Histone demethylase KDM5 regulates cardiomyocyte maturation by promoting fatty acid oxidation, oxidative phosphorylation, and myofibrillar organization.

AIMS: Human pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) provide a platform to identify and characterize factors that regulate the maturation of CMs. The transition from an immature foetal to an adult CM state entails coordinated regulation of the expression of genes involved in myofibril formation and oxidative phosphorylation (OXPHOS) among others. Lysine demethylase 5 (KDM5) specifically demethylates H3K4me1/2/3 and has emerged as potential regulators of expression of genes involved in cardiac development and mitochondrial function. The purpose of this study is to determine the role of KDM5 in iPSC-CM maturation. METHODS AND RESULTS: KDM5A, B, and C proteins were mainly expressed in the early post-natal stages, and their expressions were progressively downregulated in the post-natal CMs and were absent in adult hearts and CMs. In contrast, KDM5 proteins were persistently expressed in the iPSC-CMs up to 60 days after the induction of myogenic differentiation, consistent with the immaturity of these cells. Inhibition of KDM5 by KDM5-C70 -a pan-KDM5 inhibitor, induced differential expression of 2372 genes, including upregulation of genes involved in fatty acid oxidation (FAO), OXPHOS, and myogenesis in the iPSC-CMs. Likewise, genome-wide profiling of H3K4me3 binding sites by the cleavage under targets and release using nuclease assay showed enriched of the H3K4me3 peaks at the promoter regions of genes encoding FAO, OXPHOS, and sarcomere proteins. Consistent with the chromatin and gene expression data, KDM5 inhibition increased the expression of multiple sarcomere proteins and enhanced myofibrillar organization. Furthermore, inhibition of KDM5 increased H3K4me3 deposits at the promoter region of the ESRRA gene and increased its RNA and protein levels. Knockdown of ESRRA in KDM5-C70-treated iPSC-CM suppressed expression of a subset of the KDM5 targets. In conjunction with changes in gene expression, KDM5 inhibition increased oxygen consumption rate and contractility in iPSC-CMs. CONCLUSION: KDM5 inhibition enhances maturation of iPSC-CMs by epigenetically upregulating the expressions of OXPHOS, FAO, and sarcomere genes and enhancing myofibril organization and mitochondrial function.

Stem-like CD4+ T cells in perivascular tertiary lymphoid structures sustain autoimmune vasculitis.

Autoimmune vasculitis of the medium and large elastic arteries can cause blindness, stroke, aortic arch syndrome, and aortic aneurysm. The disease is often refractory to immunosuppressive therapy and progresses over decades as smoldering aortitis. How the granulomatous infiltrates in the vessel wall are maintained and how tissue-infiltrating T cells and macrophages are replenished are unknown. Single-cell and whole-tissue transcriptomic studies of immune cell populations in vasculitic arteries identified a CD4+ T cell population with stem cell-like features. CD4+ T cells supplying the tissue-infiltrating and tissue-damaging effector T cells survived in tertiary lymphoid structures around adventitial vasa vasora, expressed the transcription factor T cell factor 1 (TCF1), had high proliferative potential, and gave rise to two effector populations, Eomesodermin (EOMES)+ cytotoxic T cells and B cell lymphoma 6 (BCL6)+ T follicular helper-like cells. TCF1hiCD4+ T cells expressing the interleukin 7 receptor (IL-7R) sustained vasculitis in serial transplantation experiments. Thus, TCF1hiCD4+ T cells function as disease stem cells and promote chronicity and autonomy of autoimmune tissue inflammation. Remission-inducing therapies will require targeting stem-like CD4+ T cells instead of only effector T cells.

Huge Abdominopelvic Mass Arising from Cervical Fibroid around an Infected Cervical Diverticulum: A Very Rare Case and a Review of Literature.

Primary uterine diverticula are a very rare congenital anomaly of the uterus with only 21 reported cases. Even rarer is the occurrence of primary cervical diverticula with only six cases reported so far. This is a unique case of a huge abdominopelvic mass arising from cervical fibroid around an infected cervical diverticulum. A 44 year-old, P4L4 came to the OPD with a eighteen weeks size abdomino-pelvic mass. She had a failed surgery 6 months back, attempted to remove the mass. Magnetic resonance imaging revealed a cervical diverticulum which possibly had a pus collection. Relaparotomy was done. It revealed a huge cervical fibroid with dense adhesions all around the mass. A pan hysterectomy was done. In the postoperative period, she developed high-grade fever owing to the development of a pelvic collection, which had to be drained by dilating the vault sutures. Histopathology report confirmed a cervical fibroid with an infected diverticulum within. Primary uterine or cervical diverticula are a very rare anamoly which possibly arise because of a weakness in the area where the two mullerian ducts fuse. Women with this rare condition may suffer from infertility, fever and pain abdomen, acute abdomen owing to torsion or hemoperitoneum, pregnancy complications, and menorrhagia. Diverticulectomy and cervical/uterine reconstruction can be done on nulliparous women while hysterectomy can be offered to perimenopausal women. To conclude, unless known by the gynecologists, radiologists, and the pathologists, this diagnosis can be easily missed out, leading to multiple preventable complications.

Hepatocellular Carcinoma with Hepatic Vein and Inferior Vena Cava Invasion.

Hepatocellular carcinoma (HCC) invades intrahepatic vessels causing tumor thrombosis. Infrequently, there is involvement of the hepatic vein (HV) and inferior vena cava (IVC). In this review, we summarize the epidemiology, classification, clinical features, and management of HCC with HV and IVC invasion. While the involvement of HV and IVC usually portends an overall poor survival, selected patients may be candidates for aggressive treatment and thus improving outcomes.

Elimination of oncogenic KRAS in genetic mouse models eradicates pancreatic cancer by inducing FAS-dependent apoptosis by CD8+ T cells.

Oncogenic KRASG12D (KRAS∗) is critical for the initiation and maintenance of pancreatic ductal adenocarcinoma (PDAC) and is a known repressor of tumor immunity. Conditional elimination of KRAS∗ in genetic mouse models of PDAC leads to the reactivation of FAS, CD8+ T cell-mediated apoptosis, and complete eradication of tumors. KRAS∗ elimination recruits activated CD4+ and CD8+ T cells and promotes the activation of antigen-presenting cells. Mechanistically, KRAS∗-mediated immune evasion involves the epigenetic regulation of Fas death receptor in cancer cells, via methylation of its promoter region. Furthermore, analysis of human RNA sequencing identifies that high KRAS expression in PDAC tumors shows a lower proportion of CD8+ T cells and demonstrates shorter survival compared with tumors with low KRAS expression. This study highlights the role of CD8+ T cells in the eradication of PDAC following KRAS∗ elimination and provides a rationale for the combination of KRAS∗ targeting with immunotherapy to control PDAC.

The AT-hook is an evolutionarily conserved auto-regulatory domain of SWI/SNF required for cell lineage priming.

The SWI/SNF ATP-dependent chromatin remodeler is a master regulator of the epigenome, controlling pluripotency and differentiation. Towards the C-terminus of the catalytic subunit of SWI/SNF is a motif called the AT-hook that is evolutionary conserved. The AT-hook is present in many chromatin modifiers and generally thought to help anchor them to DNA. We observe however that the AT-hook regulates the intrinsic DNA-stimulated ATPase activity aside from promoting SWI/SNF recruitment to DNA or nucleosomes by increasing the reaction velocity a factor of 13 with no accompanying change in substrate affinity (KM). The changes in ATP hydrolysis causes an equivalent change in nucleosome movement, confirming they are tightly coupled. The catalytic subunit's AT-hook is required in vivo for SWI/SNF remodeling activity in yeast and mouse embryonic stem cells. The AT-hook in SWI/SNF is required for transcription regulation and activation of stage-specific enhancers critical in cell lineage priming. Similarly, growth assays suggest the AT-hook is required in yeast SWI/SNF for activation of genes involved in amino acid biosynthesis and metabolizing ethanol. Our findings highlight the importance of studying SWI/SNF attenuation versus eliminating the catalytic subunit or completely shutting down its enzymatic activity.

Single-cell chromatin accessibility profiling of acute myeloid leukemia reveals heterogeneous lineage composition upon therapy-resistance.

Acute myeloid leukemia (AML) is a heterogeneous disease characterized by high rate of therapy resistance. Since the cell of origin can impact response to therapy, it is crucial to understand the lineage composition of AML cells at time of therapy resistance. Here we leverage single-cell chromatin accessibility profiling of 22 AML bone marrow aspirates from eight patients at time of therapy resistance and following subsequent therapy to characterize their lineage landscape. Our findings reveal a complex lineage architecture of therapy-resistant AML cells that are primed for stem and progenitor lineages and spanning quiescent, activated and late stem cell/progenitor states. Remarkably, therapy-resistant AML cells are also composed of cells primed for differentiated myeloid, erythroid and even lymphoid lineages. The heterogeneous lineage composition persists following subsequent therapy, with early progenitor-driven features marking unfavorable prognosis in The Cancer Genome Atlas AML cohort. Pseudotime analysis further confirms the vast degree of heterogeneity driven by the dynamic changes in chromatin accessibility. Our findings suggest that therapy-resistant AML cells are characterized not only by stem and progenitor states, but also by a continuum of differentiated cellular lineages. The heterogeneity in lineages likely contributes to their therapy resistance by harboring different degrees of lineage-specific susceptibilities to therapy.

Dimeric p53 Mutant Elicits Unique Tumor-Suppressive Activities through an Altered Metabolic Program.

Cancer-related alterations of the p53 tetramerization domain (TD) abrogate wild-type (WT) p53 function. They result in a protein that preferentially forms monomers or dimers, which are also normal p53 states under basal cellular conditions. However, their physiologic relevance is not well understood. We have established in vivo models for monomeric and dimeric p53, which model Li-Fraumeni syndrome patients with germline p53 TD alterations. p53 monomers are inactive forms of the protein. Unexpectedly, p53 dimers conferred some tumor suppression that is not mediated by canonical WT p53 activities. p53 dimers upregulate the PPAR pathway. These activities are associated with lower prevalence of thymic lymphomas and increased CD8+ T-cell differentiation. Lymphomas derived from dimeric p53 mice show cooperating alterations in the PPAR pathway, further implicating a role for these activities in tumor suppression. Our data reveal novel functions for p53 dimers and support the exploration of PPAR agonists as therapies. SIGNIFICANCE: New mouse models with TP53R342P (monomer) or TP53A347D (dimer) mutations mimic Li-Fraumeni syndrome. Although p53 monomers lack function, p53 dimers conferred noncanonical tumor-suppressive activities. We describe novel activities for p53 dimers facilitated by PPARs and propose these are "basal" p53 activities. See related commentary by Stieg et al., p. 1046. See related article by Choe et al., p. 1250. This article is highlighted in the In This Issue feature, p. 1027.

Inhibiting androgen receptor splice variants with cysteine-selective irreversible covalent inhibitors to treat prostate cancer.

Androgen receptor (AR) and its splice variants (AR-SVs) promote prostate cancer (PCa) growth by orchestrating transcriptional reprogramming. Mechanisms by which the low complexity and intrinsically disordered primary transactivation domain (AF-1) of AR and AR-SVs regulate transcriptional programming in PCa remains poorly defined. Using omics, live and fixed fluorescent microscopy of cells, and purified AF-1 and AR-V7 recombinant proteins we show here that AF-1 and the AR-V7 splice variant form molecular condensates by liquid-liquid phase separation (LLPS) that exhibit disorder characteristics such as rapid intracellular mobility, coactivator interaction, and euchromatin induction. The LLPS and other disorder characteristics were reversed by a class of small-molecule-selective AR-irreversible covalent antagonists (SARICA) represented herein by UT-143 that covalently and selectively bind to C406 and C327 in the AF-1 region. Interfering with LLPS formation with UT-143 or mutagenesis resulted in chromatin condensation and dissociation of AR-V7 interactome, all culminating in a transcriptionally incompetent complex. Biochemical studies suggest that C327 and C406 in the AF-1 region are critical for condensate formation, AR-V7 function, and UT-143's irreversible AR inhibition. Therapeutically, UT-143 possesses drug-like pharmacokinetics and metabolism properties and inhibits PCa cell proliferation and tumor growth. Our work provides critical information suggesting that clinically important AR-V7 forms transcriptionally competent molecular condensates and covalently engaging C327 and C406 in AF-1, dissolves the condensates, and inhibits its function. The work also identifies a library of AF-1-binding AR and AR-SV-selective covalent inhibitors for the treatment of PCa.

ROLE OF CHANGE IN THE LEVELS OF INFLAMMATORY MARKERS POST DRAINAGE IN PREDICTING OUTCOME IN ACUTE CHOLANGITIS.

BACKGROUND: Acute cholangitis (AC) is a gastro-intestinal emergency associated with significant mortality. Role of change in the levels of inflammatory markers post drainage in predicting outcome in acute cholangitis is uncertain. OBJECTIVE: To evaluate the predictive value of changes in C-reactive protein (CRP) and procalcitonin levels after biliary drainage in relation to outcomes (survival or mortality) at 1 month. METHODS: A prospective observational study of consecutive adults presenting with AC was performed. At admission and at 48 hours post biliary drainage, procalcitonin and CRP were sent. RESULTS: Between August 2020 till December 2020 we recruited 72 consecutive patients of AC. The median age of the patients was 55 years (range 43-62 years) and 42 (58.33%) were females. Although the delta change in serum procalcitonin (P value<0.001) and CRP (P value<0.001) was significant, it had no bearing on the outcome. Altered sensorium and INR were independently associated with mortality at 1 month. The 30-day mortality prediction of day 0 procalcitonin was measured by receiver operating characteristic analysis which resulted in an area under the curve of 0.697 with a 95% confidence interval (95%CI) of 0.545-0.849. The optimal cut-off of procalcitonin would be 0.57ng/mL with a sensitivity and specificity of 80% and 60% respectively to predict mortality. CONCLUSION: Change in serum procalcitonin and CRP levels at 48 hours post drainage although significant, had no impact on the outcome of acute cholangitis.

Papel da mudança nos níveis de marcadores inflamatórios pós drenagem na previsão de resultado em colangite aguda / Role of change in the levels of inflammatory markers post drainage in predicting outcome in acute cholangitis

ABSTRACT Background: Acute cholangitis (AC) is a gastro-intestinal emergency associated with significant mortality. Role of change in the levels of inflammatory markers post drainage in predicting outcome in acute cholangitis is uncertain. Objective: To evaluate the predictive value of changes in C-reactive protein (CRP) and procalcitonin levels after biliary drainage in relation to outcomes (survival or mortality) at 1 month. Methods A prospective observational study of consecutive adults presenting with AC was performed. At admission and at 48 hours post biliary drainage, procalcitonin and CRP were sent. Results: Between August 2020 till December 2020 we recruited 72 consecutive patients of AC. The median age of the patients was 55 years (range 43-62 years) and 42 (58.33%) were females. Although the delta change in serum procalcitonin (P value<0.001) and CRP (P value<0.001) was significant, it had no bearing on the outcome. Altered sensorium and INR were independently associated with mortality at 1 month. The 30-day mortality prediction of day 0 procalcitonin was measured by receiver operating characteristic analysis which resulted in an area under the curve of 0.697 with a 95% confidence interval (95%CI) of 0.545-0.849. The optimal cut-off of procalcitonin would be 0.57ng/mL with a sensitivity and specificity of 80% and 60% respectively to predict mortality. Conclusion: Change in serum procalcitonin and CRP levels at 48 hours post drainage although significant, had no impact on the outcome of acute cholangitis.

RESUMO Contexto: A colangite aguda (CA) é uma emergência gastro-intestinal associada à significativa mortalidade. O papel da mudança nos níveis de marcadores inflamatórios pós drenagem na previsão do desfecho em CA é incerto. Objetivo: Avaliar o valor preditivo das alterações nos níveis de proteína reativa C (PCR) e procalcitonina após drenagem biliar em relação aos desfechos (sobrevida ou mortalidade) em um mês. Métodos Realizou-se estudo observacional prospectivo de adultos consecutivos que apresentam CA. Na admissão e após 48 horas de drenagem biliar, foram analisadas a procalcitonina e a PCR. Resultados Entre agosto de 2020 e dezembro de 2020, foram recrutados 72 pacientes consecutivos de CA. A idade mediana dos pacientes foi de 55 anos (faixa de 43 a 62 anos) e 42 (58,33%) do sexo feminino. Embora a variação delta no soro procalcitonina (valor P<0,001) e PCR (valor P<0,001) tenha sido significativa, não houve influência sobre o resultado. Sensório alterado e INR foram independentemente associados à mortalidade em 1 mês. A previsão de mortalidade de 30 dias no dia 0 da procalcitonina foi medida pela análise característica operacional receptora que resultou em uma área sob a curva de 0,697 com intervalo de confiança de 95% (IC95%) de 0,545-0,849. O corte ideal de procalcitonina seria de 0,57ng/mL com sensibilidade e especificidade de 80% e 60% respectivamente para prever a mortalidade. Conclusão: A mudança nos níveis de procalcitonina sérica e PCR em 48 horas após a drenagem, embora significativa, não teve impacto no resultado da colangite aguda.

Hepatocellular Carcinoma Evaluation and Management for Physicians: Joint Gastroenterology Research Society and Association of Physicians of India Guidelines.

Hepatocellular carcinoma (HCC) is the most common cause of, and accounts for almost 90% of all liver cancers. Data from India is limited especially due to cancer not being a reportable disease and in view of wide variation in diagnostic modalities. This document is a result of a consensus meeting comprising Hepatologists, Interventional Radiologists, Hepatobiliary surgeons, medical and surgical Oncologists nominated by the Association of Physicians of India and Gastroenterology Research Society of Mumbai. The following Clinical Practice Guidelines for practicing physicians is intended to act as an up to date protocol for clinical management of patients with hepatocellular carcinoma. The document comprises seven sections with statements and sub-statements with strength of evidence and recommendation.

Comparative evaluation of Brassica oleracea, Ocimum basilicum, and Moringa oleifera leaf extracts on lipase inhibition and adipogenesis in 3T3-L1 adipocytes.

The anti-adipogenic effects of alcohol extract of Brassica oleracea (BO), Ocimum basilicum (OB), and Moringa oleifera (MO) leaves on 3T3-L1 pre-adipocyte as well as their lipase inhibitory properties were comparatively evaluated. The polyphenol-rich extracts of MO, BO, and OB leaves were profiled for major bioactive compounds using UPLC-HRMS/MS analysis. Among the three plant extracts, BO had the highest flavonoid content with stronger radical scavenging activity. All extracts exhibited lipase inhibitory activities, dose-dependently and a non-competitive type of inhibition with altered kinetic parameters was observed. In cell culture studies, these plant extracts remarkably inhibited the intracellular triglyceride accumulation in 3T3-L1 cells and the effect was in the decreasing order of BO > OB > MO. The extracts also down-regulated the key transcription factors of adipogenesis (PPARγ, C/EPB-α, PI3k, p-Akt, and FAS enzyme) as well as modulated the release profile of leptin and adiponectins. Overall, BO extract showed an exceptional inhibitory potential against both adipogenesis and lipase activities among the evaluated plants. PRACTICAL APPLICATIONS: The present study highlights the anti-adipogenic potential of three local edible plants and herbs, that is, Brassica oleracea (BO), Ocimum basilicum (OB), and Moringa oleifera (MO) leaves, using in vitro models. The results revealed that the BO extract had remarkable activity against adipogenesis in the 3T3-L1 pre-adipocyte differentiation model as well as lipase inhibitory properties. This study elucidates the prospects of natural plant-derived compounds in managing obesity-related health issues. The outcomes of this study would be useful in developing alternative or complementary therapies that are being preferred and largely sought over pharmacological treatments and procedures for controlling and treating abnormal weight gain in humans.

Targeting the NOTCH1-MYC-CD44 axis in leukemia-initiating cells in T-ALL.

The NOTCH1-MYC-CD44 axis integrates cell-intrinsic and extrinsic signaling to ensure the persistence of leukemia-initiating cells (LICs) in T-cell acute lymphoblastic leukemia (T-ALL) but a common pathway to target this circuit is poorly defined. Bromodomain-containing protein 4 (BRD4) is implicated to have a role in the transcriptional regulation of oncogenes MYC and targets downstream of NOTCH1, and here we demonstrate its role in transcriptional regulation of CD44. Hence, targeting BRD4 will dismantle the NOTCH1-MYC-CD44 axis. As a proof of concept, degrading BRD4 with proteolysis targeting chimera (PROTAC) ARV-825, prolonged the survival of mice in Notch1 mutated patient-derived xenograft (PDX) and genetic models (ΔPTEN) of T-ALL. Single-cell proteomics analysis from the PDX model, demonstrated quantitative reduction of LICs (CD34+ CD7+ CD19-) and downregulation of the NOTCH1-MYC-CD44 axis, along with cell cycle, apoptosis and PI3K/Akt pathways. Moreover, secondary transplantation from PDX and ΔPTEN models of T-ALL, confirmed delayed leukemia development and extended survival of mice engrafted with T-ALL from ARV-825 treated mice, providing functional confirmation of depletion of LICs. Hence, BRD4 degradation is a promising LIC-targeting therapy for T-ALL.

Spatio-temporal dynamics of intra-host variability in SARS-CoV-2 genomes.

During the course of the COVID-19 pandemic, large-scale genome sequencing of SARS-CoV-2 has been useful in tracking its spread and in identifying variants of concern (VOC). Viral and host factors could contribute to variability within a host that can be captured in next-generation sequencing reads as intra-host single nucleotide variations (iSNVs). Analysing 1347 samples collected till June 2020, we recorded 16 410 iSNV sites throughout the SARS-CoV-2 genome. We found ∼42% of the iSNV sites to be reported as SNVs by 30 September 2020 in consensus sequences submitted to GISAID, which increased to ∼80% by 30th June 2021. Following this, analysis of another set of 1774 samples sequenced in India between November 2020 and May 2021 revealed that majority of the Delta (B.1.617.2) and Kappa (B.1.617.1) lineage-defining variations appeared as iSNVs before getting fixed in the population. Besides, mutations in RdRp as well as RNA-editing by APOBEC and ADAR deaminases seem to contribute to the differential prevalence of iSNVs in hosts. We also observe hyper-variability at functionally critical residues in Spike protein that could alter the antigenicity and may contribute to immune escape. Thus, tracking and functional annotation of iSNVs in ongoing genome surveillance programs could be important for early identification of potential variants of concern and actionable interventions.

Clinical, immunological and genomic characteristics of children with X-linked agammaglobulinemia from Kerala, South India.

X-linked agammaglobulinemia (XLA) is an X-linked recessive primary immunodeficiency disorder caused due to a pathogenic variant in the Bruton tyrosine (BTK) gene with an incidence of 1:379,000 live births and 1:190,000 male births. Patients affected with XLA present with recurrent infections of the gastrointestinal and respiratory tracts. Here we report the first case series of 17 XLA patients of 10 South Indian families with a wide spectrum of clinical and genetic features. In our cohort, patients presented mainly with recurrent pneumonia, gastrointestinal infection, otitis media, pyoderma, abscesses, empyema, arthritis, and osteomyelitis. Using next-generation and Sanger sequencing we have identified 10 unique pathogenic and likely pathogenic variants in 17 patients. This encompasses three nonsynonymous, two stop-gain, two frameshifts, two structural, and one splicing variant, out of which two of them are novel. Based on the type of variant, patients had variable clinical features and treatment responses. We have also evaluated Btk protein expression for six patients in comparison to the healthy individuals and determined mosaic Btk expression patterns in four mothers. We have also performed family screening in 6 families using Sanger sequencing and identified 19 carriers for the variant. The diagnosis for the patients led to the proper treatment i.e. 15 patients were on intravenous immunoglobulin (IVIG) and the other two had successful hematopoietic stem cell transplantation (HSCT). Unfortunately, two of our patients died due to sepsis, while on IVIG. We envision the present study could help in better understanding of patients with XLA and help in family screening and prenatal diagnosis. To the best of our knowledge, this is the largest case series of patients affected with XLA from South India.

Mammary-specific expression of Trim24 establishes a mouse model of human metaplastic breast cancer.

Conditional overexpression of histone reader Tripartite motif containing protein 24 (TRIM24) in mouse mammary epithelia (Trim24COE) drives spontaneous development of mammary carcinosarcoma tumors, lacking ER, PR and HER2. Human carcinosarcomas or metaplastic breast cancers (MpBC) are a rare, chemorefractory subclass of triple-negative breast cancers (TNBC). Comparison of Trim24COE metaplastic carcinosarcoma morphology, TRIM24 protein levels and a derived Trim24COE gene signature reveals strong correlation with human MpBC tumors and MpBC patient-derived xenograft (PDX) models. Global and single-cell tumor profiling reveal Met as a direct oncogenic target of TRIM24, leading to aberrant PI3K/mTOR activation. Here, we find that pharmacological inhibition of these pathways in primary Trim24COE tumor cells and TRIM24-PROTAC treatment of MpBC TNBC PDX tumorspheres decreased cellular viability, suggesting potential in therapeutically targeting TRIM24 and its regulated pathways in TRIM24-expressing TNBC.

P4HA2-induced prolyl hydroxylation suppresses YAP1-mediated prostate cancer cell migration, invasion, and metastasis.

Yes-associated protein 1 (YAP1), a key player in the Hippo pathway, has been shown to play a critical role in tumor progression. However, the role of YAP1 in prostate cancer cell invasion, migration, and metastasis is not well defined. Through functional, transcriptomic, epigenomic, and proteomic analyses, we showed that prolyl hydroxylation of YAP1 plays a critical role in the suppression of cell migration, invasion, and metastasis in prostate cancer. Knockdown (KD) or knockout (KO) of YAP1 led to an increase in cell migration, invasion, and metastasis in prostate cancer cells. Microarray analysis showed that the EMT pathway was activated in Yap1-KD cells. ChIP-seq analysis showed that YAP1 target genes are enriched in pathways regulating cell migration. Mass spectrometry analysis identified P4H prolyl hydroxylase in the YAP1 complex and YAP1 was hydroxylated at multiple proline residues. Proline-to-alanine mutations of YAP1 isoform 3 identified proline 174 as a critical residue, and its hydroxylation suppressed cell migration, invasion, and metastasis. KO of P4ha2 led to an increase in cell migration and invasion, which was reversed upon Yap1 KD. Our study identified a novel regulatory mechanism of YAP1 by which P4HA2-dependent prolyl hydroxylation of YAP1 determines its transcriptional activities and its function in prostate cancer metastasis.

Whole genome sequencing identifies novel structural variant in a large Indian family affected with X-linked agammaglobulinemia.

X-linked agammaglobulinemia (XLA, OMIM #300755) is a primary immunodeficiency disorder caused by pathogenic variations in the BTK gene, characterized by failure of development and maturation of B lymphocytes. The estimated prevalence worldwide is 1 in 190,000 male births. Recently, genome sequencing has been widely used in difficult to diagnose and familial cases. We report a large Indian family suffering from XLA with five affected individuals. We performed complete blood count, immunoglobulin assay, and lymphocyte subset analysis for all patients and analyzed Btk expression for one patient and his mother. Whole exome sequencing (WES) for four patients, and whole genome sequencing (WGS) for two patients have been performed. Carrier screening was done for 17 family members using Multiplex Ligation-dependent Probe Amplification (MLPA) and haplotype ancestry mapping using fineSTRUCTURE was performed. All patients had hypogammaglobulinemia and low CD19+ B cells. One patient who underwent Btk estimation had low expression and his mother showed a mosaic pattern. We could not identify any single nucleotide variants or small insertion/ deletions from the WES dataset that correlates with the clinical feature of the patient. Structural variant analysis through WGS data identifies a novel large deletion of 5,296 bp at loci chrX:100,624,323-100,629,619 encompassing exons 3-5 of the BTK gene. Family screening revealed seven carriers for the deletion. Two patients had a successful HSCT. Haplotype mapping revealed a South Asian ancestry. WGS led to identification of the accurate genetic mutation which could help in early diagnosis leading to improved outcomes, prevention of permanent organ damage and improved quality of life, as well as enabling genetic counselling and prenatal diagnosis in the family.

Six artificial intelligence paradigms for tissue characterisation and classification of non-COVID-19 pneumonia against COVID-19 pneumonia in computed tomography lungs.

BACKGROUND: COVID-19 pandemic has currently no vaccines. Thus, the only feasible solution for prevention relies on the detection of COVID-19-positive cases through quick and accurate testing. Since artificial intelligence (AI) offers the powerful mechanism to automatically extract the tissue features and characterise the disease, we therefore hypothesise that AI-based strategies can provide quick detection and classification, especially for radiological computed tomography (CT) lung scans. METHODOLOGY: Six models, two traditional machine learning (ML)-based (k-NN and RF), two transfer learning (TL)-based (VGG19 and InceptionV3), and the last two were our custom-designed deep learning (DL) models (CNN and iCNN), were developed for classification between COVID pneumonia (CoP) and non-COVID pneumonia (NCoP). K10 cross-validation (90% training: 10% testing) protocol on an Italian cohort of 100 CoP and 30 NCoP patients was used for performance evaluation and bispectrum analysis for CT lung characterisation. RESULTS: Using K10 protocol, our results showed the accuracy in the order of DL > TL > ML, ranging the six accuracies for k-NN, RF, VGG19, IV3, CNN, iCNN as 74.58 ± 2.44%, 96.84 ± 2.6, 94.84 ± 2.85%, 99.53 ± 0.75%, 99.53 ± 1.05%, and 99.69 ± 0.66%, respectively. The corresponding AUCs were 0.74, 0.94, 0.96, 0.99, 0.99, and 0.99 (p-values < 0.0001), respectively. Our Bispectrum-based characterisation system suggested CoP can be separated against NCoP using AI models. COVID risk severity stratification also showed a high correlation of 0.7270 (p < 0.0001) with clinical scores such as ground-glass opacities (GGO), further validating our AI models. CONCLUSIONS: We prove our hypothesis by demonstrating that all the six AI models successfully classified CoP against NCoP due to the strong presence of contrasting features such as ground-glass opacities (GGO), consolidations, and pleural effusion in CoP patients. Further, our online system takes < 2 s for inference.