MYC Gene Mutations as Causative Pathways for Development and Treatment of Hematological Malignancies

Abstract Background: Genome instability is a hallmark of cancer. Mutations in DNA repair pathway genes are frequent in a number of solid tumors. Defects in DNA repair or damage response can weaken response to conventional chemotherapy and are frequently regarded as poor prognostic markers. However, a high tumor mutation burden (TMB, number of somatic mutations per mega base) was recently found to correlate with better response to immune checkpoint inhibitors e.g. in colon cancer. Patients with defects in the DNA mismatch repair (MMR) pathway in solid tumors are among the cases with the highest TMB. Hematological malignancies are generally expected on the lower end of the TMB spectrum. We used whole genome sequencing (WGS) for 3256 patients with hematological malignancies (lymphatic and myeloid) to determine factors of genetic instability across all entities. Aim: To determine the number of known mutations in genes from the DNA repair pathway To estimate TMB using WGS and identify cases with high TMB in hematologic malignancies Methods: We investigated a cohort of 3256 patients with hematological malignancies, who were analyzed according to WHO diagnostic gold standards for routine purposes (incl. 584 acute myeloid leukemia [AML] and 635 myelodysplastic syndromes [MDS] samples). We performed amplification-free library preparation and sequencing on HiseqX and NovaSeq 6000 with a median coverage of 106x. Mapping and variant calling was performed with standard pipelines via BaseSpace (all Illumina, San Diego, CA). A pool of gender-matched genomic DNA (Promega, Madison, WI) was used for a tumor-unmatched normal variant calling. (a) In detail we evaluated 180 genes involved in DNA repair. We filtered on (likely) pathogenic variants from ClinVar and for TP53 on protein-truncating variants and (likely, possibly) pathogenic variants from the IARC database. (b) For TBM calculation we determined protein-altering changes and then subtracted all gnomAD listed variants in order to eliminate most germline variants. Results: We found 479 of 3256 (15%) patients with at least one pathogenic variant according to current database annotations in DNA repair or damage response genes. Most pathogenic variants were found in TP53 (330/3256; 10%) and ATM (25/3256, 1%), however, this is probably the effect of the already available systematic database annotation for both genes. For routine diagnostic purposes TP53 mutation status had been analyzed for 1184 patients with Sanger sequencing (7%) or amplicon next-generation sequencing (93%). A 98% and 99% concordance of the pathogenic and non-pathogenic TP53 status was found in comparison to WGS. Mutations in genes from the DNA double-strand break repair (and/or homologous DNA pairing and strand exchange) pathway were found in 93 patients (3%). Pathogenic and potentially germline MMR gene mutations were found in only 3 patients (0.1%, 2 MLH1, 1 MSH6), which equals the expected frequency in the Western population (0.05-0.3%). Next, we calculated TMB. The average was 2.4 [range: 0.4-39.2]. Only samples above the 95th percentile were defined as "TMBhi" (TMB ≥5). TMB was lowest in chronic myeloid leukemia (CML) and essential thrombocythemia (ET) (< 2) and no ET or CML patient was found among the TMBhi. We then focused on MDS, which is our largest subcohort: 56 of 635 (9%) patients were in TMBhi. Furthermore, among MDS patients, a significantly higher TMB was found in MDS-EB-2 (average 3.3 vs. 2.3 for non EB-2, p<.001) and a significantly lower TMB in MDS with isolated del(5q) (1.7 vs. 2.6 in all other MDS, p<.001). There was no association of high TMB and annotated DNA repair gene mutations. The three MMR deficient patients had an average TMB (1.8-2.2) in the hematologic sample. However, TMB was positively correlated with age (Spearman's rank correlation, p<.001) and average TMB was lower in patients under 60 years (2.3 vs. 2.5, p=0.007, t-test). Conclusions: High TMB is rare in hematological neoplasms, and our data across all entities suggest that the acquisition of mutations over age should be a major contributing factor. While TP53 and a few other factors are well studied, using genome-wide datasets in the near future will allow to deeply understand the broad patterns or signatures of genome stability (incl. copy number or structural variants) and their prognostic or predictive value. Disclosures Baer: MLL Munich Leukemia Laboratory: Employment. Hutter:MLL Munich Leukemia Laboratory: Employment. Höllein:MLL Munich Leukemia Laboratory: Employment. Walter:MLL Munich Leukemia Laboratory: Employment. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

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Immunodeficiency Genes Are Associated with EBV Positive Malignancies and Hemophagocytic Lymphohistiocytosis That Can be Cured By Allogeneic Hematopoietic Stem Cell Transplantation

Blood ◽

10.1182/blood-2021-149918 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 4908-4908

Author(s):

Yanzhi Song ◽

Caiyan Zhang ◽

Zhihui Li ◽

Erhui Yuan ◽

Qishu Qin ◽

...

Keyword(s):

Stem Cell ◽

Hematopoietic Stem Cell Transplantation ◽

Stem Cell Transplantation ◽

Hematological Malignancies ◽

Nk Cell ◽

Gene Mutations ◽

The Other ◽

Related Gene ◽

Hematopoietic Stem ◽

Hematological Diseases

Abstract Background: Epstein-Barr virus (EBV) positive hematological malignancies and EBV-associated hemophagocytic lymphohistiocytosis (HLH) are serious illness that is related with chronic active EBV infection (CAEBV). Recently, more and more immune-related gene mutations have been identified to associate with the EBV positive hematological diseases. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative therapy for above diseases. Aim: In present clinical study, the germline gene mutations associated with EBV positive hematological malignancies and EBV positive HLH were analyzed and the outcomes of allo-HSCT in those diseases were investigated. Methods: Between January 2018 and June 2021, 7 patients with EBV positive hematological diseases (lymphoma 3, aggressive NK-cell leukemia (ANKL) 1, secondary HLH 3) who received allo-HSCT in our hospital were included. Blood samples from patients, parents and potential related donors were collected before transplantation and whole exome sequencing of the blood DNA samples was performed to detect and analyze their relevance with the clinical features before and after allo-HSCT. The median age was 23 (5-31) years old. The median disease course before transplant was 6 (3-38) months. Before allo-HSCT, 3 patients (HLH 1, lymphoma 1, ANKL 1) were in non-remission (NR), 2 patients with lymphoma were in partial remission (PR) and the other two patients with HLH were in complete remission (CR). Five patients received allo-HSCT from haploidentical donors and 2 patients from unrelated donors (HLA 10/10, 9/10 matched). Myeloablative conditioning regimen with busulfan (0.8mg/kg iv q6h, 3 days)/etoposide (200mg/m 2, 3 days)/fludarabine (30mg/m 2, 5 days) /rabbit anti-T-cell globulin was used. Cyclosporine, mycophenolate mofetil and short-term methotrexate were employed for graft-versus-host disease (GVHD) prophylaxis. Results: All patients achieved durable engraftment. Plasma EBV-DNA was positive in three patients (1200, 880, 84000 copies/ml, respectively) before transplant and has been negative in all patients after transplant. One patient had gastrointestinal bleeding caused by EBV enteritis. Two patients had CMV viremia. Three patients developed grade III acute GVHD and two had extensive chronic GVHD. All GVHD was resolved with immune-suppressants. With the median follow-up time of 21 (7-36) months, all patients are alive and free of their diseases. Seven disease-related gene mutations were detected including FANCA p.S1088F, TGFBR2 p.T315M, IRF7 p.K446R, PRF1 p.R4C, MPEG1 p.P316S, STX11 p.R49Q, and UNC13D p.G863D. The first 5 gene mutations were detected in the patients with lymphoma and ANKL, and the last 2 gene mutations were detected in the patients with HLH. FANCA is associated with bone marrow failure and the other genes are associated with primary immunodeficiency, all of these gene mutations have been reported and affect in part the function of the genes. In particular, STX11 p.R49Q and UNC13D p.G863D have also been reported in HLH patients and the NK cell activity of these patients is lower than normal levels. Apart from the defined gene mutations in primary HLH, recently, many new immunodeficiency gene mutations have been reported to associate with EBV positive malignancies and HLH which suggests immunodeficiency might play a role in the etiology of the diseases. The gene mutations we reported here might be the main gene mutations associated with disease susceptibility and infection risk. Conclusion: Our preliminary results have shown the gene mutation profile related to EBV positive hematological malignancies and HLH. With our transplant protocol, durable reconstitution of hematopoiesis and immune from healthy donors has been achieved successfully which has resulted in 100% disease-free survival and no transplant-related death. Allo-HSCT is a safe and effective curative modality for EBV positive hematological malignancies and HLH. Disclosures No relevant conflicts of interest to declare.

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