HAT-PCR Enables Sensitive Quantification of Minimal Residual Disease in Chronic Lymphocytic Leukemia and Myeloma

Hughes, E; Latham, S; Kuss, B; Grist, S; Hall, R; Khong, T; Gorniak, M; Spencer, A; Tam, C; Mulligan, S; Bailey, S; Sartor, M; Carney, D; Cull, G; Gottlieb, D; Morley, A

Author(s): Hughes, E; Latham, S; Kuss, B; Grist, S; Hall, R; Khong, T; Gorniak, M; Spencer, A; Tam, C; Mulligan, S; Bailey, S; Sartor, M; Carney, D; Cull, G; Gottlieb, D; Morley, A;
Details: Publication Year 2025-08-10,Volume 26,Issue #16,Page 7720
Journal Title: International Journal of Molecular Sciences
Publication Type: Research article
Abstract: The role of HAT-PCR (High A/T or High Annealing Temperature-PCR) in the quantification of minimal residual disease (MRD) was investigated in chronic lymphocytic leukemia (CLL) and myeloma. The IGH gene sequence was determined by next-generation sequencing (NGS), either by the Lymphotrack kit or by preparing libraries using an in-house two-round PCR protocol which enabled successful sequencing in 37/37 CLL marrow samples and 34/35 myeloma marrow samples. MRD was quantified by HAT-PCR in 125 CLL marrow or blood samples from 36 patients, with 2 results being less than 10(-6) and in 63 myeloma marrow samples from 35 patients, with 10 results being less than 10(-6). Measurement of MRD in 113 pairs of CLL samples and 51 pairs of myeloma samples showed that HAT-PCR was significantly more sensitive than flow. Compared to marrow MRD, blood MRD was relatively high in CLL but very low or undetectable in myeloma. Flow-positive HAT-PCR negative samples were not seen in myeloma, although the literature review suggested that flow-positive NGS-negative myeloma samples are sometimes observed. The ability of HAT-PCR to quantify down to and below 10(-6) and the practical advantages of PCR suggest that HAT-PCR could be used widely for the quantification of MRD in lymphoid malignancy.
Publisher: MDPI
Keywords: Humans; *Leukemia, Lymphocytic, Chronic, B-Cell/genetics/diagnosis/pathology; *Multiple Myeloma/genetics/diagnosis/pathology; *Neoplasm, Residual/genetics/diagnosis; *Polymerase Chain Reaction/methods; High-Throughput Nucleotide Sequencing; Immunoglobulin Heavy Chains/genetics; Aged; Female; Male; Middle Aged; Pcr; acute lymphoblastic leukemia; chronic lymphocytic leukemia; digital PCR; flow cytometry; measurable residual disease; minimal residual disease; myeloma; next generation sequencing
Department(s): Haematology
Publisher's Version: https://doi.org/10.3390/ijms26167720
Open Access at Publisher's Site: https://doi.org/10.3390/ijms26167720
Terms of Use/Rights Notice: Refer to copyright notice on published article.

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