Thymidine phosphorylase (TP) catalyzes the conversion of
thymidine to
thymine and 2-deoxyribose-1-phosphate. The latter plays an important role in induction of angiogenesis. As such, many human
malignancies exhibit TP overexpression that correlates with increased microvessel density, formation of aggressive
tumors, and dismal prognosis. Because TP is frequently overexpressed in
cancer,
pro-drugs were developed that utilize TP activity for their bioactivation to cytotoxic drugs. In this respect, TP is indispensable for the pharmacologic activity of the chemotherapeutic
drug capecitabine, as it converts its intermediary metabolite 5'-deoxyfluorouridine to
5-fluorouracil. Thus, loss of TP function confers resistance to the
prodrug capecitabine, currently used for the treatment of metastatic
colorectal cancer and
breast cancer. However, drug resistance phenomena may frequently emerge that compromise the pharmacologic activity of
capecitabine. Deciphering the molecular mechanisms underlying resistance to TP-activated
prodrugs is an important goal toward the overcoming of such drug resistance phenomena. Here, we discovered that lack of TP
protein in
drug-resistant
tumor cells is due to unsplicing of its
pre-mRNA. Advanced bioinformatics identified the family of
heterogeneous nuclear ribonucleoproteins (
hnRNP) H/F as candidate
splicing factors potentially responsible for impaired TP splicing. Indeed, whereas parental cells lacked nuclear localization of hnRNPs H1/H2 and F,
drug-resistant cells harbored marked levels of these
splicing factors.
Nuclear RNA immunoprecipitation experiments established a strong binding of
hnRNP H1/H2 to TP
pre-mRNA, hence implicating them in TP splicing. Moreover, introduction of
hnRNP H2 into
drug-sensitive parental cells recapitulated aberrant TP splicing and 5'-deoxyfluorouridine resistance. Thus, this is the first study identifying altered function of
hnRNP H1/H2 in
tumor cells as a novel determinant of aberrant TP splicing thereby resulting in acquired chemoresistance to TP-activated fluoropyrimidine anticancer drugs.