Applications of CRISPR/Cas technology against drug-resistant lung cancers: an update.

Out of all the cancer types, the most prevalent one is lung cancer. Multiple genes and signaling pathways play role in the progression of lung cancer. Considering the wider prevalence and fatality of lung cancer it has become the focus of current cancer research. Though currently used approaches have shown positive results against lung cancer but success against non-small cell lung cancer (NSCLC) still looms as an enigma for the entire research fraternity. The development of resistance against inhibitors within a short span is one of the reasons responsible for the failure and relapse of lung cancer. Under these prevailing conditions genome/gene-editing technology using clustered regularly interspaced short palindromic repeat (CRISPR) and CRISPR associated proteins (Cas), popularly known as CRISPR/Cas technology offers a convenient and flexible method for inducing precise changes within the lung cancer cell. Additionally, CRISPR-barcoding and CRISPR knockout screens at the genome-wide level can help in the functional investigation of specific mutations and identification of novel cancer drivers respectively. Several variants of the CRISPR/Cas system are being developed to limit off-targeting with enhanced precision. The present review article updates the usefulness of CRISPR/Cas technology against various types of lung cancers.

CRISPR/Cas technology for improving nutritional values in the agricultural sector: an update.

BACKGROUND: The CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) system was initially identified in bacteria and archaea as a defense mechanism to confer immunity against phages. Later on, it was developed as a gene editing tool for both prokaryotic and eukaryotic cells including plant cells. METHODS AND RESULTS: CRISPR/Cas9 approach has wider applications in reverse genetics as well as in crop improvement. Various characters involved in enhancing economic value and crop sustainability against biotic/abiotic stresses can be targeted through this tool. Currently, CRISPR/Cas9 gene editing mechanism has been applied on around 20 crop species for improvement in several traits including yield enhancement and resistance against biotic and abiotic stresses. In the last five years, maximum genome editing research has been validated in rice, wheat, maize and soybean. Genes targeted in these plants has been involved in causing male sterility, conferring resistance against pathogens or having certain nutritional value. CONCLUSIONS: Current review summarizes various applications of CRISPR/Cas system and its future prospects in plant biotechnology targeting crop improvement with higher yield, disease tolerance and enhanced nutritional value.

Novel methylation mark and essential hypertension.

BACKGROUND: Essential hypertension (EH) is an important risk factor for various cardiovascular, cerebral and renal disorders. It is a multi-factorial trait which occurs through complex interplay between genetic, epigenetic, and environmental factors. Even after advancement of technology and deciphering the involvement of multiple signalling pathways in blood pressure regulation, it still remains as a huge global concern. Genome-wide association studies (GWAS) have revealed EH-associated genetic variants but these solely cannot explain the variability in blood pressure indicating the involvement of additional factors. The etiopathogenesis of hypertension has now advanced to the level of epigenomics where aberrant DNA methylation is the most defined epigenetic mechanism to be involved in gene regulation. Though role of DNA methylation in cancer and other mechanisms is deeply studied but this mechanism is in infancy in relation to hypertension. Generally, 5-methylcytosine (5mC) levels are being targeted at both individual gene and global level to find association with the disease. But recently, with advanced sequencing techniques another methylation mark, N6-methyladenine (6mA) was found and studied in humans which was earlier considered to be absent in case of eukaryotes. Relation of aberrant 6mA levels with cancer and stem cell fate has drawn attention to target 6mA levels with hypertension too. CONCLUSION: Recent studies targeting hypertension has suggested 6mA levels as novel marker and its demethylase, ALKBH1 as probable therapeutic target to prevent hypertension through epigenetic programming. This review compiles different methylation studies and suggests targeting of both 5mC and 6mA levels to cover role of methylation in hypertension in broader scenario.

Current paradigms in epigenetic anticancer therapeutics and future challenges.

Any alteration at the genetic or epigenetic level, may result in multiplex of diseases including tumorigenesis which ultimately results in the cancer development. Restoration of the normal epigenome by reversing the epigenetic alterations have been reported in tumors paving the way for development of an effective epigenetic treatment in cancer. However, delineating various epigenetic events has been a challenging task so far despite substantial progress in understanding DNA methylation and histone modifications during transcription of genes. Many inhibitors in the form of epigenetic drugs mostly targeting chromatin and histone modifying enzymes including DNA methyltransferase (DNMT) enzyme inhibitors and a histone deacetylases (HDACs) inhibitor, have been in use subsequent to the approval by FDA for cancer treatment. Similarly, other inhibitory drugs, such as FK228, suberoylanilide hydroxamic acid (SAHA) and MS-275, have been successfully tested in clinical studies. Despite all these advancements, still we see a hazy view as far as a promising epigenetic anticancer therapy is concerned. The challenges are to have more specific and effective inhibitors with negligible side effects. Moreover, the alterations seen in tumors are not well understood for which one has to gain deeper insight into the tumor pathology as well. Current review focusses on such epigenetic alterations occurring in cancer and the effective strategies to utilize such alterations for potential therapeutic use and treatment in cancer.

Anti-Hypertensive Potential and Epigenetics of Angiotensin II type 2 Receptor (AT2R).

BACKGROUND: Renin angiotensin system (RAS) is a critical pathway involved in blood pressure regulation. Octapeptide, angiotensin II (Ang II), is a biologically active compound of RAS pathway which mediates its action by binding to either angiotensin II type 1 receptor (AT1R) or angiotensin II type 2 receptor (AT2R). Binding of Ang II to AT1R facilitates blood pressure regulation, whereas AT2R is primarily involved in wound healing and tissue remodeling. OBJECTIVES: Recent studies have highlighted the additional role of AT2R to counterbalance the detrimental effects of AT1R. Activation of angiotensin II type 2 receptor using AT2R agonist has shown the effect on natriuresis and release of nitric oxide. Additionally, AT2R activation has been found to inhibit angiotensin converting enzyme (ACE) and enhance angiotensin receptor blocker (ARB) activity. These findings highlight the potential of AT2R as a novel therapeutic target against hypertension. CONCLUSION: The potential role of AT2R highlights the importance of exploring additional mechanisms that might be crucial for AT2R expression. Epigenetic mechanisms, including DNA methylation and histone modification, have been explored vastly with relation to cancer, but the role of such mechanisms in the expression of AT2R has recently gained interest.

COVID-19 susceptibility: potential of ACE2 polymorphisms.

Background: Angiotensin-converting enzyme 2 (ACE2) is a metallopeptidase that primarily functions as a negative regulator of renin angiotensin system (RAS) by converting angiotensin II (Ang II) to angiotensin 1-7. Contrary to this, another RAS component, angiotensin-converting enzyme (ACE) catalyzes synthesis of Ang II from angiotensin I (Ang I) that functions as active compound in blood pressure regulation. This indicates importance of ACE/ACE2 level in regulating blood pressure by targeting Ang II. An outbreak of severe acute respiratory syndrome (SARS) highlighted the additional role of ACE2 as a receptor for SARS coronavirus (SARS-CoV) infection. Main body of the abstract: ACE2 is a functional receptor for SARS-CoV and SARS-CoV-2. Activation of spike (S)-protein by either type II transmembrane serine proteases (TTSPs) or cathepsin-mediated cleavage initiates receptor binding and viral entry. In addition to TTSPs, ACE2 can also be trimmed by ADAM 17 (a disintegrin and metalloproteinase 17) that competes for the same receptor. Cleavage by TTSPs activates ACE2 receptor for binding, whereas ADAM17 releases extracellular fragment called soluble ACE2 (sACE2). Structural studies of both ACE2 and S-protein have found critical sites involved in binding mechanism. In addition to studies on structural motifs, few single-nucleotide polymorphism (SNPs) studies have been done to find an association between genetic variants and SARS susceptibility. Though no association was found in those reports, but seeing the non-reproducibility of SNP studies among different ethnic groups, screening of ACE2 SNPs in individual population can be undertaken. Short conclusion: Thus, screening for novel SNPs focussing on recently identified critical regions of ACE2 can be targeted to monitor susceptibility towards coronavirus disease 2019 (COVID-19).

Functional relevance of promoter CpG island of human Angiotensin II type 1 receptor (AT1R) gene.

Angiotensin II type 1 receptor can activate number of signalling pathways upon stimulation and consequently its involvement in cancer progression have also been revealed. But which epigenetic mechanisms are involved in its regulation, need to be further explored. In-silico analysis revealed a promoter CpG island (CGI) which was cloned and assayed for functional activity using reporter gene system. The effect of methylation on this CGI was analysed through varying degree of methyltransferase treatment of cloned fragment. Results thus obtained were validated by direct sequencing. To further establish the status of this effect, in-vivo analysis was done through screening of methylation pattern in the targeted region among hypertensives (HTN) and normotensives (NTN) using PCR-Bisulphite sequencing. Additionally, clinical details of all participants, biochemical parameters and lifestyle related information was also collected and statistically evaluated. Reporter gene assay assigned functional activity to the cloned promoter CpG island. Increased dose and durations of methyltransferase treatment decreased the expression of reporter gene indicating the role of promoter DNA methylation. Among all the human samples screened, only one of the HTN individual was found to have single hemi-methylated CpG site at a position which happens to be a part of Sp1 transcription factor binding site. To conclude, CpG island in the promoter region of AT1R (CpG.P.AT1R) gets regulated through epigenetic mechanism of DNA methylation.

Unravelling the Lesser Known Facets of Angiotensin II Type 1 Receptor.

PURPOSE OF REVIEW: Hypertension is an important risk factor in various pathologies. Despite enormous advancements in health sciences, the number of hypertensive individuals is increasing worldwide. The complex interplay between genetic and epigenetic factors seems to be a promising pathway to exploring the pathophysiology of hypertension. RECENT FINDINGS: Various single gene and genome wide association studies have generated huge but non-reproducible data that highlights the role of some additional but as yet unidentified factor(s) in disease outcome. Dietary pattern and epigenetic mechanism (mainly DNA methylation) have shown a profound effect on blood pressure regulation. Angiotensin II and its receptors are known to play an important role in maintaining blood pressure; hence, a larger section of antihypertensive drugs targets the renin-angiotensin system (RAS). Angiotensin II type 1 receptor (AT1R), besides maintaining blood pressure, also has a role in cancer progression. Besides other pathways, RAS still remains the main player in blood pressure regulation. Additionally, AT1R has recently emerged as a molecule with diverse roles ranging from physiologic to cancer progression.

Lysosomal Storage Disorders in Nonimmune Hydrops Fetalis (NIHF): An Indian Experience.

Lysosomal storage disorders (LSD) are rare inherited neurovisceral inborn errors of metabolism which may present as nonimmune hydrops fetalis (NIHF) during pregnancy. Although causes of NIHF are highly diverse, LSDs are one of the underlying causes of NIHF. The aim of this study was to elucidate most frequent causes of LSDs presenting as NIHF in Indian population. Several fetal tissues were investigated for enzymatic diagnosis of LSDs using modified fluorometric assays in the current prospective study carried out at our national tertiary center from 2006 through 2016. Other general causes of NIHF were ruled out. Twenty-one percent (7/33) of cases were confirmed to have LSDs. Two patients were diagnosed with Hurler syndrome; two had Sly syndrome and one each of Niemann-Pick disease type A/B, Gaucher's disease, and mucolipidosis. Four of eleven cases (36%) with recurrent NIHF were found to have LSDs. In spite of extreme rarity of LSDs, they should be considered as a potential cause of NIHF, especially with recurrent NIHF. Specific investigations of LSD leading to definitive diagnosis may aid the clinician in providing accurate genetic counseling and prenatal diagnosis to the patients and help in subsequent pregnancies to the families. Furthermore, early intervention and management with enzyme replacement therapy may be planned for the lysosomal storage disorders where available.