The utility of the Ion Torrent PGM next generation sequencing for analysis of the most commonly mutated genes among patients with colorectal cancer in India

Background: The requirement for the mutation analysis for Kirsten rat sarcoma viral oncogene (KRAS) in colorectal cancer (CRC) is rapidly increasing as it is a predictive biomarker and also, its absence signifies response to anti?epidermal growth factor receptor (anti?EGFR) antibody treatment. The aim of our study was to investigate the pathological diagnosis and distribution of KRAS mutations in colorectal cancer with the use of next generation sequencing platform (Ion Torrent). Methods: A total of 56 CRC samples were tested to identify the genetic mutations, especially KRAS using the primers which included ~2800 COSMIC mutations of 50 oncogenes. Ion Torrent personal genome machine (semiconductor?based sequencing) was used for the sequencing and analysis. Along with KRAS, other 49 genes were also studied for COSMIC mutations. Results: KRAS mutation 25 (44.6%) had the highest frequency, followed by TP53 10 (17.9%) and PIK3CA mutation 4 (7.1%). Of all the KRAS mutations identified, mutations in codon 12 were most frequent followed by mutations in codon 13 and 61. The most frequent substitution was glycine to aspartate mutation in codon 12 (p.Gly12Asp) followed by glycine to valine (p.Gly12Val). Combinations of mutations were also studied. Our study revealed that seven cases (12.5%) had both KRAS and TP53 mutations (highest of all the combinations). Conclusion: The analysis of KRAS mutation frequency and its mutational subtype analysis in human CRCs by using semiconductor?based platform in routine clinical practices have been performed in Indian population. The findings were similar to earlier published reports from the Western literature.

Expression of CD71 by flow cytometry in acute leukemias: More often seen in acute myeloid leukemia.

Background: CD71 is a marker that has been usually used for identifying dysplasia in the erythroid series. We have tried to evaluate the expression of CD71 in various types of acute leukemias. Materials and Methods: We studied 48 patients of acute leukemia, of which 25 were acute myeloid leukemia (AML), 13 were precursor B‑acute lymphoblastic leukemia (B‑ALL), 8 were T‑ALL, and 2 were mixed phenotype acute leukemia (T/myeloid) as per the WHO classification. Results: We found that the expression of CD71 was most prevalent in AMLs (84%), followed by T‑ALL (50%) and least in B‑ALL (30%). Conclusion: This finding clearly shows the higher expression of CD71 in AMLs compared to other common type of leukemias, such as B‑ and T‑ALL. We suggest that the high expression of CD71 in AMLs could be used as a diagnostic marker and may also be used for minimal residual disease analysis after further studies in posttreatment scenario. This study is the first of its kind in the South Asian population.

Establishing biological reference intervals for novel platelet parameters (immature platelet fraction, high immature platelet fraction, platelet distribution width, platelet large cell ratio, platelet-X, plateletcrit, and platelet distribution width) and their correlations among each other.

Aims: This study aims to establish biological reference interval for novel platelet parameters. Settings and Design: A total of 945 healthy individuals, age ranges from 18 to 64 years (881 males and 64 females) coming for voluntary blood donation from June to August 2012 (3 months) were enrolled after exclusion of rejection criteria. Materials and Methods: The samples were assayed by running in complete blood count + reticulocyte mode on the Sysmex XE-2100 hematology analyzer and the reference interval for the population was calculated using Clinical and Laboratory Standards Institute guidelines. Statistical analysis used: Tests were performed using SPSS (Statistical Product and Service Solutions , developed by IBM corporation), version 13. Student t test and pearsons correlation analysis were also used. Results: The normal range for various parameters was platelet count: 150-520 × 103/cu mm, immature platelet fraction (IPF): 0.3-8.7%, platelet distribution width (PDW): 8.3-25.0 fL, mean platelet volume (MPV): 8.6-15.5 fL, plateletcrit (PCT): 0.15-0.62%, high immature platelet fraction (H-IPF): 0.1-2.7%, platelet large cell ratio (P-LCR): 11.9-66.9% and platelet-X (PLT-X) (ch): 11.0-22.0. Negative correlation was observed between platelet count (r = −0.468 to r = −0.531; P < 0.001) and PCT (r = −0.080 to r = −0.235; P < 0.05 to P < 0.001) with IPF, PDW, MPV, H-IPF, P-LCR, and platelet-X. IPF/H-IPF showed a positive correlation among them and also with PDW, MPV, P-LCR, platelet-X (r = +0.662 to r = +0.925; P < 0.001). Conclusions: These novel platelet parameters offer newer avenues in research and clinical use. Establishing biological reference interval for different platelet parameters would help determine true high and low values and help guide treatment decisions.

Testicular swelling: A rare manifestation of chronic lymphocytic leukemia presenting with Richter's syndrome.

Richter syndrome (RS) represents the clinico-pathologic transformation of indolent lymphomas to an aggressive lymphoma, most commonly diffuse large B-cell lymphoma. Majority of the patients have a previous diagnosis of Chronic Lymphocytic Leukemia and the median time to transformation is 2-4 years. De novo RS is extremely uncommon. RS frequently arises in the lymph nodes or bone marrow and rarely presents with extra nodal involvement, common sites being the gastrointestinal tract, eye, central nervous system, lung and kidney. Involvement of testis by RS is extremely rare and we came across only one such reported case in the literature. We are reporting this case as our patient presented with de novo RS at an extremely uncommon extra nodal site, testis.

Triple composite tumor of stomach: A rare combination of alpha fetoprotein positive hepatoid adenocarcinoma, tubular adenocarcinoma and large cell neuroendocrine carcinoma.

A 50-year-old male patient presented with pain abdomen of 6 months duration. Computed tomography scan revealed a large mass in the stomach occluding the lumen. Histopathology revealed a triple composite tumor comprising of tubular adenocarcinoma arising on a background of high-grade dysplasia, hepatoid adenocarcinoma (positive for Hep Par-1 and alpha fetoprotein) and large cell neuroendocrine carcinoma (positive for synaptophysin and chromogranin) with nodal metastasis.Triple composite tumors are distinctly rare with few reports in literature.

Detection of Hb variants and hemoglobinopathies in Indian population using HPLC: Report of 2600 cases.

Background: Inherited abnormalities of hemoglobin synthesis include a myriad of disorders ranging from thalassemia syndromes to structurally abnormal hemoglobin variants. Identification of these disorders is immensely important epidemiologically and aid in prevention of more serious hemoglobin disorders. Aims: High performance liquid chromatography (HPLC) forms an important tool for accurate and speedy diagnosis of various hemoglobin disorders. About 2600 cases have been studied for identification of various hemoglobin disorders in Indian population. Material and Methods: The study was performed on BIORAD VARIANT using beta thalassemia short program. Results and conclusion: Abnormal hemoglobin fractions on HPLC were seen in 327 of the 2,600 cases displayed. Of this, the beta thalassemia trait was the predominant abnormality with a total of 232 cases (8.9%). There were 15(0.6%) cases of beta thalassemia major and 16 of thalassemia intermedia. The rest comprised of Hb D Punjab (13 cases; 0.5%), Elevated Hb F (25 cases; 0.9%), Hb E (seven cases including two Hb E homozygous and five Hb E heterozygous), Double heterozygous Hb E-beta thal trait (six cases), Hb Q India (five cases), Double heterozygous Hb Q India -beta thal trait (two cases), Hb S (total cases three including one Hb S homozygous; two Hb S -beta thal trait) and one case each of Hb J Meerut, Hb D-Iran and Hb Lepore trait. Detection of this abnormal hemoglobin, particularly keeping in mind a high prevalence of Hb A2, will help in prevention of more serious hemoglobinopathies including beta thalassemia major. HPLC forms a rapid and accurate tool in early detection and management of various hemoglobin disorders.

Congenital fibrosarcoma of the upper extremity--a case report and review of literature.

Fibrosarcoma is rare in pediatric age group. Its diagnosis in infants may be difficult clinically and histologically. A twenty-day infant presented with a rapidly growing mass in the right forearm. Histologically a gray white fleshy mass was seen occupying the entire forearm and distal arm. The tumor showed sheets of closely packed spindle cells with minimal pleomorphism. The cells were positive for vimentin, negative jor smooth muscle actin, desmin and S-100. The diagnosis of congenital fibrosarcoma was made. Most cases of infantile fibrosarcomas present in the first year of life and although they have an alarming presentation, in children they have a much better prognosis as compared to adults. Diagnosis can be made on histology with immunohistochemistry.