5-fluorouracil derivatives induce differentiation mediated by tubulin and HLA class I modulation

Neoplastic cells exhibit defects in their ability to differentiate; therefore, differentiation therapy represents a viable option to control cancer growth and progression. Rhabdomyosarcomas (RMS), a malignant tumor of skeletal muscle, is the most common soft tissue sarcoma in children and is characterized by its poor response to cytotoxic treatment and significant morbidity. Since modulation of alpha-tubulin and human leukocyte antigen (HLA) class I expression has been detected during malignant transformation, we analyzed in this study the expression pattern of both kinds of proteins after the treatment with 5-FU derivatives in the human RMS RD cell line. Cytotoxic assays, scanning and transmission electron microscopy, flow cytometry and immunocytochemical analyses were used. The compounds analyzed belonged to the following three categories: (a) symmetrical bis(5-fluorouracil-1-yl) derivatives with a linker that connects the N(1) atoms of both pyrimidine moieties by means of two amide bonds; and (b) an ester with the 5-FU base. The whole structure corresponds to the terminal fragment of the molecules included in (a) and (c) 5-fluorouracil acyclonucleoside-like structures. 1-[[3-(3-Chloro-2-hydroxypropoxy)-1-methoxy]propoxy]propyl]-5-fluorouracil (2), that belongs to the class (a) produced the highest increment of tubulin and its intense capillary distribution throughout the cytoplasm. On the other hand, N,N-bis[3-(5-fluorouracil-1-yl)-3-methoxypropanoyl]-alpha,alpha;-diamino-m-xylene (5) and 2 that are included in the class (c) caused the major percentage of marked cells by the HLA class I proteins. In short, our results showed that the 5-FU derivatives increase HLA class I expression and showed greater microtubule stability with an important network of tubulin beams related with the degree of differentiation of RD cells. These results could mean a more favorable prognosis of the patients affected with these tumors.