetic analysis was performed using PhyMLv.3.0 and lineages were classified using Pangolin and CoV-GLUE. Phylogenetic analysis showed that all but one strain belonged to clade B1.1. Four genetically linked mutations known as the globally dominant SARS-CoV-2 haplotype (C241T, C3037T, C14408T and A23403G) were found in the majority of consensus sequences. SNV signatures of previously characterized Brazilian genomes were also observed in most samples. Another 85 SNVs were found at a lower frequency (1.4-19.7%). Cancer patients displayed a significantly higher intrahost viral genetic diversity compared to HCW (p = 0.009). Intrahost genetic diversity in cancer patients was independent of SARS-CoV-2 Ct values, and was not associated with disease severity, use of corticosteroids, or use of antivirals, characteristics that could influence viral diversity. Such a feature may explain, at least in part, the more adverse outcomes to which cancer/COVID-19 patients experience. Author SummaryCancer patients are more prone to clinically evolve to more severe COVID-19 conditions, but the determinants of such a more severe outcome remain largely unknown. In this study, phylogenetic and variation analysis of SARS-CoV-2 genomes from cancer patients and non-cancer healthcare workers at the Brazilian National Cancer Institute were characterized by deep sequencing. Viral genomes showed signatures characteristic of Brazilian viruses, consistent with the hypothesis of local, community transmission rather than virus importation from abroad. Despite most genomes in patients and healthcare workers belonging to the same lineage, intrahost variability was higher in cancer patients when compared to non-cancer counterparts. The intrahost genomic diversity analysis presented in our study highlights the relaxed evolution of SARS-CoV-2 in a vulnerable population of cancer patients. The high number of minor variations can result in the selection of immune escape variants, resistance to potential drugs, and/or increased pathogenicity. The impact of this higher intrahost variability over time warrants further investigation.

microbiology↗