In addition to the somatic changes related to diagnosis and prognosis in leukemia cells, many germline mutations have been discovered in hematopoietic malignancies in recent years, namely hereditary predisposition (1, 2). The ‘Myeloid Neoplasms with Germline Predisposition’ category has been included in the revised fourth edition of the World Health Organization (WHO) classification of tumors of the hematopoietic and lymphoid tissues (3), which further emphasizes the importance of constitutional mutation in the occurrence and development of hematological neoplasms. Fusion genes are hybrid genes formed by the fusion of two previously separated genes, which are resulting from chromosomal rearrangement, including translocation, inversion, deletion, or tandem duplication (4). Fusion genes are molecular biological characteristics of many leukemia types and have been successfully used as diagnostic markers and therapeutic targets (5). However, its relationship with hereditary predisposition is still unclear.

Here, we report a case of familial acute leukemia with fusion genes and germline RecQ Like Helicase (RECQL) mutation. The proband was a 42-year-old female patient with acute promyelocytic leukemia (APL) and PML-RARα fusion gene. She has two daughters, and the younger one developed B-cell precursor acute lymphoblastic leukemia (BCP-ALL) with ETV6-RUNX1 fusion gene at age 6. As of the date of submission, the eldest daughter is still in good health. Comprehensive genetic testing showed that a single nucleotide variant (SNV) in RECQL, rs146924988, was potentially involved in this familial case with acute leukemia.

With the development of genomic sequencing methods, many germline mutations have been found in cancer patients and their immediate relatives. Germline mutations associated with the hereditary predisposition to cancers were highlighted for their clinical relevance and diagnostic values, which were also known as hereditary predisposition syndromes (HPS) (9). For hematological malignancies, hereditary predisposition was first identified in chronic lymphocytic leukemia (CLL) and acute myeloid leukemia (AML), which has led to the identification of several germline mutations, such as RUNX1, CEBPA, GATA2, ANKRD26, DDX41 and ETV6 mutations. Furthermore, genetic susceptibility was indicated in ALL and multiple myeloma, with constitutional mutations in genes such as IKZF1, SH2B3, PAX5 (ALL) and KDM1A/LSD1 (multiple myeloma). Monogenic genomic alterations have also been found to be involved in inborn errors of the immune system and confer a predisposition to hematological malignancies such as ataxia telangiectasia, Nijmegen breakage syndrome, Bloom’s syndrome, xeroderma pigmentosum, constitutional mismatch repair deficiency, Fanconi anemia, and telomere syndromes (2, 10–12). Interestingly, previously identified inherited cancer susceptibility syndromes also increase the possibility to develop hematological malignancies. For example, patients with Diamond-Blackfan anemia, severe congenital neutropenia, or familial thrombocytopenia have a significantly increased incidence of myeloid neoplasms (10). Up to 62% of children with lymphoproliferative diseases have primary immune deficiency disease (PID) (13). Compared with the general population, people with PID have a 1.5 times higher risk of developing cancer (14). At present, HPS-related genes or pathways involve transcription factors, RNA function, DNA methylation, ribosome assembly, histone modification, DNA unwinding and DNA repair (9). Other germline mutations in key transcription and translation processes and signal transduction pathways, such as the RAS pathway, cell proliferation, apoptosis, and tumor suppressor genes (e.g., TP53) were also considered as possible underlying predispositions.

In this case, the proband and her second daughter both suffered from acute leukemia at different times in their lives. Considering the incidence of acute leukemia, we speculated that there might be HPS in this case. Through WES sequencing and bioinformatics analysis, we finally focused on rs146924988, an SNV in RECQL, and the mutation frequencies in the proband and her second daughter were 45.1% and 42.9%, respectively. Further Sanger sequencing showed no such mutation of RECQL in the healthy child of the proband, which indicated that rs146924988 might play a role in this case of familial leukemia. The proband of this case was an APL patient, carrying a typical PML-RARα fusion gene. The second daughter of the proband was a BCP-ALL patient, carrying the classical ETV6-RUNX1 fusion gene. Considering the driving role of these two different fusion genes in the occurrences and development of corresponding leukemias (15, 16), it can be inferred that this RECQL gene variant (rs146924988) is potentially associated with the generation of acute leukemia. We evaluated the alteration frequency and expression levels of RECQL in acute leukemia from publicly available datasets using the cBioPortal database (17), including AML datasets from The Cancer Genome Atlas (TCGA) (18) and B-ALL datasets generated by the Therapeutically Applicable Research to Generate Effective Treatments (TARGET, https://ocg.cancer.gov/programs/target) initiative, phs000464. Samples with complete data of gene mutations, copy-number alterations (CNA) and mRNA expressions were retained after screening. The CNA frequency of RECQL was more than 5% both in ALL and AML patients, and the main variants types are deep deletion and shallow deletion that related to low gene expression ( Figures 2A, B ). Interestingly, the expression of RECQL in AML-M3 patients mostly having PML-RARα fusion gene, was significantly lower than that in other types of AML patients ( Figure 2C ). These findings highlighted the potential role of RECQL in leukemia.

RECQL gene, as a member of the RecQ helicase family located on chromosome 12p12, encodes a DNA helicase with 649 residues. RECQL protein is a highly conserved protein and plays an important role in DNA repair, replication, recombination and transcription (19). The SNV (rs146924988) detected in this case is an initiator codon variant of RECQL, which may lead to start-lost loss-of-function of RECQL (20, 21) ( Figures 3A, B ). Prior to this report, the RECQL germline mutation has been found to be associated with hereditary breast cancer and hereditary ovarian cancer syndrome (22, 23), but its role in hematological malignancies has not been reported. RECQL protein is an ATPase and helicase that can bind and unlock the structural intermediate of DNA replication and repair. RECQL unlocks double-stranded DNA and catalyzes the migration of ATP-dependent branches on Holliday junctions and removable D-loop substrates. In addition to cleavage of DNA, RECQL promotes annealing of complementary single-stranded DNA in an ATP-independent manner (24). Moreover, RECQL can also interact with proteins that are involved in DNA replication, repair and mismatch repair, such as FEN1, RPA, PARP1, and MLH1 (25–27). Deletion of RECQL resulted in increased DNA damage accumulation, which indicated that RECQL was involved in the solution of replication fork stagnation. RECQL-deficient mice and cells exhibited frequent spontaneous chromosomal breakage and chromatid translocation (28). In a recent in-vitro experiment based on MDA-MB-231 cells, RECQL was considered to be involved in regulating cell stress response (29). It has been found to mediate the recruitment of replication protein A (RPA) in DNA damage sites and to promote the activation of the ATR-Chk1 pathway in response to the DNA damage caused by gemcitabine (29). ATR-Chk1 is a classic pathway of DNA damage response (DDR) induced by DNA double-strand breaks (DSBs), involving cell cycle checkpoint regulation and DNA repair (30). It can promote ubiquitination and proteasome degradation by phosphorylating CDC25 phosphatases (CDC25A, B, C), and induce cell cycle G1-S transition and G2-M transition (31–33). ATR-Chk1 dysfunction is associated with a large number of DNA damage and chromosomal rearrangements in Ataxia-telangiectasia (A-T) patients, especially in lymphocytes, which often leads to the production of fusion genes (34, 35).

Based on these previous findings, we hypothesize that the constitutional mutation in RECQL gene (rs146924988) may inhibit the translation of RECQL mRNA and lead to a relative lack of RECQL protein, which may further impair the ATR-Chk1 pathway and resulting in functional disorders in DNA replication and DSBs repair. It is conceivable that these functional disorders could ultimately mediate the occurrence of chromosome rearrangement and fusion genes, as driving factors for leukemia ( Figure 3C ). Admittedly, we have not obtained direct evidence that RECQL gene mutation (rs146924988) will lead to decreased RECQL expression level because this study is a case report in nature, and the biological process of such mutation potentially affecting DSBs repair and mediating the production of fusion genes needs to be verified in in-vitro experiments and larger clinical cohorts in the future. Moreover, there were 7 other shared germline mutations in this case of familial leukemia, which were related to genes AK2, C7, CCDC40, NCF4, KCNQ4, GJB2 and USH2A. These genes were reported to be involved in ADP biosynthesis, complement activation, oxidation-reduction and other biological processes, and their functional annotations were shown in Table S1 . By checking the literature and public data repository, there is no direct evidence that these mutations are associated with genomic instability or genetic predisposition to cancer, but their contributions to the onset of leukemia cannot be completely excluded and needs further evaluation. The loss of RECQL function may increase the probability of the somatic events, such as the occurrence of chromosome rearrangements, in positions given by chance and modulated by other factors, resulting in different genetic fusions genes. This partly explained how germline genetic variants in RECQL lead to different types of fusions gene in these 2 cases.

Early recognition of the hereditary predisposition of hematological malignancies is critical for timely diagnosis and individualized treatment. In this study, a potential HPS-related gene mutation was identified by WES sequencing, which highlights the clinical application of WES, especially in patients with early-onset leukemia or familial leukemia. In addition to diagnostic value, screening of the HPS-related gene mutations may provide prognosis evaluation reference for an optimal intervention approach. For example, previous studies have shown that RECQL gene defects not only cause genomic instability and chromatin recombination, but also make cells more sensitive to toxic genomic stress and cytotoxic therapy (29). Particularly, polymorphic variants in RECQL was found to be related to the overall survival rate of pancreatic cancer patients treated with gemcitabine (36). RECQL helicase has also been found to protect multiple myeloma cells from melphalan and bortezomib cytotoxicity (37). Therefore, the significance of RECQL mutation in the treatment of acute leukemia needs to be evaluated in future studies.

In this study, we performed WES sequencing in a familial leukemia case, and for the first time to our knowledge, found a germline RECQL mutation potentially involved in hereditary predisposition to acute leukemia. The hypothetical biological process in which RECQL gene mutation (rs146924988) affect DSBs repair and mediate the generation of fusion genes provides a new understanding of the pathogenesis of leukemia, and highlights the necessity for next-generation sequencing-based screening of genes involved in this process in potential HPS patients.

The datasets presented in this article are not readily available because of ethical/privacy restrictions. Requests to access the datasets should be directed to the corresponding author.

This study was approved by the Medical Ethics Committee of the Department of Hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology. Written informed consent to participate in this study was provided by the participants’ legal guardian/next of kin.

All authors designed the study, interpreted the findings and revised the manuscript. WY, ZS, KS, QY, and QL carried out data management and statistical analysis and drafted the manuscript. YC and JW helped with English language editing and data management. YY performed project administration. All authors contributed to the article and approved the submitted version.