Quantitative trait loci associated with susceptibility to therapy-related acute murine promyelocytic leukemia in hCG-PML/RARA transgenic mice

Abstract

Therapy-related acute myelogenous leukemia (t-AML) is an important late adverse effect of alkylator chemotherapy. Susceptibility to t-AML has a genetic component, yet specific genetic variants that influence susceptibility are poorly understood. We analyzed an F(2) intercross (n = 282 mice) between mouse strains resistant or susceptible to t-AML induced by the alkylator ethyl-N-nitrosourea (ENU) to identify genes that regulate t-AML susceptibility. Each mouse carried the hCG-PML/RARA transgene, a well-characterized initiator of myeloid leukemia. In the absence of ENU treatment, transgenic F(2) mice developed leukemia with higher incidence (79.4% vs 12.5%) and at earlier time points (108 days vs 234 days) than mice in the resistant background. ENU treatment of F(2) mice further increased incidence (90.4%) and shortened median survival (171 vs 254 days). We genotyped F(2) mice at 384 informative single nucleotide polymorphisms across the genome and performed quantitative trait locus (QTL) analysis. Thirteen QTLs significantly associated with leukemia-free survival, spleen weight, or white blood cell count were identified on 8 chromosomes. These results suggest that susceptibility to ENU-induced leukemia in mice is a complex trait governed by genes at multiple loci. Improved understanding of genetic risk factors should lead to tailored treatment regimens that reduce risk for patients predisposed to t-AML.

Figure 1

Kaplan-Meier survival curves of treated and untreated B6C3F1 PR⁺ and F₂ mice. Untreated mice in the resistant parental background (B6C3F1 PR⁺) develop leukemia after a long latency and with low penetrance (▴). ENU treatment of PR⁺ mice in the B6C3F1 background increases leukemia penetrance and decreases latency (▾). Intercrossing SWR/J alleles has an effect equipotent to ENU treatment in B6C3F1 mice (▾ vs ■, P = .34; ▴ vs ▾ or ▴ vs ■, P < .001). Treatment with ENU further increases the rate and the incidence of leukemogenesis in F₂ mice (■ vs ●, P < .001).

Figure 2

Morphology of tumors detected in ENU-treated PR⁺ F₂ mice. (A-C) Myeloproliferative disease, (D-F) myeloid leukemia, (G-I) lymphoma, (A,D,G) peripheral blood, (B,E,H) bone marrow, and (C,F,I) spleen. (A) Normal lymphocyte and neutrophil. (B) Mild myeloid hyperplasia (secondary to the PR transgene) and normal maturation of all lineages. (C) Normal splenic architecture. (D) Clusters of immature myeloid cells in the peripheral blood. (E) Left shift and expansion of the myeloid lineage in the bone marrow. (Inset) Myeloperoxidase positive blasts. (F) Effacement of splenic architecture by leukemic blasts. (G) Immature lymphoid cells in peripheral blood. (H) Infiltration of bone marrow by malignant lymphocytes. (I) Effacement of splenic architecture by lymphoma cells. Original magnification: ×50 (A), ×40 (C,F,I); ×100 (all others). Images were obtained using a BX40 microscope (Olympus, Center Valley, PA) with UPlanFl 40×/0.75, Plan 50×/0.90 oil iris, and UplsnFl 100×/1.30 oil ∞/0.17 objective lenses. DP70 camera and DP controller software version 2.2.1.227 (Olympus) were used for image acquisition. Photoshop 7.0 (Adobe Systems, San Jose, CA) was used for image preparation.

Figure 3

Quantitative trait locus (QTL) mapping for survival and WBC count in leukemic PR⁺ F₂ mice. QTL and survival curves were generated from analysis of F₂ mice. (A) Survival QTL (Mleu1.1) in the treated model on chromosome 1 at 42.8 cM that is significant at the genomewide level. (B) Survival of ENU-treated F₂ mice is significantly (P < .001) affected by genotype at Mleu1.1. Arrows indicate ENU treatment at 9 and 10 weeks of age. (C) WBC count QTL (Wbc19.1) on chromosome 19 at 47.0 cM that is significant in the covariate model at the chromosome-wide level. (D) At Wbc19.1, SWR/J homozygotes have a higher mean WBC count than B6C3F1 homozygotes (79 ± 128 vs 48 ± 96, respectively, P < .01) independent of treatment status. Data are plotted as median (horizontal bar), 25th to 75th percentile (box) and range (whiskers).