Microsatellite instability in young patients with mucinous colorectal cancers – characterization using molecular testing, immunohistochemistry, and histological features

CONTEXT: The incidence of colorectal cancers (CRCs) in young Indian patients is higher than the international average. CRCs in young patients are commonly of mucinous type and show microsatellite instability (MSI). AIMS: To ascertain the MSI status of mucinous CRCs in patients ≤40 years of age by molecular testing and to correlate this with immunohistochemical (IHC) analysis and tumor histology. SUBJECTS AND METHODS: Archived formalin-fixed paraffin embedded tissue blocks of 30 young mucinous CRC patients were retrieved. MSI testing was done using two mononucleotide markers – BAT26 and NR24. IHC analysis was done using MLH1, MSH2, and MSH6. Histological features of all cases were studied. Data were analyzed using the SPSS software and the Pearson's chi-square test and Fisher's exact test. RESULTS: Eight out of 30 cases (26.7%) showed MSI by molecular testing. IHC identified seven of these cases. Histological features showing a statistically significant association with MSI were the presence of a well-differentiated adenocarcinoma component (P = 0.003), peritumoral lymphocytes (P = 0.002) and tumor budding (P = 0.021). CONCLUSION: The detection of defective mismatch repair (MMR) proteins using IHC for MLH1, MSH2, and MSH6 and molecular testing using BAT26 and NR24 appears to be a good protocol to detect CRCs with MSI. Histology could be useful in identifying cases that require screening for presence of MMR protein defects

V-raf murine sarcoma viral oncogene homolog B (BRAF) mutations in hairy cell leukaemia.

Introduction: Hairy cell leukemia (HCL) is a B-cell non-Hodgkin lymphoma with distinct clinical, morphological and immunophenotypic features; however, there are many other B-cell lymphomas, which closely mimic HCL. Accurate diagnosis of HCL is important as treatment with 2-chloro-2’-deoxyadenosine (cladribine) is associated with >80% chance of complete cure. The recent description of BRAF p.V600E mutations in almost all HCL cases in various studies has not only improved the pathogenetic understanding of this entity but also increased the diagnostic accuracy of this disorder. Aim: The aim of the study was to standardize a molecular test for diagnosis of HCL and compare with standard established morphological, cytochemical and immunophenotypic parameters for HCL diagnosis. Materials and Methods: The incidence of this mutation was sought in 20 patients with either classical HCL or HCL variant (HCLv) by Sanger sequencing and allele-specifi c polymerase chain reaction. BRAF p.V600E mutation was present in all HCL cases and absent in the only HCLv case. Results: A high degree of correlation was noted between the presence of BRAF p.V600E and established diagnostic criteria in 20/20 patients with HCL/ HCLv. Our data supports the observation that this mutation is present in all cases of HCL and is absent in HCLv. Hence, detection of the BRAF p. V600E mutation can be a useful adjunct in the diagnostic algorithm.

Lennert's lymphoma: Clinicopathological profile of five cases.

Background and Aim: Lennert’s lymphoma is a rare variant of peripheral T-cell lymphoma (PTCL) not otherwise specifi ed (NOS) rich in epithelioid histiocytes. This study aims to analyze the clinical, morphologic, and immunophenotypic profi le of cases of Lennert’s lymphoma from our country and determines the utility of T-cell receptor (TCR) gene rearrangement in the diagnosis. Materials and Methods: All cases diagnosed as Lennert’s lymphoma during the period of January 2001 to August 2011 were included in this study. Hematoxylin and eosin (H and E) stained slides and immunohistochemistry results were analyzed and TCR gene rearrangement was performed. Results: There were fi ve cases of Lennert’s lymphoma diagnosed in our institution during this period, which included two males and three females. All cases showed effacement of lymph node architecture by diffuse infi ltration of small lymphoid T cells [CD3+, CD4+, CD8+, T-cell intracellular antigen 1 (TIA-1+), Granzyme B−] and clusters of epithelioid histiocytes throughout the lymph node and scattered large transformed cells (CD20−, CD30+, CD15−/+). TCR rearrangement was done in three cases by polymerase chain reaction (PCR) and showed the presence of a clonal T-cell population. Conclusions: Lennert’s lymphoma constituted 0.11% of all non-Hodgkin lymphomas (NHLs) in our institution. Differentiation from classical Hodgkin’s lymphoma is sometimes diffi cult by morphology and immunohistochemistry alone and TCR gene rearrangement was extremely useful in diagnosis.

Discriminating thyroid cancers from benign lesions based on differential expression of a limited set of miRNA using paraffin embedded tissues.

Background : Micro-RNAs (miRNAs) are expressed in a tissue-specific manner and are known to demonstrate differential expression even among the various subtypes of a given tumor. This differential expression has been harnessed successfully in the development of diagnostic assays for various malignant tumors. These assays have been found to be relevant and of value as additional diagnostic tools even among thyroid tumors, particularly with regard to thyroid carcinomas of follicular morphology. Materials and Methods : A limited set of miRNA have been assessed as part of this study in an effort to use minimal number of miRNA markers (miR-187, miR-221, miR-222, and miR-224) to differentiate the benign from the malignant thyroid tumors using miRNA derived from paraffin embedded material. Results : While miR-221 and miR-222 were found to provide good accuracy as individual markers (86% and 84%), a combination of the two provided slightly better accuracy (91%). Both miR-221 and 222 were able to significantly differentiate malignant tumors from the benign samples (P< 0.001) individually and as a combination of markers. However, inclusion of miR-187 and miR-224 in the panel did not provide any additional benefit. Conclusion : While a combination of miR-221 and 222 when used in a diagnostic panel could provide fairly good accuracy additional markers may need to be investigated to augment their diagnostic utility.

Microbes in the gut: a digestable account of host-symbiont interactions.

The human bowel is host to a diverse group of bacteria with over 500 different bacterial species contributing to this diversity. Until recently these bacteria were regarded as residents without any specific functions. The last two decades have seen a radical change in our understanding of the interactions between the gut flora and their eukaryotic hosts and there is a growing appreciation of the spectrum of functions performed by these symbionts. Intestinal bacteria are recognized for their role in nutrient absorption, mucosal barrier function, angiogenesis, morphogenesis and postnatal maturation of intestinal cell lineages, intestinal motility and more importantly maturation of gut associated lymphoid tissue (GALT). Although gut flora are implicated in certain pathological disorders, their remarkable contributions to health and homeostasis of the host need to be recognized and understood.

Microbes in the gut A digestable account of host-symbiont interactions.

The human bowel is host to a diverse group of bacteria with over 500 different bacterial species contributing to this diversity. Until recently these bacteria were regarded as residents without any specific functions. The last two decades have seen a radical change in our understanding of the interactions between the gut flora and their eukaryotic hosts and there is a growing appreciation of the spectrum of functions performed by these symbionts. Intestinal bacteria are recognized for their role in nutrient absorption, mucosal barrier function, angiogenesis, morphogenesis and postnatal maturation of intestinal cell lineages, intestinal motility and more importantly maturation of gut associated lymphoid tissue (GALT). Although gut flora are implicated in certain pathological disorders, their remarkable contributions to health and homeostasis of the host need to be recognized and understood.

Comparison of disc diffusion & E test methods with agar dilution for antimicrobial susceptibility testing of Haemophilus influenzae.

BACKGROUND & OBJECTIVES: Reliable methods of detection of antimicrobial resistance are of paramount importance in the treatment and management of infections caused by Haemophilus influenzae. The objective of the present study was to compare and evaluate the performance of disc diffusion and E test (Epsilometer test) with agar dilution method for antimicrobial susceptibility testing of H. influenzae. METHODS: A total of 46 isolates of H. influenzae from various invasive sites were included as test strains. Antimicrobial susceptibility testing was performed by Kirby-Bauer disc diffusion method for ampicillin, chloramphenicol, trimethoprim-sulphamethoxazole (TMP-SMZ) and cefotaxime. Minimum Inhibitory Concentration (MIC) determination was performed by E test and agar dilution for the same set of antimicrobials. All tests were performed on Haemophilus test medium (HTM). RESULTS: Disc diffusion showed a very major (2%) and minor (4%) interpretative error with TMP-SMZ and minor interpretative errors to ampicillin (13%) and chloramphenicol (24%) when compared to agar dilution method. E test produced only minor interpretative errors to chloramphenicol (7%) and TMP-SMZ (2%) and no interpretative errors with ampicillin and cefotaxime as against agar dilution. E test showed good agreement with agar dilution for each of the antimicrobial tested. INTERPRETATION & CONCLUSION: Disc diffusion test may be used as a preliminary screen for susceptibility testing of H. influenzae. E test is simple, easy to perform and a reliable method for determination of resistance in H. influenzae. However its cost and limited availability in India may limit its use. The reference agar dilution method can be used reliably in routine susceptibility testing of H. influenzae.