In vitro study of the photocytotoxicity of some hypericin analogs on different cell lines

In the present study, hypericin analogs with an increased hydrophilic chara cter were synthesized. As chemical modifications after the lipophilicity/hy drophilicity balance together with the photophysical/chemical background of the molecule the influence of these structural changes on the cellular upt ake, retention and subcellular localization in HeLa cells was investigated. Besides, their photocytotoxic effects using three cell lines (HeLa, MCF-7, A431), as well as their plasma protein binding were also assessed. To asse ss the relative hydrophilic/lipophilic character of hypericin and analogs t heir retention times were determined on a reversed phase high performance l iquid chromatography (C-18) column. The retention time of all the hypericin analogs was <46 min, except for dibenzyltetramethylhypericin (118 min), wh ile the retention time of hypericin was >200 min (solvent system: methanol/ citrate buffer 30 mM pH 7; 70/30). Hypericin, hexa-, penta- and dibenzyltet ramethylhypericin displayed a potent antiproliferative effect at the nanomo lar range after photosensitization (3.6 J/cm(2)). On the contrary, photoact ivated tetrasulfonhypericin and fringelite D had no anti proliferative effe ct on the three cell lines, whereas hypericin polyethylene glycol showed on ly an intermediate cytotoxic effect on A431 cells. In dark conditions no an tiproliferative effect was observed for any photosensitizer. The anti proli ferative photoeffect correlated well with the intracellular accumulation as measured using HeLa cells. In general, the photocytotoxic hypericin analog s concentrated to a large extent, while the noncytotoxic compounds were not taken up by the HeLa cells. Furthermore, confocal laser microscopy reveale d that all photosensitizers mainly concentrated in the perinuclear region, probably corresponding with Golgi apparatus and the endoplasmic reticulum, except for tetrasulfonhypericin which located at the plasma membrane. In ad dition, the plasma protein binding studies illustrated that hypericin bind extensively to the low-density lipoproteins, while the other hypericin anal ogs were mainly bound to heavy proteins (mostly albumin) and to a small ext ent to low-density lipoproteins.