Epigenetic deregulations of Wnt/β-catenin and transforming growth factor beta-Smad pathways in esophageal cancer: Outcome of DNA methylation

Background: Promoter methylation of tumor suppressor genes (TSGs) is a well-reported portent in carcinogenesis; hence, it is worthy to investigate this in high-risk Northeast population of India. The study was designed to investigate methylation status of 94 TSGs in esophageal squamous cell carcinoma (ESCC). Further, the effect of OPCML promoter methylation on gene expression was analyzed by immunohistochemistry. Moreover, in silico protein–protein interactions were examined among 8 TSGs identified in the present study and 23 epigenetically regulated genes reported previously by our group in ESCC. Materials and Methods: Methylation profiling was carried out by polymerase chain reaction array and OPCML protein expression was examined by tissue microarray-based immunohistochemistry. Results: OPCML, NEUROG1, TERT, and WT1 genes were found hypermethylated and SCGB3A1, CDH1, THBS1, and VEGFA were hypomethylated in Grade 2 tumor. No significant change in OPCML expression was observed among control, Grade 1, and Grade 2 tumor. Conclusively, hypermethylation of the studied OPCML promoter in Grade 2 tumor produced no effect on expression. Unexpectedly, OPCML expression was downregulated in Grade 3 tumor in comparison to other groups signifying that downregulation of OPCML expression may lead to higher grade of tumor formation at the time of diagnosis of ESCC in patients. Significant interactions at protein level were found as VEGFA:PTK2, CTNNB1:CDH1, CTNNB1:VEGFA, CTNNB1:NEUROG1, CTNND2:CDH1, and CTNNB1:TERT. These interactions are pertinent to Wnt/β-catenin and TGF-β-Smad pathways. Conclusions: Deranged OPCML expression may lead to high-grade ESCC as well as epigenetically regulated genes, that is, CDH1, CTNNB1, CTNND2, THBS1, PTK2, WT1, OPCML, TGFB1, and SMAD4 may alter the Wnt/β-catenin and TGF-β-Smad pathways in ESCC. Further study of these genes could be useful to understand the molecular pathology of ESCC with respect to epithelial-mesenchymal transition (EMT) mediated by Wnt/β-catenin and TGF-β signaling pathways

Is CXCL10/CXCR3 axis overexpression a better indicator of leprosy type 1 reaction than inducible nitric oxide synthase.

Background & objectives: Leprosy type 1 reactions (T1R) are acute episodes of immune exacerbation that are a major cause of inflammation and nerve damage. T1R are diagnosed clinically and supported by histopathology. No laboratory marker is currently available that can accurately predict a T1R. Increased plasma and tissue expression of inducible nitric oxide synthase (i-NOS) and chemokine CXCL10 have been demonstrated in T1R. We studied the gene expression and immunoexpression of i-NOS, CXCL10 and its receptor CXCR3 in clinically and histopathologically confirmed patients with T1R and compared with non-reactional leprosy patients to understand which biomarker has better potential in distinguishing reaction from non-reaction. Methods: Gene expression of i-NOS, CXCL10 and CXCR3 was studied in 30 skin biopsies obtained from patients with borderline tuberculoid (BT), mid-borderline (BB) and borderline lepromatous (BL) leprosy with and without T1R by real-time PCR. Further validation was done by immunhistochemical expression on 60 borderline leprosy biopsies with and without T1R. Results: Of the 120 patients histopathological evaluation confirmed T1R in 65 (54.2%) patients. CXCR3 gene expression was significantly (P<0.05) higher in BT- and BB-T1R patients compared to those without T1R. The CXCL10 gene expression was significantly higher (P<0.05) in BB leprosy with T1R but the difference was not significant in patients with BT with or without T1R. Immunoexpression for CXCR3 was significant in both BB-T1R and BB (P<0.001) and BT and BT-T1R (P<0.001). Immunoexpression of CXL10 was significant only in differentiating BB from BB-T1R leprosy (P<0.01) and not the BT cases. i-NOS immunoexpression was not useful in differentiating reactional from non-reactional leprosy. Interpretation & conclusions: Both CXCL10 and CXCR3 appeared to be useful in differentiating T1R reaction in borderline leprosy while CXCR3 alone differentiated BT from BT-T1R. CXCR3 may be a potentially useful immunohistochemical marker to predict an impending T1R.

Copy number polymorphism of glutathione-S-transferase genes (GSTM1 & GSTT1) in susceptibility to lung cancer in a high-risk population from north-east India.

Background & objectives: Genetic polymorphisms in glutathione-S-transferase genes (GSTM1 and GSTT1) have been studied intensively for their potential role in lung cancer susceptibility. However, most of the studies on association between the polymorphisms and lung cancer do not distinguish between genotypes with one or two copies of the genes. The present study investigates the gene dosage effects of GSTT1 and GSTM1 copy number and their environmental interactions to examine the association of lung cancer risk with trimodular genotypes of the GSTs in a high-risk population from north-east India. Methods: A total of 154 lung cancer cases and 154 age and sex matched controls from the high risk region of north-east India were analyzed by multiplex real-time PCR to determine the trimodal genotypes (+/+, +/- and -/-) in both the genes (GSTM1 and GSTT1). Results: No significant association and gene dosage effect of GSTM1 gene copy number with lung cancer risk (Ptrend=0.13) were found. However, absence of GSTT1 conferred 68 per cent (OR=0.32;95%CI=0.15-0.71;P=0.005) reduced risk compared to the two copy number of the gene. tThere was evidence of gene dosage effect of GSTT1 gene (Ptrend=0.006). Tobacco smoking was a major environmental risk factor to lung cancer (OR=3.03;95%CI=1.73-5.31;P<0.001). However, its interaction with null genotype of GSTT1 conferred significant reduced risk to lung cancer (OR=0.30;95%CI=0.10-0.91;P=0.03). Further in only tobacco smokers, null genotype was associated with increased reduced risk [0.03(0.001-0.78)0.03; Ptrend=0.006]. No effect modification of GSTM1 was observed with lung cancer risk by environmental risk factors. Interpretation & conclusions: The results suggest that absence of GSTT1 null genotype may be associated with a reduced risk of lung cancer and the effect remains unchanged after interaction with smoking.

Implication of DNA repair genes in prostate tumourigenesis in Indian males.

Background & objectives: Prostate cancer (CaP) is the fifth most common cancer among Indian men. Tumour protein p53 (TP53) gene increases the fidelity of DNA replication and homologous recombination by transcriptional transactivation of mismatch repair (MMR) genes. DNA repair thus has a potential role in molecular carcinogenesis of CaP. The aim of the present study was to identify mutations, and polymorphisms in TP53 gene and MMR protein expression in CaP in Indian male population. Methods: TP53 codon 72 polymorphism was analysed in 105 CaP, 120 benign prostatic hyperplasia (BPH) cases and 106 normal controls. Mutational analysis of TP53 was done in DNA extracted from formalin fixed paraffin embedded tissue of 80 CaP and 24 BPH cases. Expression of MMR proteins viz. hMLH1, hMSH2, hPMS1 and hPMS2 was studied in 80 CaP, 15 prostatic intraepithelial neoplasia (PIN) and 15 BPH cases. Results: A somatic C/A variation at the intronic boundary of exon 7 in TP53 gene was observed in one each biopsy samples from CaP and BPH. A significant association of codon 72 TP53 Pro/Pro genotype was observed with the risk of CaP (OR, 2.59, P=0.02) and BPH (OR, 6.27, P<0.001). Immunohistochemical analysis of MMR proteins showed maximum loss of hPMS1 expression in cases of CaP and PIN while no loss in expression of MMR proteins was observed in BPH cases. The study also identified a significant loss of hPMS2 protein in poorly differentiated tumours (Gleason score >7) than in well differentiated tumours (Gleason score 3-6) (P<0.05). Interpretation & conclusions: The results of the present study demonstrate that TP53 codon 72 polymorphism plays significant role in the pathogenesis and susceptibility to CaP and BPH. Also, an aberrant MMR protein expression could be involved in progression of prostate cancer through PIN, early CaP to aggressive CaP. The loss of hPMS2 protein expression may serve as a marker for progression of CaP.

Expression of androgen receptor in breast cancer & its correlation with other steroid receptors & growth factors.

Background & objectives: Breast cancer is the second most common malignancy in Indian women. Among the members of the steroid receptor superfamily the role of estrogen and progesterone receptors (ER and PR) is well established in breast cancer in predicting the prognosis and management of therapy, however, little is known about the clinical significance of androgen receptor (AR) in breast carcinogenesis. The present study was aimed to evaluate the expression of AR in breast cancer and to elucidate its clinical significance by correlating it with clinicopathological parameters, other steroid receptors (ER and PR) and growth factors receptors (EGFR and CD105). Methods: Expression of AR, ER, PR, epidermal growth factor receptor (EGFR) and endoglin (CD105) was studied in 100 cases of breast cancer by immunohistochemistry (IHC). Risk ratio (RR) along with 95% confidence interval (CI) was estimated to assess the strength of association between the markers and clinicopathological characteristics. Categorical principal component analysis (CATPCA) was applied to obtain new sets of linearly combined expression, for their further evaluation with clinicopathological characteristics (n=100). Results: In 31 cases presenting with locally advanced breast cancer (LABC), the expression of AR, ER, PR, EGFR and CD105 was associated with response to neoadjuvant chemotherapy (NACT). The results indicated the association of AR+ (P=0.001) and AR+/EGFR- (P=0.001) with the therapeutic response to NACT in LABC patients. The AR expression exhibited maximum sensitivity, specificity and likelihood ratio of positive and negative test. The present results showed the benefit of adding AR, EGFR and CD105 to the existing panel of markers to be able to predict response to therapy. Interpretation & conclusions: More studies on the expression profiles of AR+, AR+/CD105+ and AR+/EGFR- in larger set of breast cancer patients may possibly help in confirming their predictive role for therapeutic response in LABC patients.

Leydig cell tumor: An unusual presentation.

Leydig cell tumor is a benign tumor of the testis and malignant transformation, if present, is rare. The case presented here showed certain features of malignancy but no infiltration beyond the capsule or metastasis. The case could not be labeled as benign or malignant and patient is on follow-up. Differential diagnosis and clinical implications of a case in the borderline zone are discussed.

An unusual presentation of primary lymphoma of the thyroid in a young male patient.

Primary lymphoma of the thyroid gland is a rare disease that is strongly associated with chronic lymphocytic thyroiditis. We report an unusual case of 20-year-old male who presented with thyroid swelling and was diagnosed on cytology as lymphocytic thyroidtis. However, histopathologicalfindings revealed non-Hodgkin's lymphoma that was further classified as B-cell type on immunohistochemical stains. In cytology, lymphomas must be considered as a differential diagnosis in case of Hashimoto's/lymphocytic thyroiditis, subacute thyroiditis and small cell carcinoma.

Gastric outlet obstruction and cutaneous metastasis in adenocarcinoid tumor of stomach - unusual presentations with cytologic and ultra structural findings.

Neuroendocrine tumors, including carcinoids account for less than 1% of gastric tumors. Various subtypes of gastric carcinoids have been reported earlier. The present case deals with two unusual presentations, diagnosis and course of a gastric neuroendocrine tumor in an adult patient. A 35-years-old male initially presented with gastric outlet obstruction for an antral growth in the emergency ward. He underwent radical gastrectomy and was diagnosed with a gastric carcinoid tumor, on histopathology. After 6 months, he developed hepatic along with nodular cutaneous lesions over the scalp. Aspiration cytology (FNAC) from these metastatic lesions showed two distinct cell types with rosette formation. Ultrastructural findings showed neurosecretory granules in some cells. Subsequently, he underwent 2 cycles of chemotherapy. After a total duration of 9 months, he finally succumbed to the disease. We present a case of a gastric adenocarcinoid tumor, with 2 rare presentations. The metastatic lesions exhibited neuroendocrine features on cytology and electron microscopy.

Multiple eccrine hidrocystomas: report of two cases treated unsuccessfully with atropine ointment.

Eccrine hidrocystomas are rare, benign, cystic lesions with a lining that resembles that of the eccrine sweat gland and may be solitary or multiple. Multiple eccrine hidrocystomas occur predominantly on the face as asymptomatic, skin-colored to bluish lesions associated with a chronic course and seasonal variability. Treatment of multiple lesions on the face is challenging. Efficacy with atropine ointment is variable. Botulinum toxin and pulsed dye laser are reported to be beneficial. Two cases of multiple eccrine hidrocystomas are reported who showed no response to 1% atropine ointment.

Metaplastic carcinoma of the breast--a rare breast tumour.

Malignant breast neoplasms consisting of mixtures of epithelial and mesenchymal elements, are a rarity. Pathogenesis of such diverse elements within obviously infiltrating carcinomas has been the subject of much controversy. After the advent of immunohistochemistry, it is now generally accepted that metaplasia of the epithelial elements of a carcinoma gives these lesions their pseudosarcomatous appearance. Hence the name metaplastic carcinoma is given to malignant breast neoplasms which show Cytokeratin positivity in both epithelial and mesenchymal elements. We recently encountered such a case, which is being presented here along with relevant review of literature.