Search

While molecular testing of hematologic malignancies is now standard of care, there is variability in practice and testing capabilities between different academic laboratories, with common questions arising on how to best meet clinical expectations. A survey was sent to hematopathology subgroup members of the Genomics Organization for Academic Laboratories consortium to assess current and future practice and potentially establish a reference for peer institutions. Responses were received from 18 academic tertiary-care laboratories regarding next-generation sequencing (NGS) panel design, sequencing protocols and metrics, assay characteristics, laboratory operations, case reimbursement, and development plans. Differences in NGS panel size, use, and gene content were reported. Gene content for myeloid processes was reported to be generally excellent, while genes for lymphoid processes were less well covered. The turnaround time (TAT) for acute cases, including acute myeloid leukemia, was reported to range from 2 to 7 calendar days to 15 to 21 calendar days, with different approaches to achieving rapid TAT described. To help guide NGS panel design and standardize gene content, consensus gene lists based on current and future NGS panels in development were generated. Most survey respondents expected molecular testing at academic laboratories to continue to be viable in the future, with rapid TAT for acute cases likely to remain an important factor. Molecular testing reimbursement was reported to be a major concern. The results of this survey and subsequent discussions improve the shared understanding of differences in testing practices for hematologic malignancies between institutions and will help provide a more consistent level of patient care.

Zhu‐Tokita‐Takenouchi‐Kim (ZTTK) syndrome is caused by de novo loss‐of‐function variants in the SON gene (MIM #617140). This multisystemic disorder is characterized by intellectual disability, seizures, abnormal brain imaging, variable dysmorphic features, and various congenital anomalies. The wide application and increasing accessibility of whole exome sequencing (WES) has helped to identify new cases of ZTTK syndrome over the last few years. To date, there have been approximately 45 cases reported in the literature. Here, we describe 15 additional individuals with variants in the SON gene, including those with missense variants bringing the total number of known cases to 60. We have reviewed the clinical and molecular data of these new cases and all previously reported cases to further delineate the most common as well as emerging clinical findings related to this syndrome. Furthermore, we aim to delineate any genotype–phenotype correlations specifically for a recurring pathogenic four base pair deletion (c.5753_5756del) along with discussing the impact of missense variants seen in the SON gene. [ABSTRACT FROM AUTHOR]

Objective: Mitochondrial DNA (mtDNA) depletion/deletions syndrome (MDDS) comprises a group of diseases caused by primary autosomal defects of mtDNA maintenance. Our objective was to study the etiology of MDDS in 4 patients who lack pathogenic variants in known genetic causes., Methods: Whole exome sequencing of the probands was performed to identify pathogenic variants. We validated the mitochondrial defect by analyzing mtDNA, mitochondrial dNTP pools, respiratory chain activities, and GUK1 activity. To confirm pathogenicity of GUK1 deficiency, we expressed 2 GUK1 isoforms in patient cells., Results: We identified biallelic GUK1 pathogenic variants in all 4 probands who presented with ptosis, ophthalmoparesis, and myopathic proximal limb weakness, as well as variable hepatopathy and altered T-lymphocyte profiles. Muscle biopsies from all probands showed mtDNA depletion, deletions, or both, as well as reduced activities of mitochondrial respiratory chain enzymes. GUK1 encodes guanylate kinase, originally identified as a cytosolic enzyme. Long and short isoforms of GUK1 exist. We observed that the long isoform is intramitochondrial and the short is cytosolic. In probands' fibroblasts, we noted decreased GUK1 activity causing unbalanced mitochondrial dNTP pools and mtDNA depletion in both replicating and quiescent fibroblasts indicating that GUK1 deficiency impairs de novo and salvage nucleotide pathways. Proband fibroblasts treated with deoxyguanosine and/or forodesine, a purine phosphatase inhibitor, ameliorated mtDNA depletion, indicating potential pharmacological therapies., Interpretation: Primary GUK1 deficiency is a new and potentially treatable cause of MDDS. The cytosolic isoform of GUK1 may contribute to the T-lymphocyte abnormality, which has not been observed in other MDDS disorders. ANN NEUROL 2024;96:1209-1224., (© 2024 The Author(s). Annals of Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association.)

Importance: Broad range assay for accurate and sensitive diagnostics., Competing Interests: The authors declare no conflict of interest.

Aims: Targeted next-generation sequencing (NGS) panels, which identify genomic alterations, are the stronghold of molecular oncology laboratories. In spite of technological advances, the quantity and quality of DNA from formalin-fixed paraffin-embedded tissue and paucicellular specimens are barriers to successful sequencing. Here, we describe an NGS assay employing single tube stem-loop inhibition mediated amplification technology that delivers highly accurate results with low input DNA. Rigorous quality metrics, regular monitoring and in-depth validation make the test attractive for clinical laboratories., Methods: The study used a customised NGS panel, targeting 48 genes across several solid tumour types. Validation, in accordance with guidelines from New York State, sequenced patient samples harbouring 136 known variants, including single-nucleotide variants (SNVs) and indels. Specimen types included formalin-fixed paraffin embedded blocks, core biopsies and cytology material. Neoplastic cellularity of the tumours ranged from 10% to 80%., Results: The assay was highly specific and sensitive with excellent accuracy, reproducibility and repeatability/precision. Concordant results for identification of SNVs and indels were obtained from specimens with DNA input of 2-3 ng, tissue with 10% neoplastic cellularity and variant allelic frequencies of 2.5%-3%. Over 99% of the target areas are shown to achieve at least 500X coverage when parsed through two bioinformatics pipelines. With over 2000 clinical specimens analysed, the success of the panel for reporting of results is 95.3% CONCLUSIONS: The advanced technology enables accurate identification of clinically relevant variants with uniformity of coverage and an impressive turn-around-time. The overall workflow and cost-effectiveness provide added value., Competing Interests: Competing interests: HF has received consulting fees from Opentrons Labworks and PIllar Biosciences. SH has received consulting fees from Opentron Labworks and Loxo Oncology and honoraria from Illumina and Medscape., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Plasmablastic lymphoma (PBL) is a rare and clinically aggressive neoplasm that typically occurs in immunocompromised individuals, including those with HIV infection and solid organ allograft recipients. Most prior studies have focused on delineating the clinicopathologic features and genetic attributes of HIV-related PBLs, where MYC deregulation and EBV infection, and more recently, mutations in JAK/STAT, MAP kinase, and NOTCH pathway genes have been implicated in disease pathogenesis. The phenotypic spectrum of post-transplant (PT)-PBLs is not well characterized and data on underlying genetic alterations are limited. Hence, we performed comprehensive histopathologic and immunophenotypic evaluation and targeted sequencing of 18 samples from 11 patients (8 males, 3 females, age range 12-76 years) with PT-PBL; 8 de novo and 3 preceded by other types of PTLDs. PT-PBLs displayed morphologic and immunophenotypic heterogeneity and some features overlapped those of plasmablastic myeloma. Six (55%) cases were EBV+ and 5 (45%) showed MYC rearrangement by fluorescence in situ hybridization. Recurrent mutations in epigenetic regulators (KMT2/MLL family, TET2) and DNA damage repair and response (TP53, mismatch repair genes, FANCA, ATRX), MAP kinase (KRAS, NRAS, HRAS, BRAF), JAK/STAT (STAT3, STAT6, SOCS1), NOTCH (NOTCH1, NOTCH3, SPEN), and immune surveillance (FAS, CD58) pathway genes were observed, with EBV+ and EBV- cases exhibiting similarities and differences in their mutational profiles. Clinical outcomes also varied, with survival ranging from 0-15.9 years postdiagnosis. Besides uncovering the biological heterogeneity of PT-PBL, our study highlights similarities and distinctions between PT-PBLs and PBLs occurring in other settings and reveals potentially targetable oncogenic pathways in disease subsets.