Molecular characterization of triple-negative myeloproliferative neoplasms by next-generation sequencing.

The role of next-generation sequencing (NGS) in identifying mutations in the driver, epigenetic regulator, RNA splicing, and signaling pathway genes in myeloproliferative neoplasms (MPNs) has contributed substantially to our understanding of the disease pathogenesis as well as disease evolution. NGS aids in determining the clonal nature of the disease in a subset of these disorders where mutations in the driver genes are not detected. There is a paucity of real-world data on the utility of this test in the characterization of triple-negative myeloproliferative neoplasms (TN-MPN). In this study, 46 samples of TN-MPN (essential thrombocythemia (ET) = 17; primary myelofibrosis (PMF) = 23; & myeloproliferative neoplasm unclassified (MPN-u) = 6) were screened for markers of clonality using targeted NGS. Among these, 25 (54.3%) patients had mutations that would help determine the clonal nature of the disease. Eight of the 17 TN-ET (47%) and 13 of the 23 TN-PMF (56.5%) patients had noncanonical mutations in the driver genes and mutations in the genes involved in epigenetic regulation. Identification of mutations categorized as high molecular markers (HMR) in 2 patients helped classify them as PMF with high risk according to the MIPSS 70 scoring system. A novel mutation in the MPIG6B (C6orf25) gene associated with childhood myelofibrosis was detected in a 14-year-old girl. The presence of clonal hematopoiesis could be confirmed in four of the six MPN-u patients in this cohort. This study demonstrates the utility of NGS in improving the characterization of TN-MPN by establishing clonality and detecting noncanonical mutations in driver genes, thereby aiding in clinical decision-making.

Serum exosomal miRNA as biomarkers for Retinoblastoma.

Retinoblastoma (RB) is a childhood eye tumor, caused by RB1 mutation. Though diagnosing RB is easier, prognosticating RB is limited to examining the patient under anesthesia and imaging technique. The aim of the study is to find exosomal miRNA biomarkers to prognosticate RB. Exosomes were isolated from one control - MIO-M1 and two RB cell lines - WERI-Rb-1 and NCC-RbC-51. Small RNA sequencing was performed on exosomal miRNA isolated from the three cell lines. miRNAs specific to each cell line were shortlisted. A total of 243, 606 and 400 miRNAs were identified in MIO-M1, WERI-Rb-1 and NCC-RbC-51 cell lines respectively. Nine miRNAs were shortlisted based on adjusted p value and literature, MIO-M1 specific (n = 1), WERI-RB-1 specific (n = 2), NCC-RbC-51 specific (n = 2) and miRNAs common to both RB cell lines (n = 4) were chosen. Validation was done using specific Taqman miRNA assays.miRNA validation was carried out on cell lines, cell line derived exosomes, primary RB tissues and exosomes isolated from serum of the RB patients. Validation of the miRNAs in cell lines and exosomes derived from the cell lines, confirmed the sequencing data. However, only 2 miRNAs - hsa-miR-301b-3p and hsa-miR-216b-5p were upregulated in the primary RB tissues. None of the miRNAs had significant expression in the serum exosomes of RB patients. Therefore, serum exosomal miRNA may not be ideal for prognosticating RB.Further research on other body fluids like CSF and vitreous could serve as potential source for biomarkers for prognosticating RB.

Pharmacokinetics and Efficacy of Generic Melphalan Is Comparable to Innovator Formulation in Patients With Multiple Myeloma Undergoing Autologous Stem Cell Transplantation.

BACKGROUND: High-dose melphalan (MEL) is the standard conditioning regimen used for autologous stem cell transplantation (ASCT) in patients with multiple myeloma (MM). Generic MEL is routinely used in various transplant centers across the world including ours due to its reduced cost and ease of availability. We compared the pharmacokinetics (PK) and the clinical efficacy of generic MEL with that of the innovator formulation in MM patients undergoing ASCT. PATIENTS AND METHODS: Sixty-three patients diagnosed with MM receiving high-dose MEL were included in this study. MEL levels in plasma were measured using a liquid chromatography tandem mass spectrometry (HPLC/MS-MS) protocol and non-linear mixed effects modeling was used to evaluate the PK of the data. RESULTS: The interindividual variability (IIV) in MEL area under the concentration versus time curve (AUC) and clearance (CL) were 4.39, 5.88-fold for generic, and 4.34, 6.85-fold for the innovator formulation, respectively. The median MEL AUC and CL were comparable between the 2 formulations. The population PK analysis showed age and creatinine CL as the only significant covariates explaining IIV in MEL AUC/CL. Analysis of MEL PK parameters with clinical outcome showed no significant differences in terms of onset and severity of mucositis, day to neutrophil and platelet engraftment, as well as response status on day 100 post ASCT between patients receiving generic or innovator formulations of MEL. In addition, neither MEL AUC nor CL was found to be associated with day +100 response. CONCLUSION: Our study suggests that the PK and efficacy of the generic MEL is comparable to the innovator formulation.