2021年 · 第101卷第13期

Chromosome-positive acute lymphocytic leukemia (Ph⁺ALL) patients, BCR-ABL1 kinase domain (KD) point mutations are the most common mechanism of tyrosine kinase inhibitor (TKIs) resistance. First-generation Sanger gene sequencing is currently the most commonly used method to screen for BCR-ABL1 KD mutations, but it has certain limitations, that is, the detection sensitivity is poor, and the composite mutations cannot be reliably identified. Second-generation gene sequencing (NGS) technology can overcome these problems. More and more diagnostic laboratories are using NGS detection as a routine application, and it is expected to become the preferred method for BCR-ABL1 KD mutation screening. In this article published by Cancer Medicine, expert consensus-based application recommendations are published on the potential value and indications of NGS in the detection of BCR-ABL1 KD resistance mutations before and during first-line TKIs treatment, in relapsed/refractory cases, and before and after allogeneic hematopoietic stem cell transplantation, and how the test results of NGS affect treatment decisions. This application proposal agrees and has been well documented to prove that the sensitivity of NGS to analyze BCR-ABL1 KD resistance mutations is significantly better than that of Sanger sequencing, and it can directly identify composite mutations; A set of NGS analysis methods and minimum technical and methodological requirements for results reporting are also given. This application proposal has good reference significance for guiding the practical application of NGS in the detection of BCR-ABL1 KD resistance mutation. On the other hand, the article also highlights the strong need for further prospective studies on BCR-ABL1 KD resistance mutations. The publication of this application recommendation will contribute to the promotion and rational application of NGS in the detection of BCR-ABL1 KD resistance mutations.

RUNX1 mutations are widespread in patients with myelodysplastic syndrome (MDS), and most reports show that RUNX1 mutations are associated with poor prognosis. However, differences still exist, and the results of univariate analysis cannot be confirmed by multivariate analysis in some cases. Therefore, this meta-analysis assessed the impact of RUNX1 mutations on the prognosis of patients with MDS. This meta-analysis study data was obtained from eligible studies from the PubMed, Embase, and Cochrane libraries. The hazard ratios for overall survival (OS) and leukemia-free survival (LFS) were calculated from the multivariate Cox proportional hazard model (HR) and its 95% confidence interval (CI）。 The study included a total of 16 studies with 5,422 patients, including 617 patients with a RUNX1 mutation and 4,805 patients with wild-type RUNX1. Results show that the total of OSHRwas 1.43 (95%CI=1.21~1.70，P<0.001), while the correspondence of LFSHRwas 1.88 (95%CI=1.42~2.51，P<0.001）。 These results suggest that RUNX1 mutations are associated with poor prognosis and shortened survival in patients with MDS. In addition, stem cell transplantation can alleviate the impact of poor prognostic factors in patients. Patients carrying these mutations should be given priority to aggressive treatment.

With the deepening of genomic research, more and more evidence shows that some patients with hematological tumors have clear genetic predisposition factors. In the updated WHO classification criteria for hematopoietic and lymphoid tissue tumors (2017 edition), "myeloid tumors with germline factors" have been listed as a separate disease classification. Germline susceptibility genes for childhood acute lymphoblastic leukemia (ALL) are also increasingly reported. In recent years, it has been found that the germline variation of ETV6 gene is associated with the risk of ALL, and it has been suggested as a new leukemia susceptibility syndrome. To clarify the role of ETV6 variants in the pathogenesis of ALL, the study performed a comprehensive analysis of 32 cases of childhood leukemia caused by this rare syndrome. A total of 34 non-synonymous ETV6 germline variants were detected in this study, and 22 of them were identified as causing impaired transcriptional repressive activity, loss of DNA binding function, and abnormal nuclear localization. The missense variant ETV6 protein retains the ability to form a dimer with the wild-type ETV6 protein and may exert a dominant negative effect. Whole transcriptome and whole genome sequencing analyses of this group of patients also revealed that ETV6 germline variants had a significant effect on the leukemia transcriptome, and that this group of patients had different patterns of somatic concomitant mutations. Among the cases carrying deleterious ETV6 germline variants, 70% had a hyperdiploid karyotype with characteristic NRAS, KRAS and PTPN11 mutations. The other 30% are diploid genomes with very high frequency of PAX5 and ETV6 copy number deletions, and their gene expression patterns are very similar to those of ALL with positive ETV6-RUNX1 fusion genes. One patient carrying the ETV6 R386fs variant developed ALL, followed by treatment-related acute myeloid leukemia (AML); Another family of carriers of the ETV6 R399H variant developed ALL and AML, respectively. Genomic analysis showed support for concomitant somatic variation determining the lineage of leukemia. In vitro studies have shown impaired tumor suppressor activity of variant ETV6. ETV6-mediated leukemia also shows a complex interaction between hereditary and acquired genomic variation in pathogenesis. The genetic predisposition factors of hematological tumors are more common than previously expected, and they have important clinical diagnosis and treatment significance, which should be paid attention to.

The tumor protein p53 (TP53) is the most common mutant gene in cancer. In patients with myelodysplastic syndrome (MDS), TP53 mutations have been associated with high-risk disease, rapid conversion to acute myeloid leukemia (AML), resistance to traditional therapies, and poor prognosis. Consistent with the tumor suppressor effect of TP53, patients carried both monoallelic and biallelic mutations. However, the biological and clinical implications of TP53 allele status have not been comprehensively studied in MDS or any other cancer type. The study analyzed TP53 mutations and allelic imbalances in 3 324 patients with MDS. 1/3 of the patients with TP53 mutations had single allelic mutations and 2/3 had multiple mutations (multi-site mutations), consistent with bi-allelic targeting. Established associations with complex karyotypes, few co-occurring mutations, high-risk manifestations, and poor prognosis are only for patients with multisite mutations. TP53 multi-site mutation status is independent of the revised International Prognostic Scoring System (IPSS-R) and predicts the risk of MDS death and leukemia transformation. However, there were no statistically significant differences in outcome and response to treatment between monoallelic patients and TP53 wild-type patients. This study suggests that consideration of TP53 allele status is critical for diagnostic and prognostic assessment of MDS.

Diffuse large B-cell lymphoma (DLBCL) is one of the most common non-Hodgkin's lymphomas with significant clinical prognostic differences and molecular heterogeneity. In recent years, a number of different molecular typing suggestions for DLBCL have been put forward based on the data of gene expression profiling and exome studies, which increasingly show the guiding significance of accurate typing based on gene indicators for the diagnosis and treatment of DLBCL. However, at present, the typing standard that is unanimously recognized by experts and easy to implement has not been reached. In order to establish a more systematic molecular typing system that is easy to implement clinically, the British Hematology and Oncology Research Network reported a population-based patient cohort study. The project conducted targeted sequencing studies of 293 hematological tumor-related genes on tumor DNA samples from 928 DLBCL patients with complete follow-up data from 14 hospitals covering a population of 4 million UK. The study employed Bernoulli mixed models for molecular typing clustering and analyzed their correlation with clinical features and prognosis. A total of five DLBCL molecular typing were identified, namely MYD88, BCL2, SOCS1/SGK1, TET2/SGK1 and NOTCH2, and an unclassified group. Among them, subtypes characterized by BCL2, NOTCH2, and MYD88 gene abnormalities suggest good, moderate, and poor prognosis, respectively, consistent with other research reports. The biological characteristics of the SOCS1/SGK1 subtype overlap with those of primary mediastinal B-cell lymphoma and have a favorable prognosis. NOTCH1 mutations, although not identified as molecular typing indicators, are associated with poor prognosis. The effects of TP53 mutations vary by molecular subtype, with no effect on prognosis in the NOTCH2 subtype and poor prognosis in the MYD88 subtype. This study further confirms the clinical prognostic significance of genetic testing and molecular typing of patients with DLBCL, as well as the importance of reaching a classification consensus.

Tyrosine kinase inhibitors (TKIs) targeting the BCR-ABL1 oncoprotein have revolutionized the treatment of leukemias carrying this fusion gene. However, after the application of TKIs, the phenomenon of BCR-ABL1 mutation resistance, mainly kinase domain mutation, was quickly discovered. BCR-ABL1 mutation resistance mostly occurs after TKIs are treated, and it can also be manifested as primary resistance, and the dynamic evolution of multiple resistant clones can occur during the treatment process (including the replacement of different TKIs). Punatinib is currently the only third-generation TKIs approved for marketing that are effective against almost all single BCR-ABL1 kinase domain mutations, including the T315I mutation that is extensively resistant to all first-and second-generation TKIs. However, with the increase of application, the phenomenon of punatinib resistance also occurs from time to time. Patients with punatinib resistance have used multiple TKIs sequentially, with a long course of disease and complex drug-resistant mutations (including compound mutations, polyclonal mutations, dynamic evolution of mutant clones, etc.), which bring challenges to detection and research. In the past, the first generation Sanger gene sequencing method was mostly used to detect BCR-ABL1 kinase domain mutations, which had limited detection sensitivity, could not be quantitatively analyzed, and it was difficult to distinguish between polyclonal mutations and composite mutations. The BCR-ABL1 mutation detection based on second-generation gene sequencing (NGS) technology reported in this study can not only directly identify polyclonal mutations and composite mutations, but also have the advantages of high detection sensitivity, quantitative analysis, and clonal evolution analysis. The results of this study show that the drug-resistant mutations of punatinib are mostly compound mutations involving the T315 codon of ABL1 gene, which occur sequentially and occur at multiple sites; Patients who are resistant to prenatinib all experience an active process of dynamic evolution and accumulation of mutations. This further emphasizes the importance of using NGS technology to monitor the dynamic evolution of drug resistance mutations, composite mutations and mutant clones when applying TKIs such as punatinib. The clonal evolution and composite mutation of drug resistance mutations have become a serious problem in the treatment of sequential TKIs. It is necessary to deeply understand the drug resistance mutation rules of various TKIs to optimize the treatment plan. The improvement of detection technology provides a powerful tool for medical research and clinical application.