Cardiac glycosides are well known in the treatment of
cardiovascular diseases; however, their application as treatment option for
cancer patients is under discussion. We showed that the
cardiac glycoside digitoxin and its analog
AMANTADIG can inhibit the growth of
renal cell carcinoma (RCC) cell lines and increase G2/M cell cycle arrest. To identify the signaling pathways and molecular basis of this G2/M arrest,
microRNAs were profiled using
microRNA arrays.
Cardiac glycoside treatment significantly deregulated two
microRNAs, miR-2278 and miR-670-5p. Pathway enrichment analysis showed that all
cardiac glycoside treatments affected the MAPK and the axon guidance pathway. Within these pathways, three genes, MAPK1, NRAS and RAC2, were identified as in silico targets of the deregulated
miRNAs. MAPK1 and NRAS are known regulators of G2/M cell cycle arrest.
AMANTADIG treatment enhanced the expression of phosphorylated MAPK1 in 786-O cells. Secondly, we studied the expression of
survivin known to be affected by
cardiac glycosides and to regulate the G2/M cell phase.
AMANTADIG treatment upregulated the expression of the pro-apoptotic survivin-2B variant in Caki-1 and 786-O cells. Moreover, treatment with
AMANTADIG resulted in significantly lower
survivin protein expression compared to 786-O control cells. Summarizing, treatment with all
cardiac glycosides induced G2/M cell cycle arrest and downregulated the miR-2278 and miR-670-5p in microarray analysis. All
cardiac glycosides affected the MAPK-pathway and
survivin expression, both associated with the G2/M phase. Because cells in the G2/M phase are radio- and
chemotherapy sensitive,
cardiac glycosides like
AMANTADIG could potentially improve the efficacy of radio- and/or
chemotherapy in RCCs.