Caratterizzazione funzionale e molecolare di inibitori farmacologici ad attività anti-istone-deacetilasica, in modelli in vitro ed in vivo di Rabdomiosarcoma

Rhabdomyosarcoma is a malignant soft tissue sarcoma typical of children and adolescents. Standard treatments are a combination of surgery, radiotherapy (RT) and chemotherapy. Unfortunately, RT promotes the formation of metastases, due to the proliferation of RT-resistant cell populations. RMS derives from mesenchymal precursors and is divided into two histological subtypes: Alveolar (ARMS) and Embryonial (ERMS). ARMS expresses the fusion proteins PAX3- or PAX7-FOXO1 (therefore they are considered, fusion-positive FP). ERMS is fusion-negative (FN) but is characterized by various mutations and genomic aberrations at the level of the RAS and RTK pathways. However, the two subtypes have anomalous pathways in common. For example, aberrant epigenetic regulation appears to play a critical role in RMS progression and survival. Histone acetylation is tightly controlled by epigenetic mechanisms regulated by acetyltransferases (HATs) and histone-deacetylases (HDACs), which make the chromatin structure transcriptionally active or inactive, respectively. This means that deregulation of HDAC expression and/or activity may be involved in the development and progression of several cancers, including RMS. It has been seen that class I and IV HDACs are deregulated in the RMS. Thus, HDAC inhibitors (HDACis) are already used with good results for hematological tumors, but not for solid tumors. They could be useful, instead, in combination with classical therapies (e.g. RT). But several studies shown that using a pan-HDAC inhibitor could be responsible for the low therapeutic efficiency of this type of drug. FK228, or Romidepsin, is a potent natural selective HDACi for class I and II. MS275, or Entinostat, is a potent selective HDACi for class I and IV. Romidepsin induces cell cycle arrest and apoptosis in various solid tumors. Entinostat appears to induce cell cycle arrest in the G0/G1 phase and promotes apoptosis. It downregulates proteins related to cell cycle progression and upregulates proapoptotic proteins. It can reduce the expression of p38 and p65 (NFkβ), thus inhibiting the pathway of MAPKs. The purpose of this work was to observe the behavior of FK228 and MS275 alone and in combination with RT in vivo and in vitro models of RMS: RH30 (ARMS) and RD (ERMS). About, FK228 as single therapy shows limited effects but appears to radiosensitize ARMS when combined with RT. It does not show many effects in ERMS; this may be due to the inability of FK228 to inhibit HDAC3, which are the major representatives of class I HDACs in ERMS and plays a crucial role in ERMS oncogenicity. MS275 has been shown to influence tumor survival by inducing non-apoptotic death and cell cycle arrest in the G1 phase. In RH30 cells it has irreversible effects, while they are reversible in RD cells. In combination with RT, the molecule is able to prevent growth even in vivo, but only in RH30 cells, because in RD cells it showed a partial inhibitory effect. However, it is important to note that targeting HDACs class I and IV may be a potential therapeutic strategy to raise awareness of the most aggressive type of RMS (FP-RMS).