Molecular Basis and Clinical Application of Growth-Factor-Independent In Vitro Myeloid Colony Formation in Chronic Myelomonocytic Leukemia

Abstract

We have originally reported that colony-forming units granulocyte/macrophage (CFU-GM) formation is an in vitro feature of chronic myelomonocytic leukemia (CMML) and a strong predictor for short survival. Elucidation of the molecular basis underlying this in vitro phenomenon could be helpful to define molecular features that predict inferior outcome in patients. We studied the correlation between the mutational landscape and spontaneous colony formation in 164 samples from 125 CMML patients. As compared to wildtype samples, spontaneous in vitro CFU-GM formation was significantly increased in samples containing mutations in NRAS, CBL and EZH2 that were confirmed as independent stimulatory factors by multiple regression analysis. Inducible expression of mutated RAS but not JAK2 was able to induce growth factor independence of Ba/F3 cells. Whereas high colony CFU-GM growth was a strong unfavorable parameter for survival (p < 0.00001) and time to transformation (p = 0.01390), no single mutated gene had the power to significantly predict for both outcome parameters. A composite molecular parameter including NRAS/CBL/EZH2, however, was predictive for inferior survival (p = 0.00059) as well as for increased risk of transformation (p = 0.01429). In conclusion, we show that the composite molecular profile NRAS/CBL/EZH2 derived from its impact on spontaneous in vitro myeloid colony formation improves the predictive power over single molecular parameters in patients with CMML.

Molecular Pathogenesis of Chronic Myelomonocytic Leukemia and Potential Molecular Targets for Treatment Approaches

“…In this analysis NRAS mutations alone did not reach statistical significance as an independent factor impacting AML-free survival, however, the combined oncogenic RAS-pathway category including NRAS, KRAS and CBL was statistically significant in a model that only included genetic factors ( 74 ). Considering the fact, that spontaneous colony formation in CMML functionally covers the most frequent RASopathy gene mutations ( 38 ) these data are in line with findings in a small study which have been previously reported. In this study patients with CMML growth-factor-independent colony formation after transformation was significantly increased as compared to CFU-GM growth before transformation ( 37 ).…”

“…Moreover, the group in Vienna could show in a small retrospective study that CMML patients with high spontaneous CFU-GM growth (>100/10 5 PB mononuclear cells) have an inferior prognosis as compared to patients with low myeloid colony formation suggesting a clinical significance of the original observation (37). These results have been recently extended in a much larger CMML patient cohort indicating that spontaneous myeloid colony-formation was, compared to other single established prognostic factors, the strongest predictor regarding overall survival (OS) (38). This may indicate that in vitro cultures using unmanipulated mononuclear cells (MNC) may be a more global test that covers different aspects of malignancy better than any of the single parameters that are currently used to characterize the behavior of a tumor.…”

Significance of cardiovascular comorbidity in patients with chronic myelomonocytic leukemia

“…We and others have shown that circulating blasts are an established adverse prognostic factor in patients with CMML [ 6 , 23 ]. We have recently demonstrated that a composite molecular parameter including NRAS/CBL/EZH2 , derived from its impact on spontaneous in vitro myeloid colony formation, was predictive for inferior survival as well as for an increased risk of transformation [ 26 ]. In the multivariable analysis reported by the Mayo group for their prognostic model, PB blasts but not BM blasts had a significant impact on leukemia-free survival [ 6 ].…”

Matrix-GLA-Protein and Vascular Calcification: Can Diet Influence the Consequences of Matrix GLA Protein Inactivation? A Review

“…Hepatic vitamin K-dependent proteins have been implicated as serine proteases in the coagulation system [88][89][90][91] .…”