• c-Myc
• diffuse large B-cell lymphoma
• DLBCL
• point mutations

Diffuse large B-cell lymphoma (DLBCL) is the most common lymphoid malignancy in adults. This type of tumor is characterized by high genetic and phenotypic heterogeneity, highlighting the need to dissect this entity at the molecular level to develop more effective and targeted therapies. Recent studies have defined a high-risk DLBCL subgroup, the so-called Molecular High-Grade (MHG), which is associated with poor clinical outcome and a significantly higher frequency of c-Myc point mutations. c-Myc is a transcriptional factor involved in many key cellular processes such as apoptosis, cell cycle and proliferation. c-Myc point mutations have been found in a variety of solid and non-solid cancers, but only few of them have been functionally tested so far. These mutations could have a previously underestimated impact on cancer evolution, as they could confer increased pro-proliferative and transforming capacity. Ultimately, investigating these mutations could be important to better understand the increased aggressiveness of the MHG high-risk DLBCL subgroup.
This master’s thesis focuses on the functional characterization of specific c-Myc point mutations identified in MHG-DLBCL patients. In particular, I studied the D2A, V6E, and P75S/P60S mutations. The latter is present in both the c-Myc isoforms expressed in human and it is located in the c-Myc Box I domain which regulates c-Myc degradation, while the other two are specific for the longer c-Myc isoform. The aim of this study is testing whether these mutations lead to an increase in c-Myc half-life and transforming capacity in vitro.
The model I adopted was a mouse embryonic fibroblast (MEF) cell line where the endogenous c-Myc is fused with the estrogen receptor (MycERT2) and therefore c-Myc is active only upon 4-hydroxytamoxifen (4-OHT) addition to the culture media. This model allowed me to specifically express and study the desired exogenous c-Myc mutant in absence of 4-OHT.
As expected, the P60S and P75S mutations were found to promote a significant increase in c-Myc protein half-life and stability and showed multiple evidence of cellular transforming ability, similarly to the known c-Myc box I point mutations. Conversely, the long c-Myc isoform-specific point mutations showed none to little cellular and biochemical effects.
These results shed light on the role of c-Myc point mutations in B-cell malignancies. and further studies on others experimental model could lead to the identification of new prognostic markers, improving diagnosis and establishing the basis for mutation specific targeted-therapies in the era of precision medicine.