Biology subjects

Saha, L. K.

Publications and source records attributed to Saha, L. K..

3 recordsLinked to original sources

bioRxiv2024

Genome-wide Mapping of Topoisomerase Binding Sites Suggests Topoisomerase 3α(TOP3A) as a Reader of Transcription-Replication Conflicts (TRC)

Both transcription and replication can take place simultaneously on the same DNA template, potentially leading to transcription-replication conflicts (TRCs) and topological problems. Here we asked which topoisomerase(s) is/are the best candidate(s) for sensing TRC. Genome-wide topoisomerase binding sites were mapped in parallel for all the nuclear topoisomerases (TOP1, TOP2A, TOP2B, TOP3A and TOP3B). To increase the signal to noise ratio (SNR), we used ectopic expression of those topoisomerases in H293 cells followed by a modified CUT&Tag method. Although each topoisomerase showed distinct binding patterns, all topoisomerase binding signals positively correlated with gene transcription. TOP3A binding signals were suppressed by DNA replication inhibition. This was also observed but to a lesser extent for TOP2A and TOP2B. Hence, we propose the involvement of TOP3A in sensing both head-on TRCs (HO-TRCs) and codirectional TRCs (CD-TRCs). In which case, the TOP3A signals appear concentrated within the promoters and first 20 kb regions of the 5 -end of genes, suggesting the prevalence of TRCs and the recruitment of TOP3A in the 5-regions of transcribed and replicated genes. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=81 SRC="FIGDIR/small/599352v1_ufig1.gif" ALT="Figure 1"> View larger version (10K): org.highwire.dtl.DTLVardef@1d1afdeorg.highwire.dtl.DTLVardef@17146eaorg.highwire.dtl.DTLVardef@1e6bed3org.highwire.dtl.DTLVardef@1fa40db_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2022

ATR inhibition augments the efficacy of lurbinectedin in small cell lung cancer

Abstract/SummarySmall cell lung cancer (SCLC) is the most lethal type of lung cancer. Specifically, MYC-driven non-neuroendocrine SCLC are particularly resistant to standard therapies. Lurbinectedin was recently approved for the treatment of relapsed SCLC, but combinatorial approaches are needed to increase the depth and duration of responses to lurbinectedin. Using high-throughput screens, we found inhibitors of ataxia telangiectasia-mutated and rad3-related (ATR) as the most effective agents for augmenting lurbinectedin efficacy. First-in-class ATR inhibitor berzosertib synergized with lurbinectedin in multiple SCLC cell lines, organoid and in-vivo models. Mechanistically, ATR inhibition abrogated S-phase arrest induced by lurbinectedin and forced cell-cycle progression causing mitotic catastrophe and cell death. High CDKN1A/p21 expression was associated with decreased synergy due to G1 arrest, while increased levels of ERCC5/XPG were predictive of increased combination efficacy. Importantly, MYC-driven non-neuroendocrine tumors which were resistant to first-line therapies displayed decreased CDKN1A/p21 expression and increased ERCC5/XPG indicating they were primed for response to lurbinectedin-berzosertib combination. The combination is being assessed in a clinical trial NCT04802174. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=198 SRC="FIGDIR/small/520783v1_ufig1.gif" ALT="Figure 1"> View larger version (63K): org.highwire.dtl.DTLVardef@19acf72org.highwire.dtl.DTLVardef@a9703forg.highwire.dtl.DTLVardef@86592org.highwire.dtl.DTLVardef@1448372_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2022

A high-throughput 384-well CometChip platform reveals a role for 3-methyladenine in the cellular response to etoposide-induced DNA damage

The Comet or single-cell gel electrophoresis assay is the gold standard for analysis of cellular, nuclear genome damage. However, low throughput limits its application for large-scale studies. To overcome these limitations, a 96-well CometChip platform was recently developed that increases throughput and reduces variation due to simultaneous processing and automated analysis of 96 samples. To advance throughput further, we developed a 384-well CometChip platform that allows analysis of [~]100 cells per well. The 384-well CometChip extends the capacity by 4-fold as compared to the 96-well system, enhancing application for larger DNA damage analysis studies. The overall sensitivity of the 384-well CometChip is consistent with that of the 96-well system, sensitive to genotoxin exposure and to loss of DNA repair capacity. We then applied the 384-well platform to screen a library of protein kinase inhibitors to probe each as enhancers of etoposide induced DNA damage and found that 3-methyladenine significantly increased levels of etoposide-induced DNA damage. Our results suggest that a 384-well CometChip is useful for large-scale DNA damage analyses, which may have increased potential in the evaluation of chemotherapy efficacy, compound library screens, population-based analyses of genome damage and evaluating the impact of environmental genotoxins on genome integrity.

cancer biology↗