Acute Myeloid Leukaemia with Gene Mutation: A Correlation with Haematological and Immunophenotypic Characteristics and Our Experience in a Tertiary Care Cancer Center in South India.

OBJECTIVE: Molecular genetic analysis of FLT3, NPM1, and CEBPA is already the standard of care in patients with acute myeloid leukaemia (AML) and represents the most frequent genetic alterations and important diagnostic and prognostic indicators. This study was undertaken to determine the frequency of FLT3 and NPM1 gene mutations in our institution and to characterize the association between gene mutations and haematological parameters as well as immunophenotypic features. MATERIAL AND METHOD: Morphological, haematological and immunophenotypic characteristics of NPM1 and FLT3 mutations in 126 patients of de novo AML including adults and children were studied. Apart from the French American British (FAB) method for classification, blasts were assessed for cuplike morphology as per strict definition for cuplike nuclei, ≥10% blasts with nuclear invaginations ≥25% of the nuclear area. RESULTS: FLT3 mutation in 31/126 (25%) and NPM1 mutation was found in 17/126 (13.4%) of the AML patients. 6 (5%) samples were positive for both NPM1 and FLT3/ITD mutations. Associations between the FLT3 and NPM1 gene mutations with haematological and immunophenotypic characteristics are reported. CONCLUSION: The results suggest that presence of distinct morphology and haematological and immunophenotypic characteristics together may serve as important indicators and surrogate for NPM1 and FLT3/ITD mutations. Further, comprehensive studies on the biological effects of NPM1 and FLT3/ITD mutations and their interactions with other genetic alterations are needed to gain insight into the molecular mechanism of these mutations involved in the pathogenesis of AML.

A 7 gene expression score predicts for radiation response in cancer cervix.

BACKGROUND: Cervical cancer is the most common cancer among Indian women. The current recommendations are to treat the stage IIB, IIIA, IIIB and IVA with radical radiotherapy and weekly cisplatin based chemotherapy. However, Radiotherapy alone can help cure more than 60% of stage IIB and up to 40% of stage IIIB patients. METHODS: Archival RNA samples from 15 patients who had achieved complete remission and stayed disease free for more than 36 months (No Evidence of Disease or NED group) and 10 patients who had failed radical radiotherapy (Failed group) were included in the study. The RNA were amplified, labelled and hybridized to Stanford microarray chips and analyzed using BRB Array Tools software and Significance Analysis of Microarray (SAM) analysis. 20 genes were selected for further validation using Relative Quantitation (RQ) Taqman assay in a Taqman Low-Density Array (TLDA) format. The RQ value was calculated, using each of the NED sample once as a calibrator. A scoring system was developed based on the RQ value for the genes. RESULTS: Using a seven gene based scoring system, it was possible to distinguish between the tumours which were likely to respond to the radiotherapy and those likely to fail. The mean score +/- 2 SE (standard error of mean) was used and at a cut-off score of greater than 5.60, the sensitivity, specificity, Positive predictive value (PPV) and Negative predictive value (NPV) were 0.64, 1.0, 1.0, 0.67, respectively, for the low risk group. CONCLUSION: We have identified a 7 gene signature which could help identify patients with cervical cancer who can be treated with radiotherapy alone. However, this needs to be validated in a larger patient population.

Prognostic significance of HPV physical status and integration sites in cervical cancer.

BACKGROUND: Human papilloma virus (HPV) infection is the major cause of cervical cancer and integration of HPV DNA into the host cell genome is believed to be essential for malignant transformation. MiRNAs are a class of 19-24 nt non-coding RNAs that regulate gene expression primarily through post transcriptional repression or m-RNA degradation in a sequence specific manner. The aim of this study was to determine the frequency of HPV16 and 18 integrated and episomal forms and to evaluate its prognostic significance in invasive cervical carcinoma cases and to detect by in-silico approach MiRNAs near HPV integration sites (within <3Mb). METHODS: HR-HPV 16 and 18 typing was performed by Nested Multiplex PCR (NMPCR) and HPV 16 and 18 physical status (integrated and episomal forms) was determined by Amplification of Papillomavirus Oncogene Transcripts (APOT) assay. Nested PCR products of the APOT assay were resolved on a 2% agarose gel and the PCR products of interest were excised and sequenced. In silico analysis was done to identify the Fragile sites and MiRNAs' near integration sites of the HPV. RESULTS: Episomal forms were more common with the HPV16 type and integrated forms with the HPV18 type (p= 0.011). Patients with tumors having the episomal forms had a better disease free survival than those with integrated forms of HPV16 type, but this did not reach statistical significance. We detected 53 miRNAs near integration sites, of which 39 have been reported to be associated with cancers. The incidence of miRNAs near HPV integration sites was 78.3%, being more common with HPV16. CONCLUSION: This is the first study from India to provide the physical status of HPV16 and HPV 18 in cervical cancers, to assess their prognostic importance and to identify FRA and MiRNAs' near HPV integration sites.