The Oncogenic Protein Kinase/ATPase RIOK1 Is Up-Regulated via the c-myc/E2F Transcription Factor Axis in Prostate Cancer.

The atypical protein kinase/ATPase RIO kinase (RIOK)-1 is involved in pre-40S ribosomal subunit production, cell-cycle progression, and protein arginine N-methyltransferase 5 methylosome substrate recruitment. RIOK1 overexpression is a characteristic of several malignancies and is correlated with cancer stage, therapy resistance, poor patient survival, and other prognostic factors. However, its role in prostate cancer (PCa) is unknown. In this study, the expression, regulation, and therapeutic potential of RIOK1 in PCa were examined. RIOK1 mRNA and protein expression were elevated in PCa tissue samples and correlated with proliferative and protein homeostasis-related pathways. RIOK1 was identified as a downstream target gene of the c-myc/E2F transcription factors. Proliferation of PCa cells was significantly reduced with RIOK1 knockdown and overexpression of the dominant-negative RIOK1-D324A mutant. Biochemical inhibition of RIOK1 with toyocamycin led to strong antiproliferative effects in androgen receptor-negative and -positive PCa cell lines with EC50 values of 3.5 to 8.8 nmol/L. Rapid decreases in RIOK1 protein expression and total rRNA content, and a shift in the 28S/18S rRNA ratio, were found with toyocamycin treatment. Apoptosis was induced with toyocamycin treatment at a level similar to that with the chemotherapeutic drug docetaxel used in clinical practice. In summary, the current study indicates that RIOK1 is a part of the MYC oncogene network, and as such, could be considered for future treatment of patients with PCa.

Cell cycle arrest and apoptotic induction in LNCaP cells by MCS-C2, novel cyclin-dependent kinase inhibitor, through p53/p21WAF1/CIP1 pathway.

The purpose of the present study was to investigate the mechanisms involved in the antiproliferative and apoptotic effects of MCS-C2, a novel analog of the pyrrolo[2,3-d]pyrimidine nucleoside toyocamycin and sangivamycin, in human prostate cancer LNCaP cells. MCS-C2, a selective inhibitor of cyclin-dependent kinase, was found to inhibit cell growth in a time- and dose-dependent manner, and inhibit cell cycle progression by inducing the arrest of the G1 phase and apoptosis in LNCaP cells. When treated with 3 microM MCS-C2, inhibited proliferation associated with apoptotic induction was found in the LNCaP cells in a concentration and time-dependent manner, and nuclear DAPI staining revealed the typical nuclear features of apoptosis. Furthermore, MCS-C2 induced cell cycle arrest in the G1 phase through the upregulated phosphorylation of the p53 protein at Ser-15 and activation of its downstream target gene p21WAF1/CIP1. Accordingly, these results suggest that MCS-C2 inhibits the proliferation of LNCaP cells by way of G1-phase arrest and apoptosis in association with the regulation of multiple molecules in the cell cycle progression.

Arabinofuranosylpyrrolo[2,3-d]pyrimidines as potential agents for human cytomegalovirus infections.

Protection of the 3'- and 5'-hydroxyl groups of the nucleoside antibiotic toyocamycin (1) with 1,3-dichloro-1,1,3,3-tetraisopropyldisiloxane was followed by (trifluoromethyl)sulfonylation of the 2'-hydroxyl group. A displacement of the resulting triflate ester moiety with lithium chloride, lithium bromide, sodium iodide, and lithium azide in hexamethylphosphoramide was followed by a removal of the disilyl moiety with tetra-n-butylammonium fluoride to afford the appropriate (2'-deoxy-2'-substituted-arabinofuranosyl)toyocamycin analogues 6a-d. Hydrolysis of the carbonitrile moieties of 6a-d with hydrogen peroxide gave the corresponding sangivamycin analogues (7a-d). A reduction of the azido moiety of 6a and 7a with 1,3-propanedithiol furnished the corresponding amino derivatives (6e and 7e). The antiproliferative activity of 6a-e and 7a-e was evaluated in L1210 cell cultures. None of these compounds caused significant inhibition of cell growth. Evaluation of these compounds for antiviral activity showed that all the toyocamycin analogues were active against human CMV, but of the sangivamycin analogues, only (2'-deoxy-2'-azidoarabinosyl)sangivamycin (7a) was active against this virus. None of the compounds were active against HSV-1 or HSV-2. (2'-Deoxy-2'-aminoarabinofuranosyl)toyocamycin (6e) was studied more extensively and showed some separation between antiviral activity and cytotoxicity as measured by effects on DNA synthesis, cell growth, and cell-plating efficiency. Although 6e also was active against murine CMV in vitro, it was not active against this virus in infected mice. We conclude that arabinosylpyrrolopyrimidines have potential as antivirals, but no members of the current series are potent enough to show significant activity in vivo.

Comparison of the cellular and RNA-dependent effects of sangivamycin and toyocamycin in human colon carcinoma cells.

The effects of the pyrrolopyrimidine antibiotics sangivamycin and toyocamycin on the synthesis of RNA and protein, ribosomal RNA processing, and cell viability were examined in colon carcinoma cell line HT-29. Exposure for 24 hr to toyocamycin caused an exponential type of cell lethality resulting in a 4-log reduction of cell viability, while sangivamycin produced a gradual and self-limiting type of cell lethality resulting in a 1-log reduction of cell viability. Toyocamycin, at a concentration of 1 microM produced total cessation of precursor rRNA processing, while 10 microM sangivamycin produced little or no effect on processing. On the contrary, sangivamycin caused a significant decrease in protein synthesis after 6 hr, while toyocamycin had less effect. The inhibition of protein synthesis by sangivamycin results from an inhibition of the formation of complexes essential to the initiation of protein synthesis. The results suggest that the mechanisms of action of these closely related agents are quite distinct. The marked loss of cell viability caused by toyocamycin correlates with its effect on rRNA processing, while the slow inhibition of protein synthesis appears to be secondary to the loss of ribosome synthesis. On the other hand, the lesser cytotoxicity produced by sangivamycin results from a more direct effect on protein synthesis. Importantly, cells are much less capable of resuming normal proliferative activity after 24 hr of impaired rRNA processing than after a similar interval of reduced protein synthesis.