miR-4716-3p and the target AKT2 Gene/rs2304186 SNP are associated with blood cancer pathogenesis in Pakistani population.

AKT2 is crucial for cancer cells' invasion, metastasis, and survival. It is a possible downstream gene target of cancer glycolysis-related microRNAs. The study investigated the role of miRNA-4716-3p, rs2304186, and the AKT2 gene in blood cancer pathogenesis. RT-qPCR was used to analyze AKT2 gene mRNA and miRNA-4716-3p expression in 200 blood cancer samples and 200 healthy controls. Furthermore, Tetra-ARMS PCR was used to examine the rs2304186 AKT2 SNP in 300 patients and 290 control samples. miRNA-4716-3p was shown to be significantly downregulated (p = 0.0294), whereas mRNA expression of the AKT2 gene was found to be significantly upregulated (p = 0.0034) in blood cancer patients compared to healthy individuals. miRNA-4716-3p downregulation (p = 0.0466) was more pronounced, while AKT2 upregulation was non-significant (p = 0.1661) in untreated patients compared to chemotherapy-treated patients. Blood cancer risk was significantly associated with the rs2304186 GT genotype (p = 0.0432), TT genotype (p = 0.0502), and mutant allele (T) frequency (p = 0.0008). Polymorphism rs2304186 was associated with an increased risk of blood cancer in dominant (p = 0.0011), recessive (p = 0.0502), and additive (p = 0.0008) genetic models. The results suggested that the rs2304186 and the deregulated expression of miRNA-4716-3p and AKT2 gene at the mRNA level may significantly increase the incidence of blood cancer, particularly in the Pakistani population. Therefore, these may function as suitable biomarkers for blood cancer diagnosis and prognosis. Additional, larger-scale investigations may be required to affirm these results.

Expression of PTEN and its correlation with proliferation marker Ki-67 in head and neck cancer.

INTRODUCTION: PTEN is part of large family of tyrosine phosphatases and has been found inactivated in a wide variety of human cancers. AIMS: In the present study we have tried to determine the association of the expression patterns of this gene with carcinogenesis. METHODS: First, a systematic review was carried out to ascertain the importance of the PTEN gene and its role in carcinogenesis. In the second phase, a case-control study was designed using different expression analysis techniques. Expression of PTEN mRNA was analyzed using reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Significantly downregulated expression of PTEN was observed in patients with head and neck cancer (HNC) compared to adjacent normal-tissue controls. These results were confirmed with quantitative polymerase chain reaction (qPCR). Significant downregulation of the gene was observed in HNC patients compared to adjacent normal-tissue controls. PTEN expression was correlated with different histopathological parameters of the study cohort by Spearman's correlation coefficient and a significant negative correlation was observed with pT stage (r = -0.271*; p<0.02) and grade (r = -0.228*; p<0.02) of HNC tissues. Furthermore, the expression variations of PTEN were correlated with the expression pattern of the proliferation marker Ki-67. Significantly (p<0.008) upregulated expression of Ki-67 was observed in HNC patients compared with adjacent normal-tissue controls This upregulation of Ki-67 was confirmed at the protein level by immunohistochemistry in HNC patients. When Spearman's correlation was carried out a significant negative correlation was observed between PTEN and Ki-67 (r = -0.230*; p<0.03). CONCLUSIONS: Our data suggest that downregulation of PTEN and overexpression of Ki-67 may contribute to the initiation and progression of HNC.

Genetic deletions of GSTM1 and GSTT1 in head and neck cancer: review of the literature from 2000 to 2012.

Head and neck cancer is one of the leading causes of deaths worldwide. Two genes GSTM1 and GSTT1 involved in phase II of carcinogen detoxification have been frequently studied in the literature. Their null genotypes are thought to be associated with increased head and neck cancer risk. However, the published reviews are not up to date and many important papers have been skipped. The current literature review was restricted to the null genotypes of the GSTM1 and GSTT1 genes with special emphasis on the genotypic status. We found that the size of study sample varied greatly and the oral cavity cancer was more influenced by GSTM1 and GSTT1 gene deletions. With respect to ethnicity Asians are more prone to head and neck cancers with these null genotypes as compared to Europeans and Americans. The current review showed significant associations (OR=9.0, 95%CI; 1.4-9.5; OR=3.7, 95%CI; 1.4-9.5) of GSTM1 and GSTT1 null genotypes with head and neck cancers. Review confirms the data of previous reviews that GSTM1 and GSTT1 gene polymorphisms may be risk factors for cancer initiation.

Sporadic mutations and expression analysis of spleen tyrosine kinase gene in breast cancer: a preliminary report.

Spleen tyrosine kinase (Syk) is an intracellular receptor protein kinase involved in cell proliferation, differentiation and phagocytosis. Syk expression has been reported in cell lines of epithelial origin. The strong expression of Syk in mammary gland prompted research into its potential role in mammary carcinogenesis. Fresh Biopsy samples were collected from different hospitals of Pakistan. Single stranded conformational polymorphism and Semi quantitative reverse transcriptase polymerase chain reaction was used to investigate somatic mutations and expression alterations in twenty five breast cancer tumor tissues along with their adjacent normal control tissue. Statistical analysis was performed to explore Syk association with breast cancer risk. In the present study, DNA from tumor tissue was analyzed and mutations in the coding sequence and intronic sequence spanning the exonic region of the Syk gene were identified. Sequence analysis revealed two missense: g61096G>A, g65967G>A, one frame shift: g87413insA and one silent mutation g42841G>A in exonic region while nucleotide variations were also observed in intronic region including one splice site mutation. These mutations in Syk are first time being reported in breast cancer. In addition, this study also revealed that Syk mRNA expression was markedly reduced in tissues of breast cancer compared to their adjacent normal control tissue.

The prevalence of HBV infection in the cohort of IDPs of war against terrorism in Malakand Division of Northern Pakistan.

BACKGROUND: Hepatitis B is an important public health problem in the Pakistani population and is the major cause of chronic hepatitis, cirrhosis, fibrosis and hepatocellular carcinoma. High prevalence of HBV infections has been observed especially in areas of low economic status. In spite of effective immunization programs, no significant change has been observed in the epidemiology of HBV in the rural areas of Pakistan (~67.5% of the total population) mainly due to lack of interest from government authorities and poor hygienic measures. The current study was aimed at estimating the prevalence and risk factors associated with HBV infection within internally displaced persons (IDPs) due to war against terrorism in the Malakand Division of Northern Pakistan. METHODS: Blood samples from 950 IDPs suspected with HBV infection (including both males and females) were collected and processed with commercial ELISA kits for HBsAg, Anti HBs, HBeAg, Anti HBe antibodies. The samples positive by ELISA were confirmed for HBV DNA by real-time PCR analysis. RESULTS: The overall prevalence of HBV observed was 21.05% of which 78.5% were males and 21.5% were females. Most confirmed HBV patients belong to the Malakand and Dir (lower) district. High-risk of infection was found in the older subjects 29.13% (46-60 years), while a lower incidence (11.97%) was observed in children aged <15 years. Lack of awareness, socioecomic conditions, sexual activities and sharing of razor blades, syringes and tattooing needles were the most common risk factors of HBV infection observed during the cohort of patients. CONCLUSION: The present study, revealed for the first time a high degree of prevalence of HBV infection in rural areas of Northern Pakistan. The noticed prevalence is gender- and age-dependent that might be due to their high exposures to the common risk factors. To avoid the transmission of HBV infection proper awareness about the possible risk factors and extension of immunization to the rural areas are recommended.

Effect of expressional alteration of KAI1 on breast cancer cell growth, adhesion, migration and invasion.

KAI1, also known as CD82, is a candidate metastasis suppressor gene and has been indicated in the disease progression of certain solid tumours, including those of breast cancer. The present study aimed to investigate the importance of KAI1 as a potential metastasis suppressor in breast cancer cells. MDA-MB-231 and MCF-7 sublines with different patterns of KAI1 expression were created by way of anti-KAI1 transgene or transfection of KAI1 expression construct. Cell adhesion was markedly increased in cancer cells showing increased expression of KAI1 (MCF-7(KAI1EXP), p=0.021 vs. control cells), while it was significantly reduced in the KAI1 knockout subline, MDA-MB-231(KAI1KO) (p=0.002 and 0.0004, respectively). Significant increase of cell migration of MCF-7(KAI1EXP) cells (p=0.024 vs. control) and restricted motility of MDA-MB-231(KAI1KO) cells (p=0.003) were observed. Furthermore, MCF-7(KAI1EXP) cells also showed reduced cell invasion (p=0.022), while MDA-MB-231(KAI1KO) cell line showed a significant increase in invasion (p=0.0063 and p=0.007, respectively). KAI1 did not affect cell growth. It is concluded therefore that KAI1 plays an important role in cell adhesion, invasion and migration of breast cancer cells, in vitro, and is a potential metastasis suppressor gene in breast cancer.

In vitro genotoxic assessment of xenobiotic diacylglycerols in an in vitro micronucleus assay.

Xenobiotic diacylglycerols (DG) may induce pathological disorders by causing abnormal chromosomal segregation, which could be aneuploid. In this study, seven xenobiotic-diacylglycerols (four of drug origin and three of pesticide origin) were evaluated for their ability to induce aneuploidy in mammalian cultures using in vitro cytokinesis blocked micronucleus (CBMN) assay coupled with kinetochore labeling and interphase fluorescent in situ hybridization. Out of seven xeno-DGs, two (ibuprofen-DG and fenbufen-DG) induced statistically significant (P < 0.001) and dose-dependent increase in micronucleus induction, but this apparent micronucleus induction was very weak in case of fenbufen-DG. These MN were produced predominantly by aneugenic and clastogenic mechanisms, respectively, confirmed by immunofluorescent labeling of kinetochores. Fluorescent in situ hybridization analysis revealed that ibuprofen-DG induced significantly higher nondisjunction for chromosomes 10, 17, and 18. Other xenobiotic diacylglycerols (indomethacin-DG, salicylic acid-DG, 4-(2-methyl-4-chlorophenoxy) butanoic acid-DG (MCPB-DG), 2-(2-methyl-4-chlorophenoxy) propanoic acid-DG (MCPP-DG) and 2-(4-dichlorophenoxy)-butanoic acid-DG (2,4 DB-DG) did not induce micronuclei, but the concentrations tested did not reach levels that caused the marked growth suppression typically required for testing for regulatory testing purposes. However, the levels of growth suppression achieved were similar to that seen with ibuprofen-DG, which was positive. This study shows that xeno-DGs, which have been neglected in the past for their possible link to any pathological disorders, need serious assessment of their mutagenic potential.

Contribution of BRCA1 germline mutation in patients with sporadic breast cancer.

Hereditary artifacts in BRCA1 gene have a significant contributory role in familial cases of breast cancer. However, its germline mutational penetrance in sporadic breast cancer cases with respect to Pakistani population has not yet been very well defined. This study was designed to assess the contributory role of germline mutations of this gene in sporadic cases of breast cancer. 150 cases of unilateral breast cancer patients, with no prior family history of breast cancer and no other disorders or diseases in general with age range 35-75 yrs, were included in this study.Mutational analysis for hot spots on Exon 2, 3 and 13 of BRCA1 was done by using Single Strand Conformational Polymorphism (SSCP). Sequence analysis revealed five variants (missense) and one novel splice site mutation at exon 13. No germline mutation was observed on the remaining exons with respect sporadic breast cancer cases in Pakistani population. A vast majority of breast cancer cases are sporadic; the present study may be helpful for designing a better genetic screening tool for germline BRCA mutations in sporadic breast cancer patients of Pakistani population. Further studies involving a screening of entire coding region of BRCA1 is required to explore the merits of genetic diagnosis and counseling in breast cancer patients.

Aneugenic and clastogenic effects of doxorubicin in human lymphocytes.

Doxorubicin, a benzanthroquinone anticancer agent, was examined for its effect on micronucleus induction in cultured human lymphocytes. A statistically significant dose-dependent increase in micronucleus frequency (P<0.001) in binucleated cells was seen and an increase in the kinetochore-positive (P<0.001) and kinetochore-negative micronuclei (P<0.001) was observed. An increase was also observed in the number of necrotic cells, but the frequency of apoptotic cells remained almost constant. This confirms that doxorubicin is both clastogenic and aneugenic.