Clinical features and survival outcomes in patients with chronic myelomonocytic leukemia arising in the context of germline predisposition syndromes
Authors: Emma C. St. Martin, Alejandro Ferrer, Kitsada Wudhikarn, Abhishek Mangaonkar, William Hogan, Ayalew Tefferi, Naseema Gangat, Terra Lasho, Jessica K. Altman, Lucy A. Godley, Mrinal M. Patnaik
Journal: American Journal of Hematology (2021)

To the Editor: Chronic myelomonocytic leukemia (CMML) is an aggressive hematologic malignancy characterized by sustained peripheral blood monocytosis with overlapping features of myelodysplastic syndromes (MDS) and myeloproliferative neoplasms (MPN).1 Chronic myelomonocytic leukemia is an aging disease, characterized by the sequential accumulation of leukemia-associated driver mutations occurring on a background of clonal hematopoiesis (CH).2 The median age at CMML diagnosis is 73 years, with a male preponderance.3 Clonal hematopoiesis involving TET2 or TET2/SRSF2 mutations skews hematopoiesis with a monocyte bias,4 and the subsequent accumulation of epigenetic (ASXL1) and signaling mutations (NRAS, CBL, PTPN11, KRAS and JAK2) contributes to CMML development and phenotypic heterogeneity.5 Germline predisposition syndromes are cancer risk disorders that occur due to constituent aberrations in important homeostatic and tumor suppressor genes. These syndromes can broadly be classified into a) general cancer predisposition syndromes (e.g., TP53, ATM, CHEK2, and CDK2NA) and b) cancer risk syndromes specifically increasing risk for hematologic malignancies (e.g., RUNX1, ANKRD26, ETV6, GATA2, CEBPA, DDX41, SAMD9, and SAMD9L).6 With better sequencing techniques and deeper understanding of cancer predisposition states, greater efforts are being placed to identify germline variants in individuals presenting with hematologic malignancies. This is especially applicable when patients present with antecedent thrombocytopenia (e.g., RUNX1, ANKRD26, and ETV6), syndromic features (e.g., GATA2, SAMD9, and SAMD9L), multiple cancers (e.g., TP53, CDK2NA, and CHEK2), or > 1 first-degree relative with hematologic malignancies.6 Although germline mutations involving PTPN11, CBL and NF1 have been documented in juvenile myelomonocytic leukemia, systematic assessment for germline risk alleles in those diagnosed with CMML have not been carried out. We performed this study to describe the prevalence of germline predisposition syndromes associated with CMML from two large hematologic referral centers in the United States.7 After obtaining IRB approval at both institutions (Mayo Clinic, Rochester, Minnesota and The University of Chicago, Chicago, Illinois) germline predisposition databases were queried for patients with a diagnosis of CMML. The Mayo Clinic germline predisposition database had 154 patients, while The University of Chicago database had 544 patients. The CMML diagnosis was based on 2016 World Health Organization diagnostic criteria and cytogenetic studies and somatic next generation sequencing (NGS) results obtained on bone marrow samples were abstracted.8 Germline testing in general was carried out using targeted capture assays, targeted exome-based slice panels, trio whole exome sequencing, and array comparative genome hybridization assays (aCGH - for detection of copy number alterations) on germline DNA as previously described, with choice of testing being driven by the presenting clinical scenario.9 Genes most commonly interrogated at both institutions included, ANKRD26; ATG2B; ATM; BLM; BTK; CASP10; CBL; CCND1; CDK2NA; CEBPA; CHEK2; C-MPL; CSF3R; DDX41; DIS3; DNAJC21; EFL1; ELANE; ETV6; FAN1; FANCA; FANCB; FANCC; FANCD1; BRCA2; FANCD2; FANCE; FANCF; FANCG; FANCI; FANCJ; BRIP1; FANCL; FANCM; FANCN; PALB2; FANCO; RAD51C; FANCP; SLX4; FANCQ; ERCC4; FANCR; RAD51; FANCS; FANCE; FANCT; UBE2T; FAS; FASLG; G6PC3; GATA1; GATA2; GBA; GSK1; GSKIP; JAK2; KDM1A; LSD1; KLHDC8B; LIG4; MBD4; MECOM; MLH1; MSH2; MSH6; NBS1; NCOA1; NF1; PAX5; PMS2; POT1; PTPN11; RBM8A; RBM8A; RECQL2; RECQL4; RPL11; RPL35A; RPL5; RPS17; RPS19; RPS24; RTEL1; RUNX1; SAMD9; SAMD9L; SBDS; SH2B3; SRP72; TERC; TERT; TET2; TP53; TSR2; WAS; WASP; HAX1; IKZF1 (100% coding region coverage with a read depth of x100-x120). In this study, germline DNA was obtained from skin biopsy-derived fibroblast cultures (n = 6) and in two patients, germline aberrations in RUNX1 were detected in blood samples and confirmed to be germline by family segregation assays. Kaplan-Meir survival analysis was carried out comparing overall survival (OS) and acute myeloid leukemia (AML)-free survival of patients with CMML arising in the context of germline predisposition states with the Mayo Clinic CMML (n = 578) cohort, with the log rank test being used to compute p values. All statistical calculations were carried out using JMP statistical software, version 16.0, SAS Institute Inc. Cary, NC, 1989–2019. We identified eight patients with CMML (five of 578 Mayo Clinic CMML patients-0.8%) that had an underlying germline predisposition syndrome (3% of Mayo Clinic and 0.5% of The University of Chicago germline predisposition database cohorts): seven males (87%), median age at CMML diagnosis 65 years (range, 54–76 years) (Table 1). Four (50%) patients presented with antecedent thrombocytopenia and were subsequently diagnosed to have germline abnormalities involving RUNX1 (n = 3; 37.5%) and ANKRD26 (n = 1; 12.5%). Two of three (67%) patients with RUNX1-FPD (familial platelet disorder) had intragenic gene deletions that were missed on somatic mutation analysis carried out by clinical amplicon-based NGS assays and were only detected by aCGH. Four (50%) patients developed CMML in the context of general cancer predisposition syndromes, including three (37.5%) with germline CHEK2 mutations and one with a germline CDKN2A (12.5%) mutation (Table 1). In seven of eight patients, presentation with CMML led to the underlying diagnosis of a germline predisposition syndrome, either due to antecedent thrombocytopenia (n = 4), or due to concomitant visceral malignancies and family histories (n = 3). There was one patient with a CDKN2A mutation that was detected a decade prior to the diagnosis of CMML due to a family history of pancreatic cancer and recurrent melanomas. Interestingly, there were no CMML cases seen in our cohorts of 16 families with GATA2 haploinsufficiency (characterized by monocytopenia), 55 families with germline DDX41 mutations (late age of onset of myeloid neoplasms) and four families with germline ETV6 mutations (antecedent thrombocytopenia).6 CMML-1 dCMML ASXL1: c.2290delC; p.(L764Yfs*8) (30%) KRAS: c.35G > A; p.(G12D) (35%) CMML-0 dCMML SRSF2: c.284C > A; p.(P95H) (48%) TET2: c.3955G > T; p.(E1319*) (42%) TET2 VUS: c.3594 + 5G > A; p.? (45%) CMML-1 pCMML JAK2: c.1849G > T; p.(V617F) (91%) SRSF2: c.284C > A; p.(P95H) (51%) ASXL1: c.1934dupG; p.(G646Wfs*12) (29%) Alive with oral GVHD Tipifarnib (K0-TIP-004 clinical trial) Hydroxyurea Ruxolitinib HCT after fludarabine and melphalan CMML-0 dCMML SRSF2: c.284C > A; p.(P95H) (44%) TET2: c.2899C > T; p.(Q967*) (37%) TET2: c.4661_4664del; p.(T1554Sfs*16) (39%) Melanoma x2 prostatic adenocarcinoma CMML-0 dCMML TET2: c.4045-2A > C; p.? (43%) TET2: c.3656A > G; p.(H1219R) (40%) SRSF2: c.161C > T; p.(S54F) (15%) PHF6: c.866C > A; p.(T289N) (9%) KRAS: c.35G > A; p.(G12D) (8%) NRAS: c.35G > A; p.(G12D) (7%) PHF6: c.719A > G; p.(Y240C) (3%) CMML-1 pCMML CMML-2 pCMML SRSF2: c.284_307del; p.(P95_R102del) (45%) TET2: c.2504C > G; p.(S835*) (46%) CBL: c.1150 T > A; p.(C384S) (90%) MPL: c.1771 T > G; p.(Y591D) (6%) TET2: c.3986 T > G; p.(L1329R) (48%) CMML-1 dCMML TET2: c.1882G > T; p.(E628*) (37%) TET2: c.4249del; p.(V1417Ffs*31) (41%) TP53: c.596G > A; p.(G199E) (42%) ZRSR2: c.52_53insCA; p.(Y18Sfs*6) (84%) Somatic mutation profiling data were available in seven of eight patients, with the most frequent mutations being TET2 (71%), SRSF2 (71%) and ASXL1 (28%). All five patients with TET2 mutations had two TET2 mutations, commensurate with prior data demonstrating the high frequency of monocytosis in patients with TET2/SRSF2 or biallelic TET2 mutations.10 Patients with germline aberrations in checkpoint regulation (CDKN2A) and DNA damage response (CHEK2) had four or more somatic mutations, in line with inherent mutagenicity of these conditions.11 The karyotype was normal in five (71%) patients, while one patient each had -Y and a balanced translocation involving chromosomes 8 and 22. Although TP53 and MPL mutations are uncommon in CMML (<1%), they were seen in two patients with germline CHEK2 mutations. Monocyte repartitioning flow cytometry was carried out in three Mayo Clinic patients with RUNX1-FPD, with all three demonstrating M01 fractions >94%; consistent with CMML.12 One patient with the balanced translocation between eight and 22 transformed to AML, whereas one patient successfully underwent reduced intensity allogenic stem cell transplantation (HCT) for proliferative CMML refractory to conventional therapies. At last follow-up two (28%) patients were deceased, one from AML-transformation and one from an unrelated cause. There was no difference in median overall (not reached vs 948 days; p = 0.28) and AML-free survival (not reached vs 687 days; p = 0.23) between CMML patients with an underlying germline predisposition state versus those without (Figure S1A,B). We comprehensively describe the clinical features and outcomes in eight patients with CMML that occurred in the context of germline predisposition states. Although 50% of cases occurred in patients with germline predisposition to hematological neoplasms (RUNX1 and ANKRD26), four (50%) cases occurred in the context of general cancer predisposition (CHEK2 and CDK2NA) syndromes, emphasizing the need for comprehensive genomic assessments extending beyond genes involved in hematologic neoplasms. Both RUNX1 probands would not have been diagnosed if copy number alterations were not assessed for by aCGH, highlighting the importance of these alterations and limitations of amplicon based NGS assays. Three patients with CHEK2 mutations, of Polish heritage, where the prevalence of heterozygous CHEK2 mutations is approximately 5%, developed CMML, with two of three (67%) having the p.I157T [with current numbering I200T] CHEK2 hotspot mutation.13 CHEK2 is a critical component of the ATM-CHEK2-TP53 pathway and although CHEK2 perturbations were associated with CMML, there were no cases of CMML in our germline ATM (n = 8) and TP53 (n = 25) mutation cohorts. To the best of our knowledge, we report the first case of CMML in a germline CDKN2A mutated patient with an expectedly high tumor mutational burden, including somatic mutations in PHF6, KRAS, and NRAS. Regardless of the cancer predisposition genes, TET2 (71%) and SRSF2 (71%) remained the most common somatic mutations associated with a CMML phenotype.1 Note, TET2 encodes a critical dioxygenase required for the conversion of 5-methylcytosine to 5-hydroxymethylcytosine and related metabolites, and SRSF2 is a serine arginine rich splicing factor playing a role in mRNA splicing.14 Our data demonstrate that assessment for germline predisposition states might be an essential part of work up of hematologic neoplasms. Detection of mutations and copy number alterations in a growing list of candidate genes has the potential to impact treatment options, donor selection for HCT, screening of additional family members and post-HCT survivorship and cancer surveillance patterns. We would like to acknowledge all patients and families who contributed to the study and the Henry J Predolin Leukemia grant at the Mayo Clinic. All authors declare no relevant conflicts of interest and disclosures. All NGS and germline sequencing data are outlined in Table 1. Raw data can be obtained by contacting the corresponding author. Figure S1- Overall and AML-free survival comparisons between CMML patients with germline predisposition syndromes and those without germline predisposition syndromes. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.