Irene E. Kochevar*1, Carmelo Garcia^1, and Nicholas E. Geacintov2
Chlorpromazine (CPZ) forms photoadducts with DNA and photosensitizes DNA strand breaks. These reactions may be responsible for the reported photomutagenicity of CPZ and for the well-known cutaneous and ocular phototoxicity associated with this drug. We have investigated whether CPZ molecules that are intercalated between base pairs in double-stranded (ds) DNA are the absorbing species for the photoaddition reaction. Quenching of CPZ fluorescence by ds-DNA gave nonlinear Stern-Volmer plots, indicating that more than one type of complex is formed. Linear dichroism spectra of CPZ in the presence of ds-DNA showed a minimum at 345 nm, indicating that the absorption maxima of intercalation complex(es) are red-shifted compared to the absorption maximum of free CPZ at 307 nm. The sum of the absorption of all CPZ complexes with ds-DNA, obtained from dialysis experiments, was broadened and maximized at about 315 nm, indicating that complexes not involving intercalation dominate the absorption spectrum at lambda
The mechanism whereby certain drugs cause adverse sunlight-induced cutaneous and ocular responses, a condition called drug phototoxicity, may involve photoreactions of the drug with cellular DNA. The most well-studied of these drugs are the psoralens that undergo [2 + 2] photocycloaddition reactions with pyrimidine bases when the planar psoralen molecule is intercalated between DNA base pairs. This photochemistry is believed to be a mechanism for the highly effective treatment for psoriasis and other cutaneous diseases using psoralens and UVA radiation. Reactions with DNA may be responsible for the phototoxicity of other drugs including chlorpromazine (CPZ)^^ (1,2), certain nonsteroidal anti-inflammatory drugs (3) and the recently reported fluoroquinolone antibiotics (4).
The wavelength dependence for photoaddition cannot be used to distinguish between externally bound CPZ and free CPZ as the chromophore for the photoaddition reaction although the slight red shift in the action spectrum from the absorption spectrum of free CPZ may favor the former species. The involvement of nonintercalated CPZ complexes in the photoaddition mechanism is supported by previous reports that CPZ photoadded more efficiently to single-stranded DNA, where intercalation is not possible, than to ds-DNA (8,11,13,17).
The apparent low reactivity of intercalated CPZ toward photoaddition contrasts with the requirement for intercalation prior to photoaddition found for psoralens. This difference cannot be rationalized solely on the basis of differences in the photoaddition mechanisms. The detailed mechanism for the formation of CPZ-base adducts is not known, although from the structure of one characterized adduct (6), the nominal mechanism involves dechlorination and subsequent addition of the phenothiazine radical to the 7-8 bond of the purine base. Unless intercalation does not allow the CPZ molecule to orient properly for this reaction, this reaction should be allowed because other molecules that intercalate into DNA undergo photoaddition by mechanisms involving (or likely to involve) free radical reactions with pyrimidines (27-28).
The more likely reason for the lack of photoadduct formation by intercalated CPZ molecules is that efficient nonradiative decay processes occur at these binding sites; the low fluorescence yields associated with CPZ-DNA intercalation complexes (Fig. 3) are consistent with this explanation. A thorough exploration of these interesting issues was beyond the scope of this work.
Acknowledgements-We gratefully acknowledge preliminary experiments by F.-L. Chung, many fruitful discussions with Mr. Rolando Oyola and support of this research by NIH grant S06-GM08216.
^^Abbreviations: CPZ, 2-chlorpromazine; ds-DNA, double-stranded DNA; LD, linear dichroism.
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'Wellman Laboratories of Photomedicine, Department of Dermatology, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA and
2Department of Chemistry, New York University, New York, NY, USA Received 28 May 1998; accepted 5 August 1998
* To whom correspondence should be addressed at: Wellman Laboratories of Photomedicine, Massachusetts General Hospital, WEL-224, 37 Fruit Street, Boston, MA 02114, USA. Fax: 617726-3192; e-mail: firstname.lastname@example.org ^ Permanent address: University of Puerto Rico Humacao, Department of Chemistry, Humacao, PR 00791, USA.
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