Triapine and a more potent dimethyl derivative induce endoplasmic reticulum stress in cancer cells
Robert Trondl1, Lea S Flocke, Christian R Kowol
1Institute of Inorganic Chemistry, University of Vienna, Vienna, Austria (R.T., L.S.F., C.R.K., M.A.J., B.K.K.); Research Platform "Translational Cancer Therapy Research" (R.T., C.R.K., P.H., U.J., M.A.J., W.B., B.K.K.), Institute of Cancer Research (P.H., U.J., W.B.), and Comprehensive Cancer Centre (P.H., U.J., W.B.), Medical University of Vienna, Vienna, Austria; Genomics Core Facility, VetCore, University of Veterinary Medicine, Vienna, Austria (G.E.M., R.S.); Department of Inorganic and Analytical Chemistry, University of Szeged, Szeged, Hungary (E.A.E.); and Hungarian Academy of Science-USZ Bioinorganic Chemistry Research Group, Szeged, Hungary (E.A.E.).
Abstract:
Triapine (3-AP; 3-aminopyridine-2-carboxaldehyde thiosemicarbazone), a ribonucleotide reductase inhibitor, has been extensively evaluated in clinical trials in the last decade. This study addresses the role of endoplasmic reticulum (ER) stress in the anticancer activity of 3-AP and the derivative N(4),N(4)-dimethyl-triapine (3-AP-Me), differing from 3-AP only by dimethylation of the terminal nitrogen. Treatment of colon cancer cells with 3-AP or 3-AP-Me activated all three ER stress pathways (PERK, IRE1a, ATF6) by phosphorylation of eIF2α and upregulation of gene expression of activating transcription factors ATF4 and ATF6. In particular, 3-AP-Me led to an upregulation of the alternatively spliced mRNA variant XBP1 (16-fold). Moreover, 3-AP and 3-AP-Me activated the cellular stress kinases c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinases, and inhibition of JNK activity antagonized the cytotoxic effect of both compounds. Subsequent to induction of the unfolded protein response, a significant upregulation of proapoptotic proteins was detected, including the transcription factor CHOP and Bim, an essential factor for ER stress-related apoptosis. In correlation with the higher degree of ER stress after 3-AP-Me treatment, also a more potent depolarization of mitochondrial membranes was found. These data suggest that 3-AP and 3-AP-Me induce apoptosis via ER stress. This was further corroborated by showing that inhibition of protein biosynthesis with cycloheximide prior to 3-AP and 3-AP-Me treatment leads to a significant reduction of the antiproliferative properties of both compounds. Taken together, this study demonstrates that induction of ER stress contributes to the mode of action of 3-AP and that terminal dimethylation leads to an even more pronounced manifestation of this effect.
Insights
Triapine (3-AP) and its derivative 3-AP-Me induce cancer cell apoptosis by activating endoplasmic reticulum (ER) stress pathways. Terminal dimethylation in 3-AP-Me enhances this ER stress-mediated anticancer effect.
Area of Science:
- Molecular Biology
- Cancer Research
- Cellular Stress Response
Background:
- Triapine (3-aminopyridine-2-carboxaldehyde thiosemicarbazone; 3-AP) is a ribonucleotide reductase inhibitor with established clinical trial evaluations.
- The role of endoplasmic reticulum (ER) stress in the anticancer mechanisms of 3-AP and its derivative N(4),N(4)-dimethyl-triapine (3-AP-Me) requires further elucidation.
Purpose of the Study:
- To investigate the involvement of ER stress in the anticancer activity of 3-AP and 3-AP-Me.
- To compare the effects of 3-AP and 3-AP-Me on ER stress pathways and downstream apoptotic events in colon cancer cells.
Main Methods:
- Treatment of colon cancer cells with 3-AP or 3-AP-Me.
- Assessment of ER stress markers including PERK, IRE1a, ATF6 pathways, eIF2α phosphorylation, ATF4 and ATF6 gene expression, and XBP1 splicing.
- Analysis of stress-activated kinases (JNK, p38), proapoptotic protein expression (CHOP, Bim), mitochondrial membrane potential, and antiproliferative effects in the presence of cycloheximide.
Main Results:
- Both 3-AP and 3-AP-Me activated all three major ER stress pathways, leading to eIF2α phosphorylation and upregulation of ATF4 and ATF6.
- 3-AP-Me induced a significant 16-fold upregulation of spliced XBP1 mRNA and a more pronounced mitochondrial membrane depolarization compared to 3-AP.
- Activation of JNK and p38 kinases was observed, and JNK inhibition antagonized the cytotoxic effects. Upregulation of CHOP and Bim indicated ER stress-induced apoptosis, which was reduced by inhibiting protein synthesis.
Conclusions:
- 3-AP and 3-AP-Me induce apoptosis in colon cancer cells primarily through the activation of ER stress.
- The terminal dimethylation in 3-AP-Me results in a more potent induction of ER stress and subsequent apoptosis compared to 3-AP.
- ER stress induction is a significant contributor to the anticancer mode of action for both Triapine and its derivative 3-AP-Me.
Related Concept Videos
01:37The Unfolded Protein Response
01:31Regulation of the Unfolded Protein Response
01:31The Intrinsic Apoptotic Pathway
03:03mTOR Signaling and Cancer Progression
The mTOR pathway or the...
01:17Role of ER in the Secretory Pathway
Components of the secretory pathway
About a third of proteins synthesized in the cell are sorted via the secretory route. They shuffle between different compartments in membrane-bound vesicles until they reach their final destination. The main intracellular compartments involved...
02:57Targeted Cancer Therapies
There are several types of targeted therapies against...