Clonal Characterization and Somatic Hypermutation Assessment by Next-Generation Sequencing in Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma: A Detailed Description of the Technical Performance, Clinical Utility, and Platform Comparison.

Somatic hypermutation status of the IGHV gene is essential for treating patients with chronic lymphocytic leukemia/small lymphocytic lymphoma. Unlike the conventional low-throughput method, assessment of somatic hypermutation by next-generation sequencing (NGS) has potential for uniformity and scalability. However, it lacks standardization or guidelines for routine clinical use. We critically assessed the performance of an amplicon-based NGS assay across 458 samples. Using a validation cohort (35 samples), the comparison of two platforms (Ion Torrent versus Illumina) and two primer sets [leader versus framework region 1 (FR1)] in their ability to identify clonotypic IGHV rearrangement(s) revealed 97% concordance. The mutation rates were identical by both platforms when using the same primer set (FR1), whereas a slight overestimation bias (+0.326%) was found when comparing FR1 with leader primers. However, for nearly all patients this did not affect the stratification into mutated or unmutated categories, suggesting that use of FR1 may provide comparable results if leader sequencing is not available and allowing for a simpler NGS laboratory workflow. In routine clinical practice (423 samples), the productive rearrangement was successfully detected by either primer set (leader, 97.7%; FR1, 94.7%), and a combination of both in problematic cases reduced the failure rate to 1.2%. Higher sensitivity of the NGS-based analysis also detected a higher frequency of double IGHV rearrangements (19.1%) compared with traditional approaches.

Defining the undetectable: The current landscape of minimal residual disease assessment in multiple myeloma and goals for future clarity.

Multiple Myeloma, the second most prevalent hematologic malignancy, yet lacks an established curative therapy. However, overall response rate to modern four-drug regimens approaches 100%. Major efforts have thus focused on the measurement of minute quantities of residual disease (minimal residual disease or MRD) for prognostic metrics and therapeutic response evaluation. Currently, MRD is assessed by flow cytometry or by next generation sequencing to track tumor-specific immunoglobulin V(D)J rearrangements. These bone marrow-based methods can reach sensitivity thresholds of the identification of one neoplastic cell in 1,000,000 (10-6). New technologies are being developed to be used alone or in conjunction with established methods, including peripheral blood-based assays, mass spectrometry, and targeted imaging. Data is also building for MRD as a surrogate endpoint for overall survival. Here, we will address the currently utilized MRD assays, challenges in validation across labs and clinical trials, techniques in development, and future directions for successful clinical application of MRD in multiple myeloma.

Marrow assessment for hemophagocytic lymphohistiocytosis demonstrates poor correlation with disease probability.

OBJECTIVES: To evaluate the amount of hemophagocytosis in 64 marrow core biopsy specimens and aspirates from 58 patients with clinical suspicion for secondary hemophagocytic lymphohistiocytosis (HLH) or reported findings of hemophagocytosis. METHODS: A review of medical records assigned patients to a low-risk (45 patients) or high-risk (13 patients) HLH group, and association with histologic findings was examined using the Fisher exact test. RESULTS: The amount of hemophagocytosis in aspirate or the core biopsy specimen did not correlate with disease probability (P = .17 and P = .63, respectively). Of the clinical/laboratory criteria assessed, the most significant correlations with HLH were highly elevated ferritin (P = .01), cytopenias (P = .02), and fever (P = .009). CONCLUSIONS: Our findings indicated that marrow histologic findings alone do not reliably predict the probability of HLH, and an isolated finding of hemophagocytosis, even when present in a high amount, lacks specificity for HLH.