Structural analysis, virtual screening and molecular simulation to identify potential inhibitors targeting 2'-O-ribose methyltransferase of SARS-CoV-2 coronavirus.

SARS-CoV-2, an emerging coronavirus, has spread rapidly around the world, resulting in over ten million cases and more than half a million deaths as of July 1, 2020. Effective treatments and vaccines for SARS-CoV-2 infection do not currently exist. Previous studies demonstrated that nonstructural protein 16 (nsp16) of coronavirus is an S-adenosyl methionine (SAM)-dependent 2'-O-methyltransferase (2'-O-MTase) that has an important role in viral replication and prevents recognition by the host innate immune system. In the present study, we employed structural analysis, virtual screening, and molecular simulation approaches to identify clinically investigated and approved drugs which can act as promising inhibitors against nsp16 2'-O-MTase of SARS-CoV-2. Comparative analysis of primary amino acid sequences and crystal structures of seven human CoVs defined the key residues for nsp16 2-O'-MTase functions. Virtual screening and docking analysis ranked the potential inhibitors of nsp16 from more than 4,500 clinically investigated and approved drugs. Furthermore, molecular dynamics simulations were carried out on eight top candidates, including Hesperidin, Rimegepant, Gs-9667, and Sonedenoson, to calculate various structural parameters and understand the dynamic behavior of the drug-protein complexes. Our studies provided the foundation to further test and repurpose these candidate drugs experimentally and/or clinically for COVID-19 treatment.Communicated by Ramaswamy H. Sarma.

Multi-omics integration of methyltransferase-like protein family reveals clinical outcomes and functional signatures in human cancer.

Human methyltransferase-like (METTL) proteins transfer methyl groups to nucleic acids, proteins, lipids, and other small molecules, subsequently playing important roles in various cellular processes. In this study, we performed integrated genomic, transcriptomic, proteomic, and clinicopathological analyses of 34 METTLs in a large cohort of primary tumor and cell line data. We identified a subset of METTL genes, notably METTL1, METTL7B, and NTMT1, with high frequencies of genomic amplification and/or up-regulation at both the mRNA and protein levels in a spectrum of human cancers. Higher METTL1 expression was associated with high-grade tumors and poor disease prognosis. Loss-of-function analysis in tumor cell lines indicated the biological importance of METTL1, an m7G methyltransferase, in cancer cell growth and survival. Furthermore, functional annotation and pathway analysis of METTL1-associated proteins revealed that, in addition to the METTL1 cofactor WDR4, RNA regulators and DNA packaging complexes may be functionally interconnected with METTL1 in human cancer. Finally, we generated a crystal structure model of the METTL1-WDR4 heterodimeric complex that might aid in understanding the key functional residues. Our results provide new information for further functional study of some METTL alterations in human cancer and might lead to the development of small inhibitors that target cancer-promoting METTLs.

Role of novel cancer gene SLITRK3 to activate NTRK3 in squamous cell lung cancer.

The development of targeted therapies that inhibit cancer-driving oncogenes has improved outcomes of patients diagnosed with lung adenocarcinoma (LUAD). In contrast, patients diagnosed with lung squamous cell carcinoma (LUSC) suffer worse survival outcomes and lack effective targeted treatment options. Identification of molecular drivers of LUSC to support development of targeted treatments is urgently needed. Addressing this need, the current report introduces the novel cancer gene SLIT- and NTRK-like family member 3 (SLITRK3) and its role in activating the neurotrophic receptor tyrosine kinase 3 (NTRK3) in LUSC cells. Multiple genome-wide data sets from patient samples were produced by us or downloaded from public databases to analyze tumor gene copy number aberrations, mRNA expression and associated survival outcomes. An accompanying mechanistic study employed LUSC cell lines and multiple methods, including in situ immunofluorescence, sphere-formation assay, and fluorescence-activated cell sorting analysis of the CD133-positive cell fraction. Altogether, the results indicate that gene amplification and consequent high expression of SLITRK3 in LUSC is associated with worse outcomes and induces SLITRK3-dependent activation of NTRK3 to promote a cancer stem cell phenotype that is inhibited by existing NTRK-targeted inhibitors. Based on a recent literature search, this is the first report of a mechanistic role for SLITRK3 in cancer.

Pan-cancer analysis of RNA methyltransferases identifies FTSJ3 as a potential regulator of breast cancer progression.

RNA methylation, catalysed by a set of RNA methyltransferases (RNMTs), modulates RNA structures, properties, and biological functions. RNMTs are increasingly documented to be dysregulated in various human diseases, particularly developmental disorders and cancer. However, the genomic and transcriptomic alterations of RNMTs, as well as their functional roles in human cancer, are limited. In this study, we utilized an unbiased approach to examine copy number alterations and mutation rates of 58 RNMTs in more than 10,000 clinical samples across 32 human cancer types. We also investigated these alterations and RNMT expression level as they related to clinical features such as tumour subtype, grade, and survival in a large cohort of tumour samples, focusing on breast cancer. Loss-of-function analysis was performed to examine RNMT candidates with important roles in growth and viability of breast cancer cells. We identified a subset of RNMTs, notably TRMT12, NSUN2, TARBP1, and FTSJ3, that were amplified or mutated in a subset of human cancers. Several RNMTs were significantly associated with breast cancer aggressiveness and poor prognosis. Loss-of-function analysis indicated FTSJ3, a 2'-O-Me methyltransferase, as a candidate RNMT with functional roles in promoting cancer growth and survival. A subset of RNMTs, like FTSJ3, represents promising novel targets for anticancer drug discovery. Our findings provide a framework for further study of the functional consequences of RNMT alterations in human cancer and for developing therapies that target cancer-promoting RNMTs in the future.

Harvest tissue source does not alter the protective power of stromal cell therapy after intestinal ischemia and reperfusion injury.

BACKGROUND: Transplantation of mesenchymal stromal cells (MSCs) may be a novel treatment for intestinal ischemia. The optimal stromal cell source that could yield maximal protection after injury, however, has not been identified. We hypothesized that (1) MSCs would increase survival and mesenteric perfusion, preserve intestinal histologic architecture, and limit inflammation after intestinal ischemia and reperfusion (I/R) injury, and (2) MSCs harvested from different sources of tissue would have equivalent protective properties to the intestine after I/R inury. METHODS: Adult male mice were anesthetized, and a midline laparotomy was performed. The intestines were eviscerated, the small bowel mesenteric root was identified, and baseline intestinal perfusion was determined using laser Doppler imaging. Intestinal ischemia was established by temporarily occluding the superior mesenteric artery for 60 min with a noncrushing clamp. After ischemia, the clamp was removed and the intestines were allowed to recover. Before abdominal closure, 2 × 10(6) human umbilical cord-derived MSCs, bone marrow-derived MSCs, or keratinocytes in 250 µL of phosphate-buffered saline vehicle were injected into the peritoneum. Animals were allowed to recover for 12 or 24 h (perfusion, histology, and inflammatory studies) or 7 d (survival studies). Survival data was analyzed using the log-rank test. Perfusion was expressed as a percentage of the baseline, and 12- and 24-h data was analyzed using one-way analysis of variance and the Student t-test. Nonparametric data was compared using the Mann-Whitney U-test. A P value of <0.05 was considered statistically significant. RESULTS: All MSCs increased 7-d survival after I/R injury and were superior to vehicle and keratinocytes (P < 0.05). All MSCs increased mesenteric perfusion more than vehicle at 12 and 24 h after injury (P < 0.05). All MSCs provided superior perfusion compared with keratinocytes at 24 h after injury (P < 0.05). Administration of each MSC line improved intestinal histology after I/R injury (P < 0.05). Multiple proinflammatory chemokines were downregulated after the application of MSCs, suggesting a decreased inflammatory response after MSC therapy. CONCLUSIONS: Transplantation of MSCs after intestinal I/R injury, irrespective of a tissue source, significantly increases survival and mesenteric perfusion and at the same time limits intestinal damage and inflammation. Further studies are needed to identify the mechanism that these cells use to promote improved outcomes after injury.

Human Adipose Stromal Cells Increase Survival and Mesenteric Perfusion Following Intestinal Ischemia and Reperfusion Injury.

OBJECTIVE: Intestinal ischemia can quickly escalate to bowel necrosis and perforation. Transplantation of stem cells presents a novel treatment modality for this problem. We hypothesized that: human adipose-derived stromal cells (hASCs) would increase survival and mesenteric perfusion to a greater degree compared with differentiated cellular controls following ischemic intestinal injury, and improved outcomes with hASC therapy would be associated with preservation of intestinal histological and tight junction architecture, and lower levels of systemic inflammation following intestinal injury. METHODS: hASCs and keratinocytes (differentiated cellular control) were cultured on polystyrene flasks at 37°C in 5% CO2 in air. Adult male C57Bl6J mice were anesthetized and a midline laparotomy performed. The intestines were eviscerated, the small bowel mesenteric root identified, and intestinal ischemia was established by temporarily occluding the superior mesenteric artery for 60 min with a noncrushing vascular clamp. Following ischemia, the clamp was removed, and the intestines were returned to the abdominal cavity. Before abdominal closure, 2 million hASCs or keratinocytes in 250 µL of phosphate-buffered saline (carrier for cells and control solution) were infused into the peritoneum. Animals were allowed to recover for 12 or 24 h (perfusion, histology, cytokine, and immunofluoresence studies), or 7 days (survival studies). Intestinal perfusion was assessed by laser Doppler imaging. Intestinal tissue segments were stained with hematoxylin and eosin, as well as antibodies for the tight junction protein claudin-1. Separate aliquots of intestine, liver, and lung tissue were homogenized and assessed for inflammatory cytokines via multiplex beaded assay. RESULTS: Animals administered hASCs following intestinal ischemia and reperfusion (I/R) injury had significantly greater 7-day survival and better postischemic recovery of mesenteric perfusion compared with vehicle or keratinocyte therapy. hASCs also abated intestinal mucosal destruction, facilitated preservation of intestinal tight junctions, and decreased the systemic inflammatory response to injury. CONCLUSIONS: Human adipose-derived stromal cells improved survival and mesenteric perfusion and attenuated the mucosal damage associated with intestinal I/R injury. hASCs should be considered as a plausible cell source for novel cellular treatment plans following intestinal ischemia.