Clinical Significance of Telomere Length and Associated Proteins in Oral Cancer

Purpose: Telomere shortening is an important event during carcinogenesis. Although studies suggest role of multiple proteins in telomere length regulation, there is dearth of reports in oral cancer which is a leading malignancy in Asian countries especially in India. Thus the present study was carried out to study these mechanisms and explore the pathways involved in telomere—telomerase regulation and identify possible prognostic markers to understand the biology of oral tumors for better treatment approaches. Methods: Telomere length was determined by Southern Hybridisation method, telomeric repeat binding factor (TRF) 1 and 2 expression was detected by Western blot method and telomerase activation by telomeric repeat amplifi cation protocol. Statistical analysis was done using SPSS (Version 10) software. Results: Significant shortening of telomeres was seen in the tumor tissues as compared to normal tissues. Poor prognosis was observed in the patients with higher telomere length in malignant tissue, higher tumor to normal telomere length ratio (T/N TRF LR). Expression of TRF-2 but not TRF-1 protein was signifi cantly higher in the malignant tissues. We also observed telomerase activation in 75 malignant tissues.Conclusions: Our results reveal signifi cant clinical usefulness of telomere length, T/N TRF LR and telomerase activation in the prognosis of oral cancer patients. TRF-2 overexpression in malignant tissues appears to play an important role in telomere length shortening in oral cancer. Abbreviations: TRF—Terminal restriction fragment; TRF-1—telomeric repeat binding factor-1; TRF-2—telomeric repeat binding factor-2; T/N TRF LR—Tumor/ Normal TRF length ratio

Towards the introduction of the ‘Immunoscore’ in the classification of malignant tumours

The American Joint Committee on Cancer/Union Internationale Contre le Cancer (AJCC/UICC) TNM staging system provides the most reliable guidelines for the routine prognostication and treatment of colorectal carcinoma. This traditional tumour staging summarizes data on tumour burden (T), the presence of cancer cells in draining and regional lymph nodes (N) and evidence for distant metastases (M). However, it is now recognized that the clinical outcome can vary significantly among patients within the same stage. The current classification provides limited prognostic information and does not predict response to therapy. Multiple ways to classify cancer and to distinguish different subtypes of colorectal cancer have been proposed, including morphology, cell origin, molecular pathways, mutation status and gene expression-based stratification. These parameters rely on tumour-cell characteristics. Extensive literature has investigated the host immune response against cancer and demonstrated the prognostic impact of the in situ immune cell infiltrate in tumours. A methodology named ‘Immunoscore’ has been defined to quantify the in situ immune infiltrate. In colorectal cancer, the Immunoscore may add to the significance of the current AJCC/UICC TNM classification, since it has been demonstrated to be a prognostic factor superior to the AJCC/UICC TNM classification. An international consortium has been initiated to validate and promote the Immunoscore in routine clinical settings. The results of this international consortium may result in the implementation of the Immunoscore as a new component for the classification of cancer, designated TNM-I (TNM-Immune). © 2013 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland

Role of nitric oxide and antioxidant enzymes in the pathogenesis of oral cancer

Background: Oral cancer is the leading malignancy in India. Nitric oxide and antioxidant enzymes play an important role in etiology of oral cancer. Therefore, the present study evaluates nitric oxide and antioxidant enzyme levels in healthy individual without tobacco habits (NHT, N=30) and healthy individuals with tobacco habits (WHT, n=90), patients with oral precancers (OPC, n=15) and oral cancer patients (n=126). Materials and Methods: Blood samples were collected from the subjects. NO 2 +NO 3 (nitrite+nitrate), superoxide dismutase (SOD) and catalase levels were estimated using highly specific spectrophotometeric methods. Statistical analysis was done by SPSS statistical software version 10. Results: Mean plasma NO 2 +NO 3 levels were elevated in patients with OPC and oral cancer patients as compared to the controls. Mean activities of erythrocyte SOD and catalase were higher in WHT than NHT. Erythrocyte SOD and catalase levels were higher in WHT and patients with OPC as compared to NHT. The erythrocyte SOD and catalase activities were lower in oral cancer patients than patients with OPC. The erythrocyte SOD activity was higher in advanced oral cancer than the early disease. Erythrocyte catalase activity was lower in poorly differentiated tumors than well and moderately differentiated tumors. Pearson\u2032s correlation analysis revealed that alterations in plasma NO 2 +NO 3 levels were negatively associated with changes in erythrocyte SOD activities. Conclusion: The data revealed that the alterations in antioxidant activities were associated with production of nitric oxide in oral cancer, which may have significant role in oral carcinogenesis

Role of nitric oxide and antioxidant enzymes in the pathogenesis of oral cancer

Background: Oral cancer is the leading malignancy in India. Nitric oxide and antioxidant enzymes play an important role in etiology of oral cancer. Therefore, the present study evaluates nitric oxide and antioxidant enzyme levels in healthy individual without tobacco habits (NHT, N=30) and healthy individuals with tobacco habits (WHT, n=90), patients with oral precancers (OPC, n=15) and oral cancer patients (n=126). Materials and Methods: Blood samples were collected from the subjects. NO 2 +NO 3 (nitrite+nitrate), superoxide dismutase (SOD) and catalase levels were estimated using highly specific spectrophotometeric methods. Statistical analysis was done by SPSS statistical software version 10. Results: Mean plasma NO 2 +NO 3 levels were elevated in patients with OPC and oral cancer patients as compared to the controls. Mean activities of erythrocyte SOD and catalase were higher in WHT than NHT. Erythrocyte SOD and catalase levels were higher in WHT and patients with OPC as compared to NHT. The erythrocyte SOD and catalase activities were lower in oral cancer patients than patients with OPC. The erythrocyte SOD activity was higher in advanced oral cancer than the early disease. Erythrocyte catalase activity was lower in poorly differentiated tumors than well and moderately differentiated tumors. Pearson′s correlation analysis revealed that alterations in plasma NO 2 +NO 3 levels were negatively associated with changes in erythrocyte SOD activities. Conclusion: The data revealed that the alterations in antioxidant activities were associated with production of nitric oxide in oral cancer, which may have significant role in oral carcinogenesis

Micronuclei and chromosomal aberrations in healthy tobacco chewers and controls: A study from Gujarat, India

Background: Tobacco chewing is attributed to oral cancer. Prediction of cancer development by genotoxicity analysis is a major challenge to identify tobacco users at greater risk. Therefore, present study aimed to analyze tobacco related genotoxic effects in chewers monitoring micronuclei (MN) and chromosome aberrations (CA). The biomarkers were compared with non chewer to (i) predict risk for genotoxicity, (ii) estimate synergistic effect of tobacco exposure with level of biomarkers, and (iii) identify best cellular site of measurements for genotoxicity assessment. Methods: Healthy tobacco chewers (n=47); and controls (n=48) were enrolled in the study. The peripheral blood lymphocyte and exfoliated buccal mucosa cells were studied for CA and micro nucleated cell count (MNC) respectively. An arbitrary unit was obtained for Lifetime Tobacco Exposure (LTE) using frequency/day multiplied by duration of years of tobacco use. Data were analyzed using SPSS statistical software. Results: MNC was significantly higher (p=0.001) in chewers than controls. CA was higher in chewers than controls. MNC can differentiate higher tobacco exposure in chewers than CA. Controls having MNC above cutoff level have greater risk of genotoxic exposition (95% C.I.; 1.462-23.26, p=0.012). Conclusion: The present study concludes that MNC is a better surrogate biomarker to predict genotoxicity than CA for tobacco exposure and DNA damage index in tobacco chewers

Detection of derivative 9 deletion by BCR-ABL fluorescence in-situ hybridization signal pattern to evaluate treatment response in CML patients

Background: To evaluate prognostic effect of submicroscopic deletions involving breakage and fusion points of the derivative chromosome 9 and 22 in chronic myeloid leukemia in untreated patients and their follow up samples to correlate with disease outcome. Methods: The study included 78 pretreatment (PT) samples from CML patients and 90 follow-up samples, classified as complete responders (CR, n=33), nonresponders (NR, n =54), and partial responder (PR, n=3) depending on the treatment status of the follow-up samples. Karyotype analysis was performed on metaphases obtained through short term cultures of bone marrow and blood. Detection of BCR-ABL fusion gene was performed using dual color dual fusion (D-FISH) translocation probes. Results: BCR-ABL fusion gene detection by D-FISH showed ABL-BCR deletion on derivative 9 in 47.8% of nonresponders which was higher as compared to pretreatment (11%). Mix D-FISH signal pattern was found in around 20% of pretreatment and non-responder samples. Average interval from chronic phase to blast crisis and accelerated phase was respectively 3.5 and 18 months and accelerated to blast crisis was 16.5 months from the time of diagnosis. The follow-up duration of 31 patients responded to therapy was significantly higher (p=0.0001) as compared to 45 patients who did not respond to therapy. Variant D-FISH signal pattern was seen at the time of diagnosis in patient who responded to therapy as well as those patients who did not respond to therapy. Conclusion: This is the first study from India reporting deletion in ABL, BCR, or ABL-BCR on derivative 9 did not correlate with response to therapy

Trisomy 8 in leukemia: A GCRI experience

Trisomy of chromosome 8 is frequently reported in myeloid lineage disorders and also detected in lymphoid neoplasms as well as solid tumors suggesting its role in neoplastic progression in general. It is likely to be a disease-modulating secondary event with underlying cryptic aberrations as it has been frequently reported in addition to known abnormalities contributing to clinical heterogeneity and modifying prognosis. Here, we share our findings of trisomy 8 in leukemia patients referred for diagnostic and prognostic cytogenetic assessment. Total 60 cases of trisomy 8, as a sole anomaly or in addition to other chromosomal aberrations, were reported (January 2005-September 2008). Unstimulated bone marrow or blood samples were cultured, followed by GTG banding and karyotyping as per the ISCN 2005. Patients with +8 were chronic myeloid leukemia (CML) (36), acute myeloid leukemia (AML) (17), and acute lymphoblastic leukemia (ALL) (7). In 7 patients, trisomy 8 was the sole anomaly, whereas in 6 patients +8 was in addition to normal clone, in 47 patients, the +8 was in addition to t(9;22), t(15;17), and others, including 3 with tetrasomy 8. Only one patient showed constitutional +8. The present study will form the basis of further cumulative studies to correlate potential differential effects of various karyotypic anomalies on disease progression and survival following a therapeutic regime. To unravel the role of extra 8 chromosome, constitutional chromosomal analysis and uniparental disomy will be considered

Trisomy 8 in leukemia: A GCRI experience

Trisomy of chromosome 8 is frequently reported in myeloid lineage disorders and also detected in lymphoid neoplasms as well as solid tumors suggesting its role in neoplastic progression in general. It is likely to be a disease-modulating secondary event with underlying cryptic aberrations as it has been frequently reported in addition to known abnormalities contributing to clinical heterogeneity and modifying prognosis. Here, we share our findings of trisomy 8 in leukemia patients referred for diagnostic and prognostic cytogenetic assessment. Total 60 cases of trisomy 8, as a sole anomaly or in addition to other chromosomal aberrations, were reported (January 2005–September 2008). Unstimulated bone marrow or blood samples were cultured, followed by GTG banding and karyotyping as per the ISCN 2005. Patients with +8 were chronic myeloid leukemia (CML) (36), acute myeloid leukemia (AML) (17), and acute lymphoblastic leukemia (ALL) (7). In 7 patients, trisomy 8 was the sole anomaly, whereas in 6 patients +8 was in addition to normal clone, in 47 patients, the +8 was in addition to t(9;22), t(15;17), and others, including 3 with tetrasomy 8. Only one patient showed constitutional +8. The present study will form the basis of further cumulative studies to correlate potential differential effects of various karyotypic anomalies on disease progression and survival following a therapeutic regime. To unravel the role of extra 8 chromosome, constitutional chromosomal analysis and uniparental disomy will be considered

Loss of sex chromosome in acute myeloid leukemia

Loss of sex chromosomes has been reported in normal and malignant marrows and its frequency increases with age in both situations. It is not clear whether the sex chromosome loss is a critical mutational event for neoplastic transformation or a genetic change related to ageing. The present study was undertaken to analyze incidence of loss of sex chromosomes in leukemia patients. Karyotypic analysis in bone marrow cells was carried out in total 270 AML patients registered at G.C.& R.I. during January 2000 to October 2003. Out of 270, 22 patients had loss of sex chromosome in addition to other disease specific chromosomal abnormalities. Out of 22 patients, 50% (11 of 22) were of the pediatric age (up to 14 years), and only 10% (3 of 22) patients were above the age of 50 years, maximum age being 65 years. On follow-up, only in patients with pathological remission normal 46XX/XY karyotypes were seen. Whereas in patients with persistent leukemic activity, clones with loss of sex chromosome were observed. The results indicate that sex chromosome loss in these cases may be equivalent of a clonal cytogenetic process rather than related to ageing process