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Background Recent massive sequencing studies have revealed that SWI/SNF complexes are among the most frequently altered functional entities in solid tumors. However, the role of SWI/SNF in acute myeloid leukemia is poorly understood. To date, SWI/SNF complexes are thought to be oncogenic in AML or, at least, necessary to support leukemogenesis. However, mutation patterns in SWI/SNF genes in AML are consistent with a tumor suppressor role. Here, we study the SWI/SNF subunit BCL7A, which has been found to be recurrently mutated in lymphomas, but whose role in acute myeloid malignancies is currently unknown. Methods Data mining and bioinformatic approaches were used to study the mutational status of BCL7A and the correlation between BCL7A expression and promoter hypermethylation. Methylation-specific PCR, bisulfite sequencing, and 5-aza-2'-deoxycytidine treatment assays were used to determine if BCL7A expression was silenced due to promoter hypermethylation. Cell competition assays after BCL7A expression restoration were used to assess the role of BCL7A in AML cell line models. Differential expression analysis was performed to determine pathways and genes altered after BCL7A expression restoration. To establish the role of BCL7A in tumor development in vivo, tumor growth was compared between BCL7A-expressing and non-expressing mouse xenografts using in vivo fluorescence imaging . Results BCL7A expression was inversely correlated with promoter methylation in three external cohorts: TCGA-LAML ( N  = 160), TARGET-AML ( N  = 188), and Glass et al. (2017) ( N  = 111). The AML-derived cell line NB4 silenced the BCL7A expression via promoter hypermethylation. Ectopic BCL7A expression in AML cells decreased their competitive ability compared to control cells. Additionally, restoration of BCL7A expression reduced tumor growth in an NB4 mouse xenograft model. Also, differential expression analysis found that BCL7A restoration altered cell cycle pathways and modified significantly the expression of genes like HMGCS1, H1-0 , and IRF7 which can help to explain its tumor suppressor role in AML. Conclusions BCL7A expression is silenced in AML by promoter methylation. In addition, restoration of BCL7A expression exerts tumor suppressor activity in AML cell lines and xenograft models.

Mutations in genes encoding subunits of the SWI/SNF chromatin remodeling complex are frequently found in different human cancers. While the tumor suppressor function of this complex is widely established in solid tumors, its role in hematologic malignancies is largely unknown. Recurrent point mutations in BCL7A gene, encoding a subunit of the SWI/SNF complex, have been reported in diffuse large B-cell lymphoma (DLBCL), but their functional impact remains to be elucidated. Here we show that BCL7A often undergoes biallelic inactivation, including a previously unnoticed mutational hotspot in the splice donor site of intron one. The splice site mutations render a truncated BCL7A protein, lacking a portion of the amino-terminal domain. Moreover, restoration of wild-type BCL7A expression elicits a tumor suppressor-like phenotype in vitro and in vivo. In contrast, splice site mutations block the tumor suppressor function of BCL7A by preventing its binding to the SWI/SNF complex. We also show that BCL7A restoration induces transcriptomic changes in genes involved in B-cell activation. In addition, we report that SWI/SNF complex subunits harbor mutations in more than half of patients with germinal center B-cell (GCB)-DLBCL. Overall, this work demonstrates the tumor suppressor function of BCL7A in DLBCL, and highlights that the SWI/SNF complex plays a relevant role in DLBCL pathogenesis.

Pediatric acute B-cell lymphoblastic leukemia (B-ALL) constitutes a heterogeneous and aggressive neoplasia in which new targeted therapies are required. Long non-coding RNAs have recently emerged as promising disease-specific biomarkers for the clinic. Here, we identified pediatric B-ALL-specific lncRNAs and associated mRNAs by comparing the transcriptomic signatures of tumoral and non-tumoral samples. We identified 48 lncRNAs that were differentially expressed between pediatric B-ALL and healthy bone marrow samples. The most relevant lncRNA/mRNA pair was AL133346.1/CCN2 (previously known as RP11-69I8.3/CTGF), whose expression was positively correlated and increased in B-ALL samples. Their differential expression pattern and their strong correlation were validated in external B-ALL datasets (Therapeutically Applicable Research to Generate Effective Treatments, Cancer Cell Line Encyclopedia). Survival curve analysis demonstrated that patients with “high” expression levels of CCN2 had higher overall survival than those with “low” levels (p = 0.042), and this gene might be an independent prognostic biomarker in pediatric B-ALL. These findings provide one of the first detailed descriptions of lncRNA expression profiles in pediatric B-ALL and indicate that these potential biomarkers could help in the classification of leukemia subtypes and that CCN2 expression could predict the survival outcome of pediatric B-cell acute lymphoblastic leukemia patients.

Mammalian SWI/SNF (SWitch/Sucrose Non-Fermentable) complexes are ATP-dependent chromatin remodelers whose subunits have emerged among the most frequently mutated genes in cancer. Studying SWI/SNF function in cancer cell line models has unveiled vulnerabilities in SWI/SNF-mutant tumors that can lead to the discovery of new therapeutic drugs. However, choosing an appropriate cancer cell line model for SWI/SNF functional studies can be challenging because SWI/SNF subunits are frequently altered in cancer by various mechanisms, including genetic alterations and post-transcriptional mechanisms. In this work, we combined genomic, transcriptomic, and proteomic approaches to study the mutational status and the expression levels of the SWI/SNF subunits in a panel of 38 lung adenocarcinoma (LUAD) cell lines. We found that the SWI/SNF complex was mutated in more than 76% of our LUAD cell lines and there was a high variability in the expression of the different SWI/SNF subunits. These results underline the importance of the SWI/SNF complex as a tumor suppressor in LUAD and the difficulties in defining altered and unaltered cell models for the SWI/SNF complex. These findings will assist researchers in choosing the most suitable cellular models for their studies of SWI/SNF to bring all of its potential to the development of novel therapeutic applications.

Missense mutations in the 12 th codon of KRAS are key drivers of lung cancer, with glycine-to-cysteine (G12C) and glycine-to-aspartic acid (G12D) substitutions being among the most prevalent. These mutations are strongly associated with poor survival outcomes. Given the critical role of KRAS in lung cancer and other cancers, it remains as a major target for the development of new and complementary treatments. We have developed a CRISPR-High Fidelity (HiFi)-Cas9-based therapy strategy that can effectively and specifically target KRAS G12C and KRAS G12D mutants, avoiding KRAS WT off-targeting and affecting KRAS downstream pathways, thereby significantly reducing tumorgenicity. The delivery of HiFiCas9 components via ribonucleoprotein particles (RNPs) and adenovirus (AdV) effectively abrogates cell viability in KRAS -mutant Non-Small Cell Lung Cancer (NSCLC) preclinical models, including 2D and 3D cell cultures, cell-derived xenografts (CDX), and patient-derived xenograft organoids (PDXO). Our in vitro studies demonstrate that HiFiCas9-based therapy achieves superior KRAS inhibition compared to Sotorasib and effectively circumvents certain resistance mechanisms associated with Sotorasib treatment. Moreover, in vivo delivery using adenoviral particles significantly suppresses tumor growth in preclinical NSCLC models. Collectively, our findings establish HiFiCas9 as an effective therapeutic strategy with promising clinical applications, especially if in vivo delivery methods are further optimized. Missense mutations in the twelfth codon of KRAS are key drivers of lung cancer. Here, the authors develop a CRISPR-High Fidelity-Cas9-based strategy to target KRAS-G12C and KRAS-G12D mutants, reducing tumourigenicity without wild type KRAS off-targeting and circumventing certain therapy resistance mechanisms in preclinical models.

A high-throughput chemical screen reveals that harmine and its analogs promote improved human pancreatic beta cell replication and function, thus identifying these molecules as a potential new class of antidiabetic agents. Types 1 and 2 diabetes affect some 380 million people worldwide. Both ultimately result from a deficiency of functional pancreatic insulin-producing beta cells. Beta cells proliferate in humans during a brief temporal window beginning around the time of birth, with a peak percentage (∼2%) engaged in the cell cycle in the first year of life 1 , 2 , 3 , 4 . In embryonic life and after early childhood, beta cell replication is barely detectable. Whereas beta cell expansion seems an obvious therapeutic approach to beta cell deficiency, adult human beta cells have proven recalcitrant to such efforts 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 . Hence, there remains an urgent need for antidiabetic therapeutic agents that can induce regeneration and expansion of adult human beta cells in vivo or ex vivo . Here, using a high-throughput small-molecule screen (HTS), we find that analogs of the small molecule harmine function as a new class of human beta cell mitogenic compounds. We also define dual-specificity tyrosine-regulated kinase-1a (DYRK1A) as the likely target of harmine and the nuclear factors of activated T cells (NFAT) family of transcription factors as likely mediators of human beta cell proliferation and differentiation. Using three different mouse and human islet in vivo –based models, we show that harmine is able to induce beta cell proliferation, increase islet mass and improve glycemic control. These observations suggest that harmine analogs may have unique therapeutic promise for human diabetes therapy. Enhancing the potency and beta cell specificity of these compounds are important future challenges.

Reversible transition between the epithelial and mesenchymal states are key aspects of carcinoma cell dissemination and the metastatic disease, and thus, characterizing the molecular basis of the epithelial to mesenchymal transition (EMT) is crucial to find druggable targets and more effective therapeutic approaches in cancer. Emerging studies suggest that epigenetic regulators might endorse cancer cells with the cell plasticity required to conduct dynamic changes in cell state during EMT. However, epigenetic mechanisms involved remain mostly unknown. Polycomb Repressive Complexes (PRCs) proteins are well-established epigenetic regulators of development and stem cell differentiation, but their role in different cancer systems is inconsistent and sometimes paradoxical. In this study, we have analysed the role of the PRC2 protein EZH2 in lung carcinoma cells. We found that besides its described role in CDKN2A-dependent cell proliferation, EZH2 upholds the epithelial state of cancer cells by repressing the transcription of hundreds of mesenchymal genes. Chemical inhibition or genetic removal of EZH2 promotes the residence of cancer cells in the mesenchymal state during reversible epithelial-mesenchymal transition. In fitting, analysis of human patient samples and tumour xenograft models indicate that EZH2 is required to efficiently repress mesenchymal genes and facilitate tumour colonization in vivo. Overall, this study discloses a novel role of PRC2 as a master regulator of EMT in carcinoma cells. This finding has important implications for the design of therapies based on EZH2 inhibitors in human cancer patients.

Plakophilin 1 (PKP1) is a member of the arm-repeat (armadillo) and plakophilin gene families and it is an essential component of the desmosomes. Although desmosomes have generally been associated with tumor suppressor functions, we have consistently observed that PKP1 is among the top overexpressed proteins in squamous cell lung cancer. To explore this paradox, we developed in vivo and in vitro functional models of PKP1 gain/loss in squamous cell lung cancer. CRISPR-Cas9 PKP1 knockout severely impaired cell proliferation, but it increased cell dissemination. In addition, PKP1 overexpression increased cell proliferation, cell survival, and in vivo xenograft engraftment. We further investigated the molecular mechanism of the mainly oncogenic function of PKP1 by combining transcriptomics, proteomics, and protein-nucleic acid interaction assays. Interestingly, we found that PKP1 enhances MYC translation in collaboration with the translation initiation complex by binding to the 5′-UTR of MYC mRNA. We propose PKP1 as an oncogene in SqCLC and a novel posttranscriptional regulator of MYC. PKP1 may be a valuable diagnostic biomarker and potential therapeutic target for SqCLC. Importantly, PKP1 inhibition may indirectly target MYC, a primary anticancer target.

Hematological malignancies are a highly heterogeneous group of diseases with varied molecular and phenotypical characteristics. SWI/SNF (SWItch/Sucrose Non-Fermentable) chromatin remodeling complexes play significant roles in the regulation of gene expression, being essential for processes such as cell maintenance and differentiation in hematopoietic stem cells. Furthermore, alterations in SWI/SNF complex subunits, especially in ARID1A/1B/2, SMARCA2/4, and BCL7A, are highly recurrent across a wide variety of lymphoid and myeloid malignancies. Most genetic alterations cause a loss of function of the subunit, suggesting a tumor suppressor role. However, SWI/SNF subunits can also be required for tumor maintenance or even play an oncogenic role in certain disease contexts. The recurrent alterations of SWI/SNF subunits highlight not only the biological relevance of SWI/SNF complexes in hematological malignancies but also their clinical potential. In particular, increasing evidence has shown that mutations in SWI/SNF complex subunits confer resistance to several antineoplastic agents routinely used for the treatment of hematological malignancies. Furthermore, mutations in SWI/SNF subunits often create synthetic lethality relationships with other SWI/SNF or non-SWI/SNF proteins that could be exploited therapeutically. In conclusion, SWI/SNF complexes are recurrently altered in hematological malignancies and some SWI/SNF subunits may be essential for tumor maintenance. These alterations, as well as their synthetic lethal relationships with SWI/SNF and non-SWI/SNF proteins, may be pharmacologically exploited for the treatment of diverse hematological cancers.