Neocarzinostatin as a probe for DNA protection activity—molecular interaction with caffeine

Abstract

Neocarzinostatin (NCS), a potent mutagen and carcinogen, consists of an enediyne prodrug and a protein carrier. It has a unique double role in that it intercalates into DNA and imposes radical-mediated damage after thiol activation. Here we employed NCS as a probe to examine the DNA-protection capability of caffeine, one of common dietary phytochemicals with potential cancer-chemopreventive activity. NCS at the nanomolar concentration range could induce significant single- and double-strand lesions in DNA, but up to 75 ± 5% of such lesions were found to be efficiently inhibited by caffeine. The percentage of inhibition was caffeine-concentration dependent, but was not sensitive to the DNA-lesion types. The well-characterized activation reactions of NCS allowed us to explore the effect of caffeine on the enediyne-generated radicals. Postactivation analyses by chromatographic and mass spectroscopic methods identified a caffeine-quenched enediyne-radical adduct, but the yield was too small to fully account for the large inhibition effect on DNA lesions. The affinity between NCS chromophore and DNA was characterized by a fluorescence-based kinetic method. The drug-DNA intercalation was hampered by caffeine, and the caffeine-induced increases in DNA-drug dissociation constant was caffeine-concentration dependent, suggesting importance of binding affinity in the protection mechanism. Caffeine has been shown to be both an effective free radical scavenger and an intercalation inhibitor. Our results demonstrated that caffeine ingeniously protected DNA against the enediyne-induced damages mainly by inhibiting DNA intercalation beforehand. The direct scavenging of the DNA-bound NCS free radicals by caffeine played only a minor role.

A positive feedback loop of SIRT1 and miR17HG promotes the repair of DNA double-stranded breaks

“…All three RNAi fragments significantly suppressed the miR17HG expression (>90%) but exhibited a minor effect on the expression of two other miR17HG isoforms (Figure 3(a)). NCS (neocarzinostatin) is a radiomimetic antitumor drug, which generates DSBs in cells [29]. We then monitored the γH2A.X levels in cells transfected with simiR17HG-1/2 or siNC with or without NCS treatment (0.5μg/ml).…”

The theory of interceptor-protector action of DNA binding drugs

“…1): d protector (Skamrova et al, 2014;Traganos et al, 1991aTraganos et al, , 1991bEvstigneev et al, 2005Evstigneev et al, , 2006aEvstigneev et al, , 2006bEvstigneev et al, , 2008Evstigneev et al, , 2011aHernandez Santiago et al, 2009;Buchelnikov et al, 2012;Davies et al, 2001;Chin et al, 2012;Maria Johnson et al, 2003a), i.e. competition of the drug and interceptor for DNA binding sites leading to removal of the drug from DNA as a result of interceptor-DNA binding, and d interceptor (Traganos et al, 1991a(Traganos et al, , 1991b(Traganos et al, , 1993Hill et al, 2011;Skamrova et al, 2014;Woziwodzka et al, 2011Woziwodzka et al, , 2013bKapuscinski et al, 2002;Ganapathi et al, 1986;Lyles and Cameron, 2002;Bedner et al, 2001;Borowik et al, 2018;Guo, 1992, 1993;Evstigneev et al, 2005Evstigneev et al, , 2006aEvstigneev et al, , 2006bEvstigneev et al, , 2008Evstigneev et al, , 2011aHernandez Santiago et al, 2009;Buchelnikov et al, 2012;Davies et al, 2001;Chin et al, 2012;Maria Johnson et al, 2003a;Piosik et al, 2002Piosik et al, , 2003Piosik et al, , 2005Piosik et al, , 2010Ulanowska et al, 2005<...>…”

“…The most typical aromatic interceptor molecules, for which the interceptor-protector hypothesis is commonly discussed as being superior to other mechanisms, belong to four groups (Fig. 2): d methylxanthines (caffeine, theophylline, pentoxifylline) (Woziwodzka et al, 2011(Woziwodzka et al, , 2013a(Woziwodzka et al, , 2013bEvstigneev, 2013;Traganos et al, 1991aTraganos et al, , 1991bTraganos et al, , 1993Hill et al, 2011;Skamrova et al, 2014;Sadzuka et al, 1995;Kakuyamanee Iwazaki and Sadzuka, 2001;Kapuscinski et al, 2002;Ganapathi et al, 1986;Lyles and Cameron, 2002;Bedner et al, 2001;Evstigneev et al, 2006aEvstigneev et al, , 2006bEvstigneev et al, , 2008Evstigneev et al, , 2011aHernandez Santiago et al, 2009;Buchelnikov et al, 2012;Davies et al, 2001;Chin et al, 2012;Maria Johnson et al, 2003a;Piosik et al, 2002Piosik et al, , 2003Piosik et al, , 2005Piosik et al, , 2010Ulanowska et al, 2005Ulanowska et al, , 2007Gołu nski et al, , 2015Gołu nski et al, , 2016Banerjee et al, 2011); d polyphenols (vitamin B 2 ) (Woziwodzka et al, 2013a;Evstigneev, 2013;Evstigneev et al, 2005Evstigneev et al, , 2008; Kharasch and Novak, 1981;Raiczyk and Pinto, 1988;…”

Physicochemical Mechanisms of Synergistic Biological Action of Combinations of Aromatic Heterocyclic Compounds

“…To date, it is known that, on simultaneous addition or right after the main drug, CAF diminishes the toxicity of the antitumour antibiotics doxorubicin [30,31,33,[37][38][39], mitoxantrone [32,33,37,39], ellipticine [33,38], amsacrine [34], camptothecins [38,40], and phenothiazine drugs [41] as well as the aromatic mutagen ethidium bromide [42] and aromatic neurotoxin tetrahydropyridine [43]. The "protector" effect has also been reported with respect to the aromatic mutagen quinacrine, ICR-191, ICR-170, IQ-type heterocyclic aromatic amines, and neocarzinostatin in the presence of caffeine and its derivatives [44][45][46][47]. Two basic mechanisms responsible for the observed effects in cells have initially been suggested [32,33]: hetero-association of caffeine and the aromatic drug and competitive binding of CAF with DNA ( Figure 3).…”