Biological properties of 4-methyl-2,7-diamino-5,10-diphenyl-4,9-diazapyrenium hydrogensulfate (ADAP).
OBJECTIVE
4-Methyl-2,7-diamino-5,10-diphenyl-4,9-diazapyrenium hydrogensulfate (ADAP) is a potential antitumor compound because of its DNA and RNA intercalating ability. In this study, cellular uptake, intracellular distribution as well as mechanism of action, antitumor activity in vitro and toxicity in vivo of ADAP were investigated.


METHODS
Based on the fluorescence properties of ADAP, its entry and distribution into live cells were analyzed by fluorescence microscopy. The in vitro antiproliferative activity was determined using MTT test. For screening of topoisomerase II-targeted effects of ADAP, the cell-free assay and immunoband depletion assay were used. Expression of the genes c-mos, c-N-ras, c-Ki-ras, c-H-ras, p53 and caspase 3 in Caco-2 cells treated with ADAP was examined by RT-PCR. Toxicity in vivo was determined using C3HHf/Bu Zgr/Hr mice treated by single or multiple doses of ADAP at a concentration of 25 mg/kg.


RESULTS
ADAP in microM concentrations entered into MIAPaCa-2 cell's cytoplasm in 5 min and into nuclei in 60 min after administration. Intracellular distribution of ADAP depended on the period of treatment time. ADAP (0.1-100 microM) strongly inhibited the growth of both mouse (FsaR, SCCVII) and human tumor cells (HeLa, Caco-2, HT-29, MIAPaCa-2, HBL, HEp-2, SW620, MCF-7) compared to its weak cytotoxicity on controls and normal cells (WI38). Results of both topoisomerase II assays showed that ADAP is not a topoisomerase II poison. Expression of investigated genes was dependent on the incubation time, except for p53 and c-H-ras. Morphological changes in tissues and organs of mice were not observed. Results of patohistological analysis have been confirmed by hematological and clinical-chemical analysis of blood of treated and non-treated animals.


CONCLUSIONS
ADAP is a strongly bioactive compound with antitumor potential in vitro. The antitumor potential in vivo remains to be identified.
Marczi
Saska
S
Department of Nuclear Medicine and Pathophysiology, Clinical Hospital Osijek, Osijek, Croatia.
Glavas-Obrovac
Ljubica
L
Belovari
Tatjana
T
Stojković
Ranko
R
Ivanković
Sinisa
S
Serić
Vatroslav
V
Piantanida
Ivo
I
Zinić
Mladen
M
D000230:Adenocarcinoma / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics; D000634:Aminoquinolines / Q000494:pharmacology* / Q000633:toxicity; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000633:toxicity; D018938:Caco-2 Cells; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D003110:Colonic Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics; D004250:DNA Topoisomerases, Type II / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004354:Drug Screening Assays, Antitumor; D005260:Female; D005562:Formazans / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D007364:Intercalating Agents / Q000494:pharmacology* / Q000633:toxicity; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D011806:Quinolinium Compounds / Q000494:pharmacology* / Q000633:toxicity; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D013778:Tetrazolium Salts / Q000378:metabolism
C532299:4-methyl-2,7-diamino-5,10-diphenyl-4,9-diazapyrenium; D000634:Aminoquinolines; D000970:Antineoplastic Agents; D005562:Formazans; D007364:Intercalating Agents; D011806:Quinolinium Compounds; D012333:RNA, Messenger; D013778:Tetrazolium Salts; C079782:MTT formazan; D004250:DNA Topoisomerases, Type II