Direct photocleavage of HIV-DNA by quinacridine derivatives triggered by triplex formation

Amino-p-quinacridine compounds (PQs) have been shown to stabilize strongly and specifically triple-helical DNA. Moreover, these derivatives display ph otoactive properties that make them efficient DNA cleavage agents. We explo ited these two properties (triplex-specific binding and photoactivity) to s electively cleave a double-stranded (ds)DNA sequence present in the HIV-1 g enome. Cleavage was first carried out on a linearized plasmid (3300 bp) con taining the HIV polypurine tract (PPT) that allowed targeting by a triplex- forming oligonucleotide (TFO). PQ(3), the most active compound of the serie s, efficiently cleaved double-stranded DNA in the vicinity of the PPT when this sequence had formed a triplex with a 16-mer TFO. Investigation of the cleavage at the molecular level was addressed on a short DNA fragment (56 b p); the photoinduced cleavage by PQ(3) occurred only in the presence of the triple helix. Nevertheless, unusual cleavage patterns were observed: damag e was observed at guanines located 6-9 bp away from the end of the triple h elical site. This cleavage is very efficient (up to 60%), does not require alkaline treatment, and is observed on both strands. A quinacridine-TFO con jugate produced the same cleavage pattern. This observation, along with oth ers, excludes the hypothesis of a triplex-induced allosteric binding site o f PQ(3) adjacent to the damaged sequence and indicates that PQ(3) preferent ially binds in the vicinity of the 5'-triplex junction. Irradiation in the presence of TFO-conjugates with acridine (an intercalative agent) and with the tripeptide lys-tryp-lys led to a complete inhibition of the photocleava ge reaction. These results are interpreted in terms of competitive binding and of electron-transfer quenching. Together with the findings of simple me chanistic investigations, they led to the conclusion that the photoinduced damage proceeds through a direct electron transfer between the quinacridine and the guanines. This study addresses the chemical mechanism leading to s trand breakage and characterizes the particular photosensitivity of the HIV -DNA target sequence which could be an oxidative hot spot for addressed pho toinduced strand scission by photosensitizers.