High applicability of ASO-RQPCR for detection of minimal residual disease in multiple myeloma by entirely patient-specific primers/probes

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Epigenetic Inactivation of the miR-124-1 in Haematological Malignancies

miR-124-1 is a tumour suppressor microRNA (miR). Epigenetic deregulation of miRs is implicated in carcinogenesis. Promoter DNA methylation and histone modification of miR-124-1 was studied in 5 normal marrow controls, 4 lymphoma, 8 multiple myeloma (MM) cell lines, 230 diagnostic primary samples of acute myeloid leukaemia (AML), acute lymphoblastic leukaemia (ALL), chronic myeloid leukaemia (CML), chronic lymphocytic leukaemia (CLL), MM, and non-Hodgkin's lymphoma (NHL), and 53 MM samples at stable disease or relapse. Promoter of miR-124-1 was unmethylated in normal controls but homozygously methylated in 4 of 4 lymphoma and 4 of 8 myeloma cell lines. Treatment of 5-Aza-2′-deoxycytidine led to miR-124-1 demethylation and re-expression of mature miR-124, which also associated with emergence of euchromatic trimethyl H3K4 and consequent downregulation of CDK6 in myeloma cells harboring homozygous miR-124-1 methylation. In primary samples at diagnosis, miR-124-1 methylation was absent in CML but detected in 2% each of MM at diagnosis and relapse/progression, 5% ALL, 15% AML, 14% CLL and 58.1% of NHL (p<0.001). Amongst lymphoid malignancies, miR-124-1 was preferentially methylated in NHL than MM, CLL or ALL. In primary lymphoma samples, miR-124-1 was preferentially hypermethylated in B- or NK/T-cell lymphomas and associated with reduced miR-124 expression. In conclusion, miR-124-1 was hypermethylated in a tumour-specific manner, with a heterochromatic histone configuration. Hypomethylation led to partial restoration of euchromatic histone code and miR re-expression. Infrequent miR-124-1 methylation detected in diagnostic and relapse MM samples showed an unimportant role in MM pathogenesis, despite frequent methylation found in cell lines. Amongst haematological cancers, miR-124-1 was more frequently hypermethylated in NHL, and hence warrants further study

Leptomeningeal leukemia masquerading as cauda equina syndrome: Appraisal by magnetic resonance imaging

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Cerebral leukostasis manifesting as multifocal intracerebral hemorrhage

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Study of gene promoter methylation in acute promyelocytic leukaemia

published_or_final_versiontocabstractMedicineMasterDoctor of Medicin

The role of aberrant gene promoter methylation in multiple myeloma

abstractpublished_or_final_versionMedicineDoctoralDoctor of Philosoph

Disease detection by next-generation sequencing in multiple myeloma

Copyright © 2019 Yao, Bai, Orfao and Chim.Next-generation sequencing (NGS) has been applied to monitor minimal residual disease (MRD) in multiple myeloma (MM). Standardized DNA input and sequencing depth is essential for achieving a uniform sensitivity in NGS-based MRD study. Herein, the sensitivity of 10−5 was verified by a standardized experimental design based on triplicate measurements of 1 μg DNA input and 1 million sequencing reads using the LymphoTrack-MiSeq platform. MRD level was defined as the mean MRD burden of the triplicates. Two spike-in controls at concentrations of 0.001% tumor plasma cells (PC) for verifying the sensitivity of 10−5 and 0.01% (or 0.005%) for MRD normalization were systematically analyzed. The spike-in control of 0.001% MRD was consistently detected in all samples, confirming a sensitivity of 10−5. Moreover, this standardized NGS approach yielded MRD measurements concordant with serological response and comparable to allele-specific oligonucleotide (ASO) real-time quantitative (RQ)-PCR. Moreover, NGS showed an improved sensitivity and provided quantification of MRD for cases assigned “positive but not quantifiable” (PNQ) by ASO RQ-PCR, even without the use of patient-specific probes/primers. Issues regarding the specificity of myeloma-specific sequences as MRD target, optimal input for spike-in normalization, and interpretation of MRD from triplicates are discussed. Herein, the standardized LymphoTrack-MiSeq-based method is verified to carry a sensitivity of 10−5, hence an effective tool for MRD monitoring in MM. As only a small number of samples are tested here, further study with a larger number of patients is warranted.This work was supported by Hong Kong Blood Cancer Foundation and National Natural Science Foundation of China (81470369) awarded to CC

Aberrant gene methylation implicated in the progression of monoclonal gammopathy of undetermined significance to multiple myeloma

Malignant transformation is a multistep process that may involve dysregulation of oncogenes and tumour suppressor genes, and monoclonal gammopathy of undetermined significance (MGUS) is believed to be a precursor of multiple myeloma. To investigate whether aberrant promoter methylation might be involved in the evolution of MGUS to multiple myeloma, we examined the p16, protein tyrosine phosphatase, non‐receptor type 6 (SHP1), death‐associated protein (DAP) kinase, E‐cadherin and oestrogen receptor genes, most being tumour suppressor genes, by methylation‐specific polymerase chain reaction. In 32 cases of multiple myeloma and 19 cases of MGUS, significantly more frequent methylation of p16 (p = 0.001), SHP1 (p⩽0.001) and E‐cadherin (p⩽0.001) genes was found in multiple myeloma than in MGUS. Methylation of DAP kinase and oestrogen receptor genes was comparable in multiple myeloma and MGUS. In conclusion, methylation of p16, SHP1 and E‐cadherin genes might be involved in the progression of MGUS to multiple myeloma