Distinct Phenotypes of Myeloid Neoplasia Associated with Truncating C-CBL Mutations

Introduction. Canonical missense (MS) c-CBL mutations affecting linker and zinc finger domains impair protein ubiquitination function and are the most encountered in myeloid neoplasia (MN). However, c-CBL truncations can be also observed with consequential impairment of ubiquitin-association and PI3K/AKT activation function. Due to their rarity, the phenotype and clinical implications of these variants have been poorly investigated. Therefore, we took advantage of a very large and annotated cohort of MN cases to analyze clinical, mutational, and cytogenetic characteristics of patients with the intent of describing the clinical phenotype associated with such genetic variant.

Methods. To that end we reviewed genotypic and clinically data of 11,553 cases from our institution and sources (7221 AML; 3448 MDS; 575 MPN/MDS; 309 MPN)123456. Pathogenicity of the variants was assessed according to in silico prediction tools and literature review allowing for classification of variants into tiers. All the analyses were performed with tier 1 and 2 hits but, to prove our point, truncating [not in-frame indel, nonsense, splice-site; (T)] and frameshift (FS) mutations not previously described (19 variants) were considered as tier 2 and thus, were included as well.

Results. We identified 322 c-CBL mutations in 296 patients. Based on our criteria, we focused on the variants of 184 patients harboring 202 tier 1-2 mutations (131 harboring one MS, 8 two MS, 2 three MS mutations; 36 harbored one T/FS, 1 two T/FS, 4 MS and T/FS, 1 with two MS and 1 harboring T/FS mutations). Biallelic hits were found in 46% in patients with MS and 36% in patients with T/FS. There were significant differences in the distribution of c-CBL T/FS and MS mutations among disease phenotypes: T/FS vs canonical MS mutations were more common in AML in general (78% vs 53%, P= 0.005) with enrichment in primary AML (pAML, 90% vs 68%, P=0. 023). Considering the high fraction of AML in the T/FS group, we decided to focus our analysis on this phenotype. AML patients with T/FS mutations tended to harbor abnormal karyotype compared to patients with MS mutations (31% vs 53%, P= 0.09), had higher percentage of complex karyotype (19% vs 4%, P= 0.016), lower percentage of co-mutations in the splicing factor SRSF2 (7% vs 27%, P= 0.025) and showed a trend to co-occur with del(17p) (8% vs 1%, P= 0.10) and TP53 mutations (10% vs 3%, P= 0.10). Although, no difference was detected in clonal hierarchy, c-CBL mutations appear to be more frequently sub-clonal (T/FS 63% vs MS 47%, P= 0.16). When we conducted survival analysis, AML patients with T/FS mutations had a lower median OS (although not significant) vs patients with MS mutations (11 vs 16 months, P= 0.161) vs patients with wild type c-CBL AML (11 vs 14.3 months, P= 0.20). However, Cox regression model showed an independent role of T/FS mutations as risk factor for poor outcome (Hazard ratio:1.642, 95%CI:1.002-2.690; P= 0.049). T/FS mutations occur often in pAML and convey a less favorable prognosis independently from the co-occurrence of high-risk features, such as complex karyotype, del(17p) and TP53 mutation possibly suggesting other mechanisms explaining the aggressive nature of T/FS.

Conclusions. Our study describes the differences associated with “non canonical” c-CBL mutations and T/FS variants whose impact could be underestimated and suggests that AML harboring different types of c-CBL mutations present profound differences in disease phenotype, co-mutational landscape, and outcome. Interestingly, the hierarchical analysis of c-CBL mutations, which showed that, roughly, in 1/3 of cases (30% T/FS, 35% MS mutations) c-CBL is the only mutation detected or the founder lesion, suggests that both types of c-CBL hits could represent primary leukemogenic drivers.