Abstract
Somatic heterozygous mutations in genes encoding for RNA splicing factors (SF) SRSF2, U2AF1, and SF3B1 are frequently mutated in patients with hematologic malignancies, representing a unique ...genetic vulnerability for targeted therapy. In the current study, we performed a focused drug screen with inhibitors targeting different DNA damage response and DNA metabolic pathways to identify novel therapeutic vulnerabilities generated by SF mutations. We generated a murine leukemia model by overexpressing the MLL-AF9 fusion oncogene on an Srsf2P95H/+ background, a mutational combination that is found in ~10% of MLL-rearranged leukemias. We surprisingly found that MLL-AF9 Srsf2P95H/+ mutant leukemias are sensitive to inhibitors targeting ADP-ribosyltransferases (PARP). PARP inhibitor sensitivity was also observed in isogenic murine MLL-AF9 U2af1s34/+ cells compared to MLL-AF9 U2af1+/+ cells. Second, murine Srsf2P95H leukemias showed improved prolonged survival when treated with olaparib (PARPi) compared to vehicle treatment in vivo. Third, human primary AML patient samples that harbor SF mutations are sensitive to PARPi compared to non-SF mutant samples. These data highlight that both SRSF2P95H and U2AF1S34F mutations create a common vulnerability that is dependent on PARP activity for survival. To evaluate PARP activity, we used isogenic K562 leukemia cells expressing SRSF2P95H and U2AF1S34F mutations from their endogenous loci and monitored ADP-ribosylation (ADPr) levels, a marker of PARP activity. Both SRSF2P95H and U2AF1S34F cells exhibited elevated levels of ADPr compared to wildtype cells in a PARP1- dependent manner. PARPi preferentially induced DNA damage and cell death in SF mutant cells. Surprisingly, we found that SRSF2P95H and U2AF1S34F cells are not defective in homologous recombination repair. Instead, the increased PARP1-mediated ADPr in SF-mutant cells is caused by accumulated R loops, a group of transcription intermediates containing RNA:DNA hybrids and displaced single-stranded DNA. To determine whether PARPi sensitivity is due to R-loop accumulation, we overexpressed RNase H1, an enzyme that specifically cleaves the RNA moiety within RNA:DNA hybrids in U2AF1S34F cells. Overexpression of RNase H1 significantly reduced ADPr levels and suppressed the PARPi-induced U2AF1S34F cell growth inhibition. Collectively, these results suggest that spliceosome mutants induce R-loop accumulation and elicit an R-loop-associated PARP1 response to promote cell survival. In summary, our data establish a previously unknown link between R-loop-induced PARP1 response and RNA splicing perturbation and provide a mechanistic rationale to evaluate the clinical efficacy of PARP inhibitors in spliceosome-mutant malignancies. Furthermore, our study highlights a new therapeutic potential of targeting the R-loop tolerance pathways caused by different spliceosome gene mutations.
Citation Format: Dang Hai Nguyen, Sayantani Sinha, Zhiyan Silvia Liu, Maxwell Henry Bannister, Erica Arriaga-Gomez, Axia Song, Dawei Zong, Martina Sarchi, Victor Corral, Wannasiri Chiraphapphaiboon, Jennifer Yoo, Matthew McMahon, Cassandra Leibson, Derek L. Stirewalt, H Joachim Deeg, Sumit Rai, Matthew Walter, Timothy A. Graubert, Sergei Doulatov, Stanley C. Lee. PARP inhibitors preferentially sensitize splicing factor mutant myeloid neoplasms. abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6183.
If sexual compulsivity and other addictive behaviours share common aetiology, contemporary proposals about the role of attentional processes in understanding addictive behaviours are relevant.
To ...examine attentional biases for sex-related words among sexually active individuals and the relationship between sexual compulsivity and sexual behavioural engagement with attentional bias, 55 sexually active individuals completed a modified Stroop task and the sexual compulsivity scale.
Findings showed attentional bias towards sex-related stimuli among sexually active participants. In addition, among those with low levels of sexual compulsivity, levels of attentional bias were the same across all levels of sexual experience. Among those with higher levels of sexual compulsivity, greater attentional bias was linked with lower levels of sexual experience.
Attentional preference for concern-related stimuli varies as a function of the interaction between how long a person has been active sexually and how compulsive their sexual behaviour is.
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The study of single cells has evolved over the past several years to include expression and genomic analysis of an increasing number of single cells. Several studies have demonstrated wide spread ...variation and heterogeneity within cell populations of similar phenotype. While the characterization of these populations will likely set the foundation for our understanding of genomic- and expression-based diversity, it will not be able to link the functional differences of a single cell to its underlying genomic structure and activity. Currently, it is difficult to perturb single cells in a controlled environment, monitor and measure the response due to perturbation, and link these response measurements to downstream genomic and transcriptomic analysis. In order to address this challenge, we developed a platform to integrate and miniaturize many of the experimental steps required to study single-cell function. The heart of this platform is an elastomer-based integrated fluidic circuit that uses fluidic logic to select and sequester specific single cells based on a phenotypic trait for downstream experimentation. Experiments with sequestered cells that have been performed include on-chip culture, exposure to various stimulants, and post-exposure image-based response analysis, followed by preparation of the mRNA transcriptome for massively parallel sequencing analysis. The flexible system embodies experimental design and execution that enable routine functional studies of single cells.
A major obstacle to achieving long-term antiretroviral (ART) free remission or functional cure of HIV infection is the presence of persistently infected cells that establish a long-lived viral ...reservoir. HIV largely resides in anatomical regions that are inaccessible to routine sampling, however, and non-invasive methods to understand the longitudinal tissue-wide burden of HIV persistence are urgently needed. Positron emission tomography (PET) imaging is a promising strategy to identify and characterize the tissue-wide burden of HIV. Here, we assess the efficacy of using immunoPET imaging to characterize HIV reservoirs and identify anatomical foci of persistent viral transcriptional activity using a radiolabeled HIV Env-specific broadly neutralizing antibody,
Zr-VRC01, in HIV-infected individuals with detectable viremia and on suppressive ART compared to uninfected controls (NCT03729752). We also assess the relationship between PET tracer uptake in tissues and timing of ART initiation and direct HIV protein expression in CD4 T cells obtained from lymph node biopsies. We observe significant increases in
Zr-VRC01 uptake in various tissues (including lymph nodes and gut) in HIV-infected individuals with detectable viremia (N = 5) and on suppressive ART (N = 5) compared to uninfected controls (N = 5). Importantly, PET tracer uptake in inguinal lymph nodes in viremic and ART-suppressed participants significantly and positively correlates with HIV protein expression measured directly in tissue. Our strategy may allow non-invasive longitudinal characterization of residual HIV infection and lays the framework for the development of immunoPET imaging in a variety of other infectious diseases.
The recruitment of asymptomatic volunteers has been identified as a critical factor that is delaying development and validation of preventive therapies for Alzheimer’s disease (AD). Typical ...recruitment strategies involve the use of convenience samples or soliciting participation of older adults with a family history of AD from clinics and outreach efforts. However, high risk groups, such as ethnic/racial minorities, are traditionally less likely to be recruited for AD prevention studies, thus limiting the ability to generalize findings for a significant proportion of the aging population. A community-engagement approach was used to create a registry of 2,311 research-ready, healthy adult volunteers who reflect the ethnically diverse local community. Furthermore, the registry’s actual commitment to research was examined, through demonstrated participation rates in a clinical study. The approach had varying levels of success in establishing a large, diverse pool of individuals who are interested in participating in pharmacological prevention trials and meet criteria for primary prevention research trials designed to delay the onset of AD. Our efforts suggest that entry criteria for clinical trials need to be carefully considered to be inclusive of African Americans, and that sustained effort is needed to engage African Americans in pharmacological prevention approaches.
According to the study results, 50 maternal-infant pairs had concurrent MRSA colonization: 76% shared isolates of the same pulsed-field type and 30% shared USA300 isolates.
Fungal pathogens such as Candida albicans pose a significant threat to human health with limited treatment options available. One strategy to expand the therapeutic target space is to identify genes ...important for pathogen growth in host-relevant environments. Here, we leverage a pooled functional genomic screening strategy to identify genes important for fitness of C. albicans in diverse conditions. We identify an essential gene with no known Saccharomyces cerevisiae homolog, C1_09670C, and demonstrate that it encodes subunit 3 of replication factor A, Rfa3. Furthermore, we apply computational analyses to identify functionally-coherent gene clusters and predict gene function. Through this approach, we predict the cell cycle-associated function of C3_06880W, a previously uncharacterized genes required for fitness specifically at elevated temperatures, and follow-up assays confirm C3_06880W encodes Iml3, a component of the C. albicans kinetochore with roles in virulence in vivo. Overall, this work reveals insights into the vulnerabilities of C. albicans.
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•Genomic screen reveals genes important for C. albicans growth in diverse conditions•C1_09670C encodes Rfa3 with roles in DNA damage repair•C3_08880W is required for C. albicans fitness and virulence at elevated temperature•C3_08880W encodes Iml3, a component of the inner kinetochore
Xiong et al. perform a functional genomic screen to identify genes important for fitness in diverse environments in the fungal pathogen Candida albicans. Through their investigations, they characterize C1_09670C as subunit 3 of replication factor A (Rfa3), and C3_08880W as kinetochore component Iml3 with important roles in C. albicans virulence.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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