Pharmacophore modeling for identification of anti-IGF-1R drugs and in-vitro validation of fulvestrant as a potential inhibitor.

Insulin-like growth factor 1 receptor (IGF-1R) is an important therapeutic target for breast cancer treatment. The alteration in the IGF-1R associated signaling network due to various genetic and environmental factors leads the system towards metastasis. The pharmacophore modeling and logical approaches have been applied to analyze the behaviour of complex regulatory network involved in breast cancer. A total of 23 inhibitors were selected to generate ligand based pharmacophore using the tool, Molecular Operating Environment (MOE). The best model consisted of three pharmacophore features: aromatic hydrophobic (HyD/Aro), hydrophobic (HyD) and hydrogen bond acceptor (HBA). This model was validated against World drug bank (WDB) database screening to identify 189 hits with the required pharmacophore features and was further screened by using Lipinski positive compounds. Finally, the most effective drug, fulvestrant, was selected. Fulvestrant is a selective estrogen receptor down regulator (SERD). This inhibitor was further studied by using both in-silico and in-vitro approaches that showed the targeted effect of fulvestrant in ER+ MCF-7 cells. Results suggested that fulvestrant has selective cytotoxic effect and a dose dependent response on IRS-1, IGF-1R, PDZK1 and ER-α in MCF-7 cells. PDZK1 can be an important inhibitory target using fulvestrant because it directly regulates IGF-1R.

Association of rs1801157 single nucleotide polymorphism of CXCL12 gene in breast cancer in Pakistan and in-silico expression analysis of CXCL12-CXCR4 associated biological regulatory network.

BACKGROUND: C-X-C chemokine ligand 12 (CXCL12) has important implications in breast cancer (BC) pathogenesis. It is selectively expressed on B and T lymphocytes and is involved in hematopoiesis, thymocyte trafficking, stem cell motility, neovascularization, and tumorigenesis. The single nucleotide polymorphism (SNP) rs1801157 of CXCL12 gene has been found to be associated with higher risk of BC. METHODS: Our study focuses on the genotypic and allelic distribution of SNP (rs1801157; G/A) in Pakistani population as well as its association with the clinico-pathological features. The association between rs1801157 genotypes (G/A) and BC risks was assessed by a multivariate logistic regression (MLR) analysis. Genotyping was performed in both healthy individuals and patients of BC using PCR-restriction fragment length polymorphism (PCR-RFLP) method. Furthermore, in-silico approaches were adapted to investigate the association of CXCL12 and its receptor CXCR4 with genes/proteins involved in BC signalling. RESULTS: Significant differences in allelic and genotypic distribution between BC patients and healthy individuals of genotype (G/G) and (A/G) (p < 0.05) were observed. The frequency of the allele G in the BC group (77%) was significantly higher as compared to control group (61%) (p = 0.01). The association of genotype GG with clinico-pathological features including age, stages of cancer and organ (lung, liver, bones and brain) metastasis (p > 0.05) was assessed. In a MLR analysis, a number of variables including age, weight of an individual, affected lymph nodes, hormonal status (estrogen and progesterone receptor), alcohol consumption and family history associated with the GG genotype (GG:AA, odds ratio (OR) = 1.30, 95% CI [1.06-1.60]) were found to be independent risk factors for BC. Our in-vitro results suggest that genotype GG is possibly increasing the risk of BC in Pakistani cohorts. in-silico analysis finds that CXCL12-CXCR4 is associated with an increased expression of PDZK1, PI3k and Akt which lead the breast tumor towards metastasis. CONCLUSION: Multiple targets such as CXCL12, CXCR4, PDZK1, PI3k and Akt can be inhibited in combined strategies to treat BC metastasis.

Formal modeling and analysis of ER-α associated Biological Regulatory Network in breast cancer.

BACKGROUND: Breast cancer (BC) is one of the leading cause of death among females worldwide. The increasing incidence of BC is due to various genetic and environmental changes which lead to the disruption of cellular signaling network(s). It is a complex disease in which several interlinking signaling cascades play a crucial role in establishing a complex regulatory network. The logical modeling approach of René Thomas has been applied to analyze the behavior of estrogen receptor-alpha (ER-α) associated Biological Regulatory Network (BRN) for a small part of complex events that leads to BC metastasis. METHODS: A discrete model was constructed using the kinetic logic formalism and its set of logical parameters were obtained using the model checking technique implemented in the SMBioNet software which is consistent with biological observations. The discrete model was further enriched with continuous dynamics by converting it into an equivalent Petri Net (PN) to analyze the logical parameters of the involved entities. RESULTS: In-silico based discrete and continuous modeling of ER-α associated signaling network involved in BC provides information about behaviors and gene-gene interaction in detail. The dynamics of discrete model revealed, imperative behaviors represented as cyclic paths and trajectories leading to pathogenic states such as metastasis. Results suggest that the increased expressions of receptors ER-α, IGF-1R and EGFR slow down the activity of tumor suppressor genes (TSGs) such as BRCA1, p53 and Mdm2 which can lead to metastasis. Therefore, IGF-1R and EGFR are considered as important inhibitory targets to control the metastasis in BC. CONCLUSION: The in-silico approaches allow us to increase our understanding of the functional properties of living organisms. It opens new avenues of investigations of multiple inhibitory targets (ER-α, IGF-1R and EGFR) for wet lab experiments as well as provided valuable insights in the treatment of cancers such as BC.

Significant correlation of TLR4 expression with the clinicopathological features of invasive ductal carcinoma of the breast.

Toll-like receptor 4 belongs to the family of pattern recognition receptors and is therefore an important part of innate immunity. Since the last decade, Toll-like receptors (TLRs) have gained much interest in cancer research due to their role in tumor progression. However, there is scarcity of data evaluating TLR4 expression in different cell populations of tumor in relation to clinicopathological parameters of invasive ductal carcinoma of the breast. Therefore, we investigated TLR4 expression in 50 primary invasive ductal carcinoma and 17 lymph node sections through immunohistochemistry. Significant association of TLR4 expression with local metastasis and absence of necrosis was observed. The incidence of lymph node metastasis was higher in patients with high TLR4 expression in the tumor cells. We also observed an interesting inverse relationship of TLR4 with necrosis. In endothelial and lymphocytic population constituting breast tumors, a significant association of TLR4 expression was seen with the tumor grade. Therefore, we conclude that TLR4 expression is of biological interest both as a prognostic marker as well as a possible therapeutic target.

A case of lymphangioleiomyomatosis with membranous nephropathy and likely systemic lupus.

Membranous glomerulonephritis is most often idiopathic, but it can be secondary to systemic lupus erythematosus, viral hepatitis, and drugs. A number of malignancies have also been associated with membranous glomerulonephritis, although a causal link has not been established yet. A young patient with lymphangioleiomyomatosis is described who developed massive proteinuria and was found to have membranous glomerulonephritis on renal biopsy. There were many pathological features and some clinical features that suggested the patient also suffered from systemic lupus erythematosus.