RESUMO
BACKGROUND: Ovarian carcinoma is one of the leading causes of cancer-related death among females. K-ras codon 12 mutations are commonly occurring mutations in different types of cancers and leads to resistance against anti-EGFR therapeutics. Hence, determination of mutations in k-ras gene is crucial for predicting response to anti-EGFR therapies. This study aimed to evaluate the prevalence of k-ras gene mutations and CA125 tumor marker in patients with ovarian carcinoma in Tabriz city. METHODS: Genomic DNA was extracted from 67 tissues pertained to women with ovarian carcinoma. Mutations in codon 12 and 13 of k-ras gene were analyzed by Nested PCR and RFLP methods. Titer of CA125 tumor marker was determined by ELISA. RESULTS: Among the 67 patients, 7 patients (10.4%) had mutation in k-ras codon 12 while none of them had mutation in k-ras codon 13. Based on the results, the frequency of various genotypes were 89.55%, 10.44%, and 0%, for GG, GA, and AA alleles, respectively. The p-value for stage I and Grade I tumors were 0.022 and 0.04, respectively, indicating a statistically significant correlation between codon 12 mutation and stage I and Grade I tumors. Furthermore, we found significant correlations among CA125 tumor marker titers and histological grade (p<0.000) and stage (p<0.000). The mean serum CA125 tumor marker levels in malignant carcinomas were 499.84 U/ml. CONCLUSION: The results in this study indicated high prevalence of k-ras codon 12 mutations and high titer of CA125 tumor marker in patients with ovarian carcinoma in the study region.
RESUMO
BACKGROUND: Overexpression of EGFR is associated with carcinogenesis in more than 70% of head and neck cancers. Anti-EGFR monoclonal antibodies bind to the extracellular domain of EGFR and block the EGFR downstream signaling pathway, which results in the suppression of the growth of the tumor cells. Escherichia coli is the preferred system for expressing various recombinant proteins, including single chain antibodies, but the formation of inclusion bodies negatively affects the efficacy of this system. Several strategies have been suggested to solve this problem, notably the utilization of molecular chaperones. OBJECTIVES: In this study, we attempted to increase the soluble expression of huscfv antibody via co-expression with the cytoplasmic chaperones. MATERIALS AND METHODS: To achieve this purpose, chaperones plasmids pG-KJE8, pGro7, pKjE7, pTf16 and pG-Tf2 encoding cytoplasmic chaperones were co-expressed with the humanized anti-EGFR scFv construct in E. coli. Different temperatures, incubations times, and concentrations of IPTG were used to produce an active antibody with the highest solubility. Results were analyzed by SDS-PAGE. Soluble huscFv was purified by Ni-NTA column and the biologic activity of the recombinant protein was determined by ELISA. RESULT: The results indicated that the highest concentrations of humanized anti-EGFR scFv were obtained by co-expression of huscFv via chaperone plasmid pG-KJE8 with 0.2 mM concentration of inducer (IPTG), culture temperature of 25 °C, and 4 h incubation time after induction. CONCLUSION: In conclusion, co-expression with chaperones could be used as an efficient strategy to produce soluble active ScFvs in E. coli.
RESUMO
The cell cycle is controlled by precise mechanisms to prevent malignancies such as cancer, and the cell needs these tight and advanced controls. Cyclin dependent kinase inhibitor p27 (also known as KIP1) is a factor that inhibits the progression of the cell cycle by using specific molecular mechanisms. The inhibitory effect of p27 on the cell cycle is mediated by CDKs inhibition. Other important functions of p27 include cell proliferation, cell differentiation and apoptosis. Post- translational modification of p27 by phosphorylation and ubiquitination respectively regulates interaction between p27 and cyclin/CDK complex and degradation of p27. In this review, we focus on the multiple function of p27 in cell cycle regulation, apoptosis, epigenetic modifications and post- translational modification, and briefly discuss the mechanisms and factors that have important roles in p27 functions.
