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Molecular diagnosis of myeloid neoplasms (MN) is based on the detection of multiple genetic alterations using various techniques. Next-generation sequencing (NGS) has been proved as a useful method for analyzing many genes simultaneously. In this context, we analyzed diagnostic samples from 121 patients affected by MN and ten relapse samples from a subset of acute myeloid leukemia patients using two enrichment-capture NGS gene panels. Pathogenicity classification of variants was enhanced by the development and application of a custom onco-hematology score. A total of 278 pathogenic variants were detected in 84% of patients. For structural alterations, 82% of those identified by cytogenetics were detected by NGS, 25 of 31 copy number variants and three out of three translocations. The detection of variants using NGS changed the diagnosis of seven patients and the prognosis of 15 patients and enabled us to identify 44 suitable candidates for clinical trials. Regarding AML, six of the ten relapsed patients lost or gained variants, comparing with diagnostic samples. In conclusion, the use of NGS panels in MN improves genetic characterization of the disease compared with conventional methods, thus demonstrating its potential clinical utility in routine clinical testing. This approach leads to better-adjusted treatments for each patient.

Chimerism refers to the relative proportion of donor and recipient DNA after hematopoietic stem cell transplantation (HSCT) and its quantitative follow-up is of great clinical utility in this setting. PCR of short tandem repeats (STR-PCR) constitutes the gold standard method for chimerism quantification, although more sensitive PCR techniques (such as qPCR) have recently arisen. We compared the sensitivity and the quantification capacity of both techniques in patient samples and artificial mixtures and demonstrated adequate performance of both methods, with higher sensitivity of qPCR and better quantification skills of STR-PCR. By qPCR, we then prospectively followed up 57 patients that were in complete chimerism (CC) by STR-PCR. Twenty-seven patients (59%) showed 0.1&ndash, 1% recipient DNA in the bone marrow. Only 4 patients presented 0.1&ndash, 1% recipient DNA in peripheral blood (PB), and one of them relapsed. Finally, by qPCR, we retrospectively studied the last sample that showed CC by STR-PCR prior to relapse in 8 relapsed patients. At a median of 59 days prior to relapse, six patients presented mixed chimerism by qPCR in PB. Since both approaches have complementary characteristics, we conclude that different techniques should be applied in different clinical settings and therefore propose a methodological algorithm for chimerism follow-up after HSCT.