Abnormal global longitudinal strain and reduced serum inflammatory markers in cardiac AL amyloidosis patients without significant amyloid fibril deposition.

Background: Cardiac dysfunction in AL amyloidosis is thought to be partly related to the direct impact of AL LCs on cardiomyocyte function, with the degree of dysfunction at diagnosis as a major determinant of clinical outcomes. Nonetheless, mechanisms underlying LC-induced myocardial toxicity are not well understood. Methods: We identified gene expression changes correlating with human cardiac cells exposed to a cardiomyopathy-associated κAL LC. We then sought to confirm these findings in a clinical dataset by focusing on clinical parameters associated with the pathways dysregulated at the gene expression level. Results: Upon exposure to a cardiomyopathy-associated κAL LC, cardiac cells exhibited gene expression changes related to myocardial contractile function and inflammation, leading us to hypothesize that there could be clinically detectable changes in GLS on echocardiogram and serum inflammatory markers in patients. Thus, we identified 29 patients with normal IVSd but abnormal cardiac biomarkers suggestive of LC-induced cardiac dysfunction. These patients display early cardiac biomarker staging, abnormal GLS, and significantly reduced serum inflammatory markers compared to patients with clinically evident amyloid fibril deposition. Conclusion: Collectively, our findings highlight early molecular and functional signatures of cardiac AL amyloidosis, with potential impact for developing improved patient biomarkers and novel therapeutics.

Whole-exome sequencing of Indian prostate cancer reveals a novel therapeutic target: POLQ.

PURPOSE: Prostate cancer is the second most common cancer diagnosed worldwide and the third most common cancer among men in India. This study's objective was to characterise the mutational landscape of Indian prostate cancer using whole-exome sequencing to identify population-specific polymorphisms. METHODS: Whole-exome sequencing was performed of 58 treatment-naive primary prostate tumors of Indian origin. Multiple computational and statistical analyses were used to profile the known common mutations, other deleterious mutations, driver genes, prognostic biomarkers, and gene signatures unique to each clinical parameter. Cox analysis was performed to validate survival-associated genes. McNemar test identified genes significant to recurrence and receiver-operating characteristic (ROC) analysis was conducted to determine its accuracy. OncodriveCLUSTL algorithm was used to deduce driver genes. The druggable target identified was modeled with its known inhibitor using Autodock. RESULTS: TP53 was the most commonly mutated gene in our cohort. Three novel deleterious variants unique to the Indian prostate cancer subtype were identified: POLQ, FTHL17, and OR8G1. COX regression analysis identified ACSM5, a mitochondrial gene responsible for survival. CYLC1 gene, which encodes for sperm head cytoskeletal protein, was identified as an unfavorable prognostic biomarker indicative of recurrence. The novel POLQ mutant, also identified as a driver gene, was evaluated as the druggable target in this study. POLQ, a DNA repair enzyme implicated in various cancer types, is overexpressed and is associated with a poor prognosis. The mutant POLQ was subjected to structural analysis and modeled with its known inhibitor novobiocin resulting in decreased binding efficiency necessitating the development of a better drug. CONCLUSION: In this pilot study, the molecular profiling using multiple computational and statistical analyses revealed distinct polymorphisms in the Indian prostate cancer cohort. The mutational signatures identified provide a valuable resource for prognostic stratification and targeted treatment strategies for Indian prostate cancer patients. The DNA repair enzyme, POLQ, was identified as the druggable target in this study.

Multidimensional Mutational Profiling of the Indian HNSCC Sub-Population Provides IRAK1, a Novel Driver Gene and Potential Druggable Target.

Head and neck squamous cell carcinomas (HNSCC) include heterogeneous group of tumors, classified according to their anatomical site. It is the sixth most prevalent cancer globally. Among South Asian countries, India accounts for 40% of HNC malignancies with significant morbidity and mortality. In the present study, we have performed exome sequencing and analysis of 51 Head and Neck squamous cell carcinoma samples. Besides known mutations in the oncogenes and tumour suppressors, we have identified novel gene signatures differentiating buccal, alveolar, and tongue cancers. Around 50% of the patients showed mutation in tumour suppressor genes TP53 and TP63. Apart from the known mutations, we report novel mutations in the genes AKT1, SPECC1, and LRP1B, which are linked with tumour progression and patient survival. A highly curated process was developed to identify survival signatures. 36 survival-related genes were identified based on the correlation of functional impact of variants identified using exome-seq with gene expression from transcriptome data (GEPIA database) and survival. An independent LASSO regression analysis was also performed. Survival signatures common to both the methods led to identification of 4 dead and 3 alive gene signatures, the accuracy of which was confirmed by performing a ROC analysis (AUC=0.79 and 0.91, respectively). Also, machine learning-based driver gene prediction tool resulted in the identification of IRAK1 as the driver (p-value = 9.7 e-08) and also as an actionable mutation. Modelling of the IRAK1 mutation showed a decrease in its binding to known IRAK1 inhibitors.

hg19KIndel: ethnicity normalized human reference genome.

BACKGROUND: The most widely used human genome reference assembly hg19 harbors minor alleles at 2.18 million positions as revealed by 1000 Genome Phase 3 dataset. Although this is less than 2% of the 89 million variants reported, it has been shown that the minor alleles can result in 30% false positives in individual genomes, thus misleading and burdening downstream interpretation. More alarming is the fact that, significant percentage of variants that are homozygous recessive for these minor alleles, with potential disease implications, are masked from reporting. RESULTS: We have demonstrated that the false positives (FP) and false negatives (FN) can be corrected for by simply replacing nucleotides at the minor allele positions in hg19 with corresponding major allele. Here, we have effectively replaced 2.18 million minor alleles Single Nucleotide Polymorphism (SNPs), Insertion and Deletions (INDELs), Multiple Nucleotide Polymorphism (MNPs) in hg19 with the corresponding major alleles to create an ethnically normalized reference genome called hg19KIndel. In doing so, hg19KIndel has both corrected for sequencing errors acknowledged to be present in hg19 and has improved read alignment near the minor alleles in hg19. CONCLUSION: We have created and made available a new version human reference genome called hg19KIndel. It has been shown that variant calling using hg19KIndel, significantly reduces false positives calls, which in-turn reduces the burden from downstream analysis and validation. It also improved false negative variants call, which means that the variants which were getting missed due to the presence of minor alleles in hg19, will now be called using hg19KIndel. Using hg19KIndel, one even gets a better mapping percentage when compared to currently available human reference genome. hg19KIndel reference genome and its auxiliary datasets are available at https://doi.org/10.5281/zenodo.2638113.

Early splicing functions of fission yeast Prp16 and its unexpected requirement for gene Silencing is governed by intronic features.

Prp16 is a DEAH box pre-mRNA splicing factor that triggers a key spliceosome conformational switch to facilitate second step splicing in Saccharomyces cerevisiae. However, Prp16 functions are largely unexplored in Schizosaccharomyces pombe, an attractive model with exon-intron architecture more relevant to several other eukaryotes. Here, we generated mis-sense alleles in SpPrp16 whose consequences on genome-wide splicing uncover its nearly global splicing role with only a small subset of unaffected introns. Prp16 dependent and independent intron categories displayed a striking difference in the strength of intronic 5' splice site (5'SS)-U6 snRNA and branch site (BS)-U2 snRNA interactions. Selective weakening of these interactions could convert a Prp16 dependent intron into an independent one. These results point to the role of SpPrp16 in destabilizing 5'SS-U6snRNA and BS-U2snRNA interactions which plausibly trigger structural alterations in the spliceosome to facilitate first step catalysis. Our data suggest that SpPrp16 interactions with early acting factors, its enzymatic activities and association with intronic elements collectively account for efficient and accurate first step catalysis. In addition to splicing derangements in the spprp16F528S mutant, we show that SpPrp16 influences cell cycle progression and centromeric heterochromatinization. We propose that strong 5'SS-U6 snRNA and BS-U2 snRNA complementarity of intron-like elements in non-coding RNAs which lead to complete splicing arrest and impaired Seb1 functions at the pericentromeric loci may cumulatively account for the heterochromatin defects in spprp16F528S cells. These findings suggest that the diverse Prp16 functions within a genome are likely governed by its intronic features that influence splice site-snRNA interaction strength.

A novel molecular mechanism for a long non-coding RNA PCAT92 implicated in prostate cancer.

The role of many lncRNAs in cancer remains elusive including that for a Prostate Cancer Associated Transcript 92 (PCAT92). PCAT92 shares the locus on chromosome 13 with ABCC4 gene, known to be implicated in prostate cancer. It has been shown that PCAT92 and ABCC4 are up-regulated in prostate cancer samples from multiple transcriptome datasets. Among the prostate cancer cell-lines LNCaP showed maximum overexpression of PCAT92 compared to control cell-line RWPE-1. We have shown that knockdown of PCAT92 in LNCaP cells reduces cell viability and proliferation and down-regulates ABCC4 transcript/protein expression. The shared region between PCAT92 and ABCC4 has a binding site for an oncogenic transcription factor (ZIC2) which is also upregulated in the majority of datasets studied here. ZIC2 binding to the predicted ABCC4 promoter has been confirmed using pull-down assay. Interestingly, under PCAT92 knockdown condition, there is a reduction in the ZIC2 binding to ABCC4 promoter indicating the potential involvement of PCAT92 in the recruitment of ZIC2. We have identified distinct regions on PCAT92 with potential to bind to ZIC2 non-DNA binding Zinc-finger domain and potential for triplex formation near ABCC4 promoter region, which have been experimentally validated. Together, these observations and localization in the nucleus suggests that PCAT92 may play a role in prostate cancer by increasing the local concentration of ZIC2 by forming RNA-DNA triplex near ABCC4 promoter thus helping in recruitment of ZIC2 for ABCC4 regulation.