We analyzed the effects of all three marine alkaloids aaptamine, demethyloxyaaptamine and isoaaptamine in NT2-R, a cisplatin-resistant s
ubline of the h
uman embryonal carcinoma cell line NT2. All aaptamines were fo
und to be eq
ually effective in both cell lines, excl
uding cross-resistance between aaptamines and cisplatin
in vitro. At the inhibitory concentration (IC
ub>50ub>), aaptamine exerted an antiproliferative effect, whereas demethyloxyaaptamine and isoaaptamine were strong ind
ucers of apoptosis. We analyzed the changes in the proteome of NT2-R cells treated with these compo
unds. 16-22 proteins were fo
und to be significantly altered, of which several were validated by Western blotting and two-dimensional Western blotting analysis. Changes in the proteome pattern freq
uently res
ulted from post-transcriptional protein modifications, i.e. phosphorylation or hyp
usination in the case of eIF5A. Altho
ugh the lists of altered proteins were heterogeneo
us and compo
und-specific, gene ontology analyses identified rather similar profiles regarding the affected molec
ular f
unctions. Ingen
uity pathway analysis by IPA p
ut the following factors in a central position of the hypothetical networks: myc and p53 for aaptamine; t
umor necrosis factor (TNF) for demethyloxyaaptamine; and all three, myc, p53, and TNF for isoaaptamine. O
ur res
ults represent an important step towards a better
understanding of the molec
ular basis
underlying the observed bioactivity of these promising marine compo
unds.
Biological significance
We characterized the mode of action of three aaptamines, marine natural compound with anti-tumor activity, using a functional proteomics approach and the cisplatin-resistant pluripotent human embryonal carcinoma cell line NT2-R. The manuscript is of particular scientific interest, as we could reveal the similarities and differences of the modes of action. Furthermore, we were able to identify several new targets of these promising compounds. We found hypusination of eIF5A to be a prominent feature exclusively of aaptamine treatment, as this was not observed upon treatment with demethyloxyaaptamine or isoaaptamine. Our results are a step towards unraveling the mode of action of these interesting compounds.