ORIGINAL RESEARCH

Abstract

Antikanser ilaçların çoğunun hücrede sıklıkla bulunan DNA topoizomeraz reaksiyonları ile girişim yaparak etkinliklerini ortaya koyduğu gösterilmiştir. Nitekim kamfotekinin (camptothecin) topoizomeraz I enzimini hedefleyen bileşik olarak tanımlanmasından itibaren sentetik ve doğal bileşiklerin farmasötik önemini değerlendirmek üzere biyolojik çalışmalarda bu enzimlerden geniş ölçüde yararlanılmıştır. Önemli sayıda bileşiğin topoizomeraz reaksiyonları üzerinden sitotoksik etkinliğe sahip olmaları nedeniyle asetonaftonun ve türevinin daha önce rapor edilen fizyolojik aktivitelerinin topoizomeraz reaksiyonlarını içerip içermediklerinin araştırılması hedeflenmiştir. Başlangıç molekülü olan asetonaftonun (AN) biyoaktivitesi ile karşılaştırılmak üzere bileşikte bulunan aminometilin biyoaktiviteye etkisi, Mannich bazı türevi olan piperidinopropiyonafton hidroklorür bileşiği (MB) ile topoizomeraz aktivite ve sitostatik aktivite ölçümleri kullanılarak değerlendirilmiştir. MB laboratuvarımızda sentezlenip karakterize edilmiştir. Memeli DNA topoizomerazları üzerindeki biyolojik aktivitelerini değerlendirmek üzere süpersarmal plazmid relaksasyon ve dekatenasyon deneyleri gerçekleştirilmiştir. Ayrıca HeLa, MCF7 ve A431 hücre hatlarında sitostatik aktivite araştırılmıştır. Verilerimize göre sitostatik değerlendirmelerden bağımsız olarak MB, tip I ve tip II DNA topoizomeraz üzerinde dikkate değer bir etki göstermiştir. MB, MCF7 hücrelerindeki proliferasyona karşı 27.62 µM olan IC50 değeri ile ortalama bir etki göstermiştir. Bununla birlikte MB varlığı topoizomeraz II dekatenasyon aktivitesini etkilemiştir. In vitro koşullarda sonuçlarımız DNA topoizomerazlar ve kanser hücrelerinin proliferasyonu üzerindeki etkilerini direkt olarak açıklamamaktadır. Sonuçlarımız Mannich bazının ilaç geliştirme çalışmalarındaki potansiyel önemi üzerinden değerlendirilmiştir.

The interference of piperidinopropionaphthone hydrochloride in mammalian type I and type II DNA topoisomerase reactions

Abstract

Majority of anti-cancer drugs were shown to exert their activities by interfering with DNA topoisomerase reactions. Since the identification of Camptothecin as the topoisomerase I targeting compound, these enzymes are widely utilized in biological assays to assess the pharmaceutical significance of the synthetic and natural agents. Because a considerable number of compounds were shown to have cytostatic activities via blocking topoisomerase reactions, we aimed to identify if the previously-reported physiological activities of acetonapthones involves the interference with topoisomerase reactions. We covered topoisomerase activity and cytostatic activity evaluation of piperidinopropionaphthone hydrochloride type Mannich base (MB) to compare its bioactivities to the starting propionaphtone in order to assess the contribution of aminomethyl moiety of the compound on its bioactivity. MB was synthesized and characterized in our laboratory. Supercoiled plasmid relaxation and decatenation assays were carried out to evaluate their biological activities in mammalian DNA topoisomerases. We also assayed the cytostatic activities using HeLa, MCF7 and A431 cell lines. Our data showed a considerable inhibition of MB on type I and type II DNA topoisomerases without a correlation to cytostatic assays. MB exerted a modest activity against the proliferation of MCF7 cells with an IC50 value of 27.62 μM. The presence of MB inhibited topo II decatenation activity as well. Results offer no direct explanation for the contradictory effects on the DNA topoisomerases and the proliferation of cancer cells in vitro. Our results are discussed in relation to potential significance of aminomethyl group of Mannich base in the course of drug-development studies. 

Keywords

• Mannich base
• Anti-cancer drugs
• Decatenation

References

1. Denton JJ, Turner RJ, Neier WB, Lawson VA, Schedl HP. Antispasmodics. I. Substituted β -amino ketones. J Am Chem Soc, 1949; 71:2048-2050.
2. Philippot E, Denys W, Dallemagne MJ. Ganglioplegic action of several derivatives of 2,2,6,6-tetramethylpiperidine. Arch Int Pharmacodyn Ther, 1962; 135:273-280.
3. Flynn GA, Lee SA, Faris M, Brandt DW, Chakravarty S. Preparation of aryl ketone derivatives as intracellular kinase inhibitors. PCT / WO 2007; 2007/136790 (A2).
4. Gordon PN, Johnston JD, English AR. β-Amino ketones as anti-infective agents. In: Hobby, GL (ed), Antimicrobial Agents and Chemotherapy. Am Soc Microbiol, Bethesda, 1965; pp 165-167.
5. Dimmock JR, Sidhu KK, Chen M, Reid RS, Allen TM, Kao GY, Truitt GA. Evaluation of some Mannich bases of cycloalkanones and related compounds for cytotoxic activity. Eur J Med Chem 1993; 28:313-322.
6. Gul HI, Gul M, Erciyas E. Syntheses and Stability Studies of Some Mannich Bases of Acetophenones and Evaluation of their Cytotoxicity Against Jurkat Cells. Arzneimittel-Forsch 2002; 52:628-635.
7. Tramontini M, Angiolini L, Ghedini N. Mannich bases in polymer chemistry. Polymer 1988; 29:771-788.
8. Friedman M, Cavins JF, Wall JS. Relative Nucleophilic Reactivities of Amino Groups and Mercaptide Ions in Addition Reactions with α, β-Unsaturated Compounds. J Am Chem Soc 1965; 87:3672-3682.

9. Erciyas E, Erkaleli HI, Cosar G. Antimicrobial Evaluation of Some Styryl Ketone Derivatives and Related Thiol Adducts. J Pharm Sci 1994; 83:545-548.
10. Barranco SC, Townsend CM Jr, Weintraub B, Beasley EG, MacLean KK, Shaeffer J, Liu NH, Schellenberg K. Changes in glutathione content and resistance to anticancer agents in human stomach cancer cells induced by treatments with melphalan in vitro. Cancer Res 1990; 50:3614-3618.
11. -
12. Medh RD, Gupta V, Awasthi YC. Reversal of melphalan resistance in vivo and in vitro by modulation of glutathione metabolism. Biochemical Pharmacology 1991; 42:439-441.
13. Schauenstein E, Noehammer G, Rauch HJ, Kresbach H. Significant decreases in the intensity of staining for proteins and protein thiols in basal-cell epitheliomas (basaliomas) as compared to normal skin. Histochemistry 1985; 83:451-454.
14. Stern H. Sulfhydryl groups and cell division. Science (Washington DC) 1956; 124:1292-1293.
15. Alpan AS, Gunes HS, Topcu Z. 1H-Benzimidazole derivatives as mammalian DNA Topoisomerase I inhibitors. Acta Biochim Pol 2007; 54:561-565.
16. Chhatriwala H, Jafri N, Salgia R. A Review of Topoisomerase Inhibition in Lung Cancer. Cancer Biol Ther 2006; 12:1600-1607.
17. Ishar MPS, Singh G, Singh S, Sreenivasan KK, Singh G. Design, synthesis, and evluation of novel 6-chloro-/fluorochromone derivatives as potential topoisomerase inhibitor anticancer agents. Bioorg Med Chem Lett 2006; 16:1366–1370.
18. Kucukoglu O, Ozturk B, Kamataki T, Topcu Z. Inhibitory Activities of Helichrysum taxa on Mammalian Type I DNA topoisomerase. Pharm Bio 2006; 44:189-193.
19. López-Lázaro M, Martín-Cordero C, Toro MV, Ayuso MJ. Flavonoids As DNA Topoisomerase I Poisons. J Enzyme Inhib Med Chem 2002; 17:25-29.
20. Martín-Cordero C, López-Lázaro M, Gálvez M, Ayuso MJ. Curcumin as a DNA Topoisomerase II Poison. J Enzyme Inhib Med Chem 2003; 18:505-509.
21. Topcu Z, Ozturk B, Kucukoglu O, Kilinc E. Flavonoids in Helichrysum pamphylicum Inhibit Mammalian Type I DNA Topoisomerase. Z Naturforsch C 2008; 63:69-74.
22. Topcu Z, Borden KLB. The yeast two-hybrid system and its pharmaceutical significance. Pharm Res 2000;17:1049-1055.
23. Wang JC. DNA topoisomerases. Ann Rev Biochem 1996; 65:635-692.
24. Bjornsti MA, Osheroff N. DNA topology and enzymes. Humana Press, Totowa, New Jersey, 1999.
25. Dimmock JR, Erciyas E, Bigam GE, Kirkpatrick DL, Duke MM. Intramolecular cyclization and cytotoxicities of some Mannich bases of styryl ketones. Eur J Med Chem 1989; 24:379-383.
26. Istanbullu H, Erzurumlu Y, Ballar Kirmizibayrak P, Erciyas E. Evaluation of Alkylating and Intercalating Properties of Mannich Bases for Cytotoxic Activity. Letters in Drug Des Discove, 2014:11:1096-1106.
27. Topcu Z and Castora FJ. Mammalian Mitochondrial DNA Topoisomerase I Preferentially Relaxes Supercoils in Plasmids Containing Specific Mitochondrial DNA Sequences. Biochim Biophys Acta 1996; 1264:377-387.
28. Sobhani AM., Ebrahimi SA, Mahmoudian M. An In Vitro Evaluation of Human DNA Topoisomerase I Inhibition by Peganum harmala L. Seeds Extract and Its β-Carboline Alkaloids. J Pharm Pharmaceut Sci 2002; 5:19-23.
29. Marini JC, Miller KG, Englund PT. Decatenation of Kinetoplast DNA by Topoisomerases. J Biol Chem 1980; 255:4976-4979.
30. Mosmann T. Rapid Colorimetric Assay for Cellular Growth and Survival: Application to Proliferation And Cytotoxicity Assays. J Immunol Methods 1983; 65:55-63.
31. Dimmock JR, Erciyas E, Kirkpatrick DL, King KM. Evaluation of some azines of aminomethylacetophenones and related quaternary ammonium compounds versus the EMT6 tumor. Pharmazie 1988; 43:614-616.
32. Réthy B, Zupkó I, Minorics R, Hohmann J, Ocsovszki I, Falkay G. Investigation of Cytotoxic Activity on Human Cancer Cell Lines of Arborinine and Furanoacridones Isolated from Ruta Graveolens. Planta Med 2007; 73:41-48.