Forecasting most deleterious nsSNPs in human <i>TLR9</i> gene and their cumulative impact on biophysical features of the protein using <i>in silico</i> approaches

In women, the uterine cervix and corpus uteri are two main suspects, playing a major role in cancer-associated-mortality. Immunologically, Toll-like receptors (TLRs) associated with the innate immune system, can recognize pathogens and induce immune responses against pathogens. Cellularly, TLR9 expression occurs in immune system cells including macrophages, natural killer cells, dendritic cells, and other antigen-presenting cells. TLR9 recognizes and interacts with viral and bacterial DNA comprising cytosine-phosphate-guanine (CpG) dideoxynucleotide motif. The current study is designed to identify the most deleterious nonsynonymous single nucleotide polymorphisms (nsSNPs) in the TLR9 gene and to delineate their deleterious effect on the structural and functional features of proteins at the molecular level. Based on the implementation of various computational tools and algorithms eight most deleterious nsSNPs (P139H, R257C, C265Y, L283P, G514D, L544Q, H566Y, and W670R) have been identified in the human TLR9 gene as potentially damaging SNPs. Further, our study suggests highly conserved patterns at deleterious nsSNPs sites could influence protein stability and its functional features. Additionally, this study identifies two nsSNPs (G514D and W670R) associated with the severity of Uterine corpus endometrial carcinoma. In support of our computational findings, the validation of key results using polymerase chain reaction and other experimental methods is warranted in the Indian population. In general, this study might be able to delineate the guideline for identifying the most damaging SNPs and enhances the understating of the risk factors for cancer and disease susceptibilities.</p

Demographic characteristics of the population studied.

<p>Demographic characteristics of the population studied.</p

Clinical Impact of De-Regulated Notch-1 and Notch-3 in the Development and Progression of HPV-Associated Different Histological Subtypes of Precancerous and Cancerous Lesions of Human Uterine Cervix

<div><p>Background</p><p>Cervical cancer is the leading cause of cancer related deaths among women in India. Limited reports are available for Notch-1 and Notch-3 protein in cervical carcinoma, which play crucial role in cell proliferation, differentiation, and apoptosis.</p><p>Methods</p><p>This study was designed to evaluate the role of Notch-1 and Notch-3 with context to HPV infection in cervical carcinoma. A total of 168 tissue biopsy samples comprising of tumor specimens (n = 98), precancer (n = 30) and non-neoplastic cervical tissues (n = 40) were screened for HPV infection by PCR and expression of Notch-1 and Notch-3 protein by Immunohistochemistry and Immunoblotting.</p><p>Results</p><p>80% (24/30) were found to be positive for HPV in precancer and 86.7% (85/98) in cancer patients. Notch-1 expression of precancer and cancer cases was found to be significantly down-regulated with severity of disease in nuclear (3.43±0.29; 2.04±0.19, p = 0.0001, p = 0.0001) and cytoplasm (3.07±0.29; 2.29±0.17, p = 0.0001, p = 0.0001) obtained from different stages as compared to normal cervix tissue (5.40±0.19, 4.97±0.15; p<0.001; p<0.001). However, Notch-3 expression of above cases was significantly up-regulated with severity of disease and showed intense nuclear (4.17±0.39; 4.74±0.18, p = 0.0001, p = 0.0001) and cytoplasm (3.67±0.36; 4.48±0.18, p = 0.0001, p = 0.0001) of different stages as compared to normal cervix tissue (0.95±0.20, 0.70±0.20; p<0.001; p<0.001) respectively.</p><p>Conclusions</p><p>These findings suggest that Notch-1 and Notch-3 may play an important role with synergistic effect of HPV in regulating development and proliferation of cervical cancer through the deregulation of Notch signalling. This study also shows the clinical utility of both proteins which may be used as predictable biomarkers in diagnosing different histological sub-types of HPV associated cervical cancer. Nevertheless, abnormal activation of this pathway may provide legitimate targets for cervical cancer therapy.</p></div

Total score (Intensity score + Percentage positivity, Mean ± S.E) of Notch-1 and Notch-3 expression in normal, precancerous and cancer tissues.

<p>p-values are calculated using the chi-square test.</p><p>* p≤0.05 was considered significant.</p>▾<p>Downregulation;^▴Upregulation.</p

Awareness about cervical cancer, HPV and HPV vaccine.

<p>*Number indicate participants who had knowledge about cervical cancer and know the causative agent.</p><p>**Number indicate participants who had both knowledge and know the causative agent.</p><p>***General opinion among all participants.</p><p>Awareness about cervical cancer, HPV and HPV vaccine.</p

Detection of Human Papilloma virus genotypes 16 and 18 in precancer & cervical cancer patients by In vitro nucleic acid amplification test.

<p>(<b>A</b>) β-globin, 288bp (<b>B</b>) HPV-L1, 450bp (<b>C</b>) HPV-16, 217bp (<b>D</b>) HPV-18, 100bp as indicated in Materials and Methods section.</p

Comparison of awareness among Females and Males.

<p>Comparison of awareness among Females and Males.</p

Awareness about cervical cancer and HPV according to living area.

<p>Awareness about cervical cancer and HPV according to living area.</p

Correlation of HPV infection with Notch-3 protein expression in precancer.

<p>*p≤0.05 is considered as significant.</p

Correlation of HPV infection with Notch-3 protein expression in ISCC.

<p>*p≤0.05 is considered as significant.</p