A novel kinase mutation in VEGFR-1 predisposes its αC-helix/activation loop towards allosteric activation: Atomic insights from protein simulation.

Vascular endothelial growth factor receptor 1 (VEGFR-1) has been implicated in diverse pathologies, including cancers. Although VEGFR-1 is considered as functionally impaired kinase, its decoy characteristics make it an important regulator of VEGFR-mediated signaling, particularly in tumor angiogenesis. VEGFR-1 conveys signaling via its tyrosine kinase (TK) domain whose activation is regulated by phosphorylation of specific tyrosine residues. Thus dysregulation of VEGFR-1 signaling, as reported in most of the cancers, might be a consequence of altered phosphorylation that could be attributed to genotypic variations in its TK domain. Considering the importance of TK domain of VEGFR-1, we carried out its mutational screening in 84 clinically validated and histopathologically confirmed colorectal cancer patients. By means of direct DNA sequencing and SNP analyses, eight novel variations, including one synonymous, two deletion, one missense and four intronic variations, were reported in the TK domain of VEGFR-1. rs730882263:C>G variation specifically reported in colon cancer, representing a single-atomic change (Sulfur to Oxygen) in the predicted (p.Cys1110Ser) protein, was observed as potentially deleterious variation as assessed by multiple single-nucleotide polymorphism prediction servers. Molecular dynamics simulations of VEGFR-1 Wt and (p.Cys1110Ser) variant models revealed major conformational changes in variant protein presumptuously generating an open conformation thereby exposing the activation domain and consequently increasing the probability of phosphorylation events: a condition frequently reported in cancers.

Tectorigenin ablates the inflammation-induced epithelial-mesenchymal transition in a co-culture model of human lung carcinoma.

OBJECTIVES: Tumors not only manage to escape from the host immune system, but they effectively contrive to benefit from infiltrating immune cells by modifying their functions so as to create a pro-inflammatory microenvironment favorable for tumor progression and metastasis. In this study we investigated if tectorigenin could suppress lung cancer-induced pro-inflammatory response generated from monocytes. MATERIALS AND METHODS: A549:THP1 co-culture model was set-up favoring release of pro-inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor alpha (TNF-α). Effect of tectorigenin on A549 imparted invasive phenotype of A549:THP-1 co-culture was monitored by cytokine release from monocytes, and metastasis/epithelial-mesenchymal transitiom (EMT) in A549 cells. RESULTS: In a contact A549:THP1 co-culture model, THP-1 cells were activated by A549 cells favoring secretion of pro-inflammatory cytokines, TNF-α and IL-6. However, priming of A549 cells with tectorigenin for 24h repressed A549 cell-induced secretion of TNF-α and IL-6 by THP-1 cells. Tectorigenin induced change in functional phenotype of A549 cells rendered THP-1 cells non-responsive for the secretion of IL-6 and TNF-α in a contact co-culture setup. Additionally, conditioned media from this non-responsive A549:THP-1 co-culture suppressed metastatic potential of A549 cells as confirmed by the wound healing and transwell migration assays. These finding were further corroborated by decrease in expression of Snail with a concomitant increase in E-cadherin, the two signature markers of EMT. CONCLUSION: These results clearly demonstrate the therapeutic potential of tectorigenin to prevent lung cancer elicited inflammatory and pro-metastatic response in monocytes and warrants further investigations to elucidate its mechanism of action.

Affinity proteomics led identification of vimentin as a potential biomarker in colon cancers: insights from serological screening and computational modelling.

Proteomic analysis using multiplex affinity reagents is perhaps the most reliable strategy to capture differentially expressed proteins that are slightly or immensely modified. In addition to expressional variation, it is comprehensively evident that the immunogenicity of a protein can be a deciding factor for instigating an inflammation afflicted-carcinogenesis. Considering both these factors, a simple and systematic strategy was designed to capture the immunogenic cancer biomarkers from sera of colorectal cancer patients. The affinity reagent, in the form of an antibody repertoire against the secretome of the HT29 cell line was used to grade the sera samples on the basis of the degree of immuno-reactivity and to capture differentially expressed antigens from the patient sera. Following affinity based 2DE-MALDI-TOF; the proteins were identified as (1) soluble vimentin; and (2) TGF-beta-inhibited membrane-associated protein (PP16B), in colon cancer sera and (3) keratin, type II cytoskeletal protein in rectal cancer sera. Pathway reconstruction and protein-protein networking of identified proteins predicted only Vimentin to be physically and genetically engaged in close proximity with the most established colorectal cancer associated tumorigenic pathways. Furthermore, our findings suggest that a possible surface stoichiometric shift in the structure of protein could be due to mutations in the coding sequence of Vimentin that may elicit its enhanced secretion possibly due to protein-hyperphosphorylation. Of the three proteins identified, only Vimentin showed higher expression in sera of colon cancer patients alone. Thus, it could be argued that vimentin might help in predicting individuals at higher risk of developing colon cancers. Our data are therefore suggestive of using vimentin as an antigen for tumor vaccination in an autologous set-up for colon cancers.