Association Between a High Gene Expression Variant of Chymase and Increased Risk for Basal Cell Carcinoma.

BACKGROUND/AIM: Previous studies have associated certain variations in genes encoding factors of renin-angiotensin system (RAS), indirectly leading to higher angiotensin II (AngII) levels, with greater risk for basal cell carcinoma (BCC) development. Chymase (CMA1) is the main regulator of the RAS-independent AngII generation pathway and numerous studies have shown its oncogenic potential in several cancer types including BCC. In this study, we investigated the possible association between BCC pathogenesis and the functional DNA polymorphism A1903G (rs1800875) that affects expression of the CMA1 gene. PATIENTS AND METHODS: We genotyped 199 DNA samples, isolated from 100 BCC patients and 99 age, sex, and ethnicity-matched healthy controls for the CMA1 A1903G polymorphism. Genotyping was performed with PCR amplification, followed by MboI enzyme digestion and agarose gel electrophoresis of the resulted DNA fragments. RESULTS: The variant G allele that possibly increases CMA1 gene expression was not detected at a significantly different frequency between the groups of BCC patients and healthy controls. However, the AG heterozygous genotype was significantly increased in BCC patients compared with controls (p<0.001). CONCLUSION: The high expression CMA1 G allele carriers have an increased risk for BCC and elevated levels of chymase in the skin may have a carcinogenic effect.

Lack of Association Between the ACE2 G8790A Gene Variation and Risk for Basal Cell Carcinoma.

BACKGROUND/AIM: The G8790A (rs2285666) functional polymorphism of the angiotensin-converting enzyme 2 (ACE2) gene influences alternative mRNA splicing and quantitatively affects the enzyme's production. Specifically, the presence of the A allele has been associated with higher ACE2 plasma levels. In this study, we investigated the possible association of the functional polymorphism ACE2-G8790A with the pathogenesis of basal cell carcinoma (BCC). PATIENTS AND METHODS: A total of 190 DNA samples were studied, including 91 BCC patients and 99 controls of Greek origin. Molecular genotyping for the ACE2 G8790A polymorphism was carried out by PCR amplification, followed by AluI enzyme digestion and agarose gel electrophoresis of the DNA fragments. RESULTS: The allelic and genotypic frequencies presented no statistical difference between the patient and the control group. CONCLUSION: There is no association between the ACE2 G8790A polymorphism and pathogenesis of BCC.

Association study indicates combined effect of interleukin-10 and angiotensin-converting enzyme in basal cell carcinoma development.

Cytokines involved in inflammatory and immune response have been associated with risk for development of basal cell carcinoma (BCC). In this study, three functional DNA polymorphisms affecting gene expression were investigated in 54 BCC patients and 111 healthy controls: interleukin-1b (IL-1b) +3953C/T, interleukin-10 (IL-10) - 1082G/A and angiotensin-converting enzyme (ACE) insertion/deletion (I/D) polymorphisms. Significant increase of the variant alleles was observed in IL-10 - 1082G (P = 0.019) and in ACE D (P = 0.003) in BCC patients in comparison to controls. Multivariate logistic regression models evaluated the contribution of homozygous and heterozygous variant polymorphisms to the risk for BCC development. The studied polymorphisms influencing the expression of IL-10 and ACE genes were recognized as potential predictive factors for BCC. These findings suggest a possible molecular mechanism leading to BCC development that is likely to involve the activation of angiotensin receptors in combination with increased plasma levels of IL-10 in patients.

Recurrence in Oral Premalignancy: Clinicopathologic and Immunohistochemical Analysis.

Oral leukoplakia (OL) has a propensity for recurrence and malignant transformation (MT). Herein, we evaluate sociodemographic, clinical, microscopic and immunohistochemical parameters as predictive factors for OL recurrence, also comparing primary lesions (PLs) with recurrences. Thirty-three patients with OL, completely removed either by excisional biopsy or by laser ablation following incisional biopsy, were studied. Selected molecules associated with the STAT3 oncogenic pathway, including pSTAT3, Bcl-xL, survivin, cyclin D1 and Ki-67, were further analyzed. A total of 135 OL lesions, including 97 PLs and 38 recurrences, were included. Out of 97 PLs, 31 recurred at least once and none of them underwent MT, during a mean follow-up time of 48.3 months. There was no statistically significant difference among the various parameters in recurrent vs. non-recurrent PLs, although recurrence was most frequent in non-homogeneous lesions (p = 0.087) and dysplastic lesions recurred at a higher percentage compared to hyperplastic lesions (34.5% vs. 15.4%). Lower levels of Bcl-xL and survivin were identified as significant risk factors for OL recurrence. Recurrences, although smaller and more frequently homogeneous and non-dysplastic compared to their corresponding PLs, exhibited increased immunohistochemical expression of oncogenic molecules, especially pSTAT3 and Bcl-xL. Our results suggest that parameters associated with recurrence may differ from those that affect the risk of progression to malignancy and support OL management protocols favoring excision and close monitoring of all lesions.

Association of Polymorphisms in the Genes of Angiotensinogen and Angiotensin Receptors With Risk for Basal Cell Carcinoma.

BACKGROUND/AIM: Basal cell carcinoma (BCC) has been genetically associated with an increased expression of angiotensin-converting enzyme (ACE), an important factor of the renin-angiotensin system which produces vasoconstrictor angiotensin II. Other factors of this system include angiotensinogen (AGT) and angiotensin receptors AGTR1, AGTR2. We investigated the possible association of BCC with genetic variability in the AGT, AGTR1 and AGTR2 genes. MATERIALS AND METHODS: DNA samples of 190 Greeks were studied, including 91 patients with BCC and 99 matched healthy controls. Molecular genotyping of patients and controls was performed for the polymorphisms AGT M235T, AGTR1 A1166C and AGTR2 G1675A. RESULTS: The mutant T allele that increases AGT gene expression was detected in two-fold increased frequency in BCC patients in comparison to healthy controls (p <0.001). On the contrary, no significant difference was observed in AGTR1 and AGTR2 variants between patients and controls. CONCLUSION: Increased expression of AGT may be associated with BCC.

The platelet glycoprotein Ibalpha VNTR polymorphism is associated with risk for oral cancer.

BACKGROUND: We have previously found an association of platelet glycoprotein Ia polymorphism with increased risk for oral cancer. The purpose of this study was to investigate the possible relation of another platelet glycoprotein, Iba (GPIbalpha), with oral oncogenesis. PATIENTS AND METHODS: The variable number of tandem repeats (VNTR) polymorphism of the GPIbalpha gene, which affects the protein's structure and function, was examined in 162 Greek and German patients with oral squamous cell carcinoma and 225 healthy controls of equivalent age, gender and ethnicity. RESULTS: The B allele frequency detected, representing higher platelet activation, in the patient group and in the subgroups of patients without family history either of cancer or thrombophilia were significantly elevated in comparison with that of the control group (p = 0.03, p = 0.016 and p = 0.036, respectively). The D allele frequency (lower platelet activation) was significantly lower in comparison with controls only in patients with family history of thrombophilia. The frequency of B/B homozygotes was significantly increased in the total group of patients and the subgroup of patients with a family history of thrombophilia, in comparison with the control group (p = 0.042 and p = 0.043, respectively), while the frequency of heterozygotes for the C/B alleles was significantly lower in the subgroups of patients with a family history of cancer and thrombophilia (p = 0.036 and p = 0.027, respectively) compared to the control group. CONCLUSION: The VNTR polymorphism of the GPIbalpha gene, which affects the structure and function of this platelet glycoprotein, seems to be associated with risk for oral cancer, especially in patients without a family history of cancer.

Increased risk of oral cancer in diabetic animals is not associated with c-jun activation pathway.

PURPOSE: The expression of oncogenic protein c-jun was investigated in an experimental model of chemically induced carcinogenesis in normal and diabetic (type I) Sprague-Dawley rats. MATERIAL AND METHODS: Thirteen diabetic and twelve normal rats developed cancer after 4-nitroquinoline-N-oxide treatment, while six diabetic and six normal animals were used as controls. The biopsies were classified pathologically from oral mucosal dysplasia to moderately differentiated oral squamous cell carcinoma (OSCC) and studied immunohistochemically using monoclonal antibody against c-jun protein. RESULTS: Higher c-jun levels were observed in non-cancerous and precancerous stages of normal rats compared with diabetic rats, while in different tumour stages, the expression of c-jun was practically identical for both groups. CONCLUSION: It seems that diabetes does not affect the c-jun N-terminal kinase (JNK)/c-jun pathway.

Mutations of Two Major Coagulation Factors Are Not Associated with Male Infertility.

BACKGROUND: Thrombophilia-related mutations, such as coagulation factor V Leiden and factor II (G20210A), have been associated with female infertility due to spontaneous abortions during pregnancy. The possible role of mutations of these two factors in male infertility has not been studied to date. MATERIALS AND METHODS: A total of 208 unrelated Greek men were investigated, including 108 infertile men with idiopathic oligozoospermia, azoospermia, and oligozoospermia of various etiologies, as well as 100 fertile male controls. DNA extracted from participants' sperm or blood was analyzed for factor V Leiden and factor II G20210A mutations. RESULTS: There were no significant differences in factor V and factor II genotypes between infertile men and normal controls. CONCLUSION: An association of the two common thrombophilia-related mutations with male infertility was not observed in this preliminary study.

Cerebrovascular Aneurysms May Be Associated with Thrombophilia-predisposing Mutations in Patients with Familial Risk.

BACKGROUND: Pathogenesis of cerebral aneurysms implicates several risk factors. Three common thrombophilia-predisposing mutations were studied in patients with cerebrovascular aneurysms. PATIENTS AND METHODS: A total of 186 Greeks (66 patients with intracranial aneurysm and 120 healthy controls) were studied. Fifteen patients had a family history of thrombophilia, while two of them had a first-degree relative with an aneurysm. Genetic analysis for thrombophilia-predisposing mutations factor V Leiden, factor II (prothrombin) G20210A and methylenetetrahydrofolate reductase C677T was performed in all subjects. RESULTS: Genotypic distributions and allelic frequencies were compatible with the Hardy-Weinberg equilibrium. There was no significant difference between healthy individuals and patients in mutant allelic frequencies of thrombophilia mutations. Nevertheless, the mutant allelic frequencies of factor V and II mutations were significantly increased in the sub-group of patients with a positive family history of thrombophilia compared to controls (p≤0.003). CONCLUSION: Certain thrombophilia-related mutations may contribute to pathogenesis of intracranial aneurysms in a subset of the general population.

Association of angiotensin-converting enzyme gene insertion/deletion polymorphism with decreased risk for basal cell carcinoma.

The incidence of basal cell carcinoma (BCC) is significantly reduced in individuals treated with inhibitors of angiotensin I-converting enzyme (ACE) that produces angiotensin II. The objective of this study was to investigate the possible association of a functional polymorphism in the ACE gene, which affects its transcription, with risk for BCC. In DNA samples of 92 patients with BCC and 103 healthy controls of Greek origin and comparable age and gender, we studied the ACE gene insertion/deletion (I/D) polymorphism. Fisher's exact test was used for comparison of allele and genotype frequencies between the control and patients' groups. The detected low expression I allele frequency in the group of BCC patients was significantly decreased compared to controls (15.8 vs. 31.1 %, respectively; P = 0.001). ID heterozygotes exhibited 3.06 times lower BCC risk, compared with DD homozygotes (P = 0.001; OR = 0.327, 95 % CI = 0.174-0.615). The protective role of I allele was particularly prominent in women (P = 0.007, OR = 0.299, 95 % CI = 0.125-0.716), while for men it exhibited a marginal level (P = 0.041). These findings indicate that the low expression ACE I allele carriers have a decreased risk for BCC. The protective effect of the ID genotype against BCC may be explained by a possible underlying mechanism involving the effect of produced angiotensin II levels on its receptors due to putatively different binding affinity.

Oral carcinogenesis is not achieved in different carcinogen-treated PAI-1 transgenic and wild-type mouse models.

AIM: In an effort to assess the role of plasminogen activator inhibitor-1 (PAI-1) in oral squamous cancer development and progression, two different carcinogen treatment protocols were conducted. MATERIALS AND METHODS: Protocol I included mice from a PAI-1 transgenic (Tg) breed (n=56) and their wild-type (WT) counterparts (n=56), divided into one control group and two main experimental groups, treated with 7,12-dimethylbenz[a]anthracene (DMBA) for 8 and 16 weeks, respectively. Protocol II included the same number and types of animals and groups, which were similarly treated with 4-Nitroquinoline 1-oxide (4-NQO) in drinking water. Two drugs that affect plasma PAI-1 levels, enalapril and pravastatin, were administered to certain subgroups of animals in both protocols. RESULTS: None of the animals developed macroscopically-visible oral cancer lesions. Eleven animals under Protocol I and 52 animals under Protocol II died. Skin lesions were noted only in DMBA-treated animals (n=9). Almost all animals administered with 4-NQO developed alopecia and lost weight, while two of them developed stomach tumours, and one female mouse developed a large ovarian cyst. CONCLUSION: Transgenic mice may respond differently when used in well-established carcinogen models and oral carcinogenesis is hard to achieve in these rodents.

Comparison of cytology, colposcopy, HPV typing and biomarker analysis in cervical neoplasia.

BACKGROUND: Cervical cancer is the leading cause of mortality among women worldwide, despite existing prevention programs. In light of the recent development of anti-HPV vaccines, the aim of this study was to evaluate concurrently the efficacy of four methods for risk assessment (cytology, colposcopy, HPV molecular typing and detection of biomarkers in cervical biopsies) in an attempt to define the most efficient combination. PATIENTS AND METHODS: The studied group included 62 women with abnormal Pap tests and cervical lesions ranging from cervicitis and condylomas to intraepithelial neoplasias and invasive cancer. All women underwent full colposcopy assessment and colposcopically-taken biopsies were selected for histological examination, immunohistochemical identification of p16, p53, Bcl-2 biomarkers, as well as molecular detection and typing of HPV genomes. RESULTS: Cytology and colposcopy showed very high sensitivity in detecting CIN and cancer (91.7% and 94.4%, respectively), but low specificity (34.6% and 50%, respectively). The detection of the 3 biomarkers reached an impressive sensitivity (83.3%) and a moderate specificity (65.4%). HPV detection and typing achieved 77.8% sensitivity, and the highest specificity of 80.8% in detecting CIN and cancer cases. HPV DNA testing had the highest positive prognostic value (84.9%; confidence interval, CI: 67.4%- 94.3%) and cytology the lowest (66.0%; CI: 51.2%- 78.4%). Coupled HPV typing and colposcopy proved to be the most efficient combination, increasing sensitivity to 97.2% and negative prognostic value to 92.3%. The estimation of cervical neoplasia or cancer in women with high-risk HPV types increased approximately 15-fold (odds ratio, OR: 14.70; CI: 4.30-50.09, p<0.001), ~23-fold in the case of combined positive biomarkers (OR: 23.18; CI: 4.97- 104.23, p<0.001), and 35-fold in case of colposcopically detected cervical neoplasia (OR: 35.00; CI: 5.16- 225.07, p<0.001). CONCLUSION: The most efficient combination among all tested methodologies was found to be HPV typing with colposcopy.