Phototoxicity of 5-aminolevulinic acid in the HeLa cell line as an indicative measure of photodynamic effect after topical administration to gynecological lesions of intraepithelial form.
OBJECTIVE
The depth-resolved pattern of aminolevulinic acid (ALA) concentration in excised vaginal tissue was determined after in vitro application of an ALA-loaded bioadhesive patch. From this data, the tissue concentration of ALA achievable at a specified depth from the surface could be related to the concentration needed to elicit a photodynamic effect in a model gynecological tumor cell line (HeLa).


METHODS
Excised vaginal tissue was mounted in a modified Franz diffusion cell and exposed to a water-soluble, ALA-loaded, bioadhesive patch. After a period of time, the tissue was cryostatically sectioned and the stratal concentration of radiolabeled ALA determined using scintillation spectroscopy. HeLa cells were cultured in media containing specific concentrations of ALA and exposed to standard photodynamic protocols of light exposure.


RESULTS
An ALA concentration of 65.6 mM was achievable at 2.375 mm from the tissue surface after application of ALA-loaded patch. The photodynamic effectiveness of this concentration was demonstrated in HeLa with exposure to concentrations exceeding 1.0 mM ALA bringing about reductions in viable cell numbers by 90%. An enhancement of PpIX production using adjunctive EDTA over the clinically relevant 4 h application time interval was shown to be minimal in HeLa. Instead, PpIX production was more closely correlated with ALA concentration, with 100 mM ALA producing approximately 3100 ng PpIX mg(-1) protein in the same time period.


CONCLUSIONS
Given that vaginal intraepithelial neoplasias can extend to 2.0 mm from the lesion surface, the ALA permeability derived from a bioadhesive patch is sufficient to induce photosensitization suitable for light induced destruction at deep sites of this type of lesion.
McCarron
Paul A
PA
School of Pharmacy, Queens University Belfast, Medical Biology Centre, Belfast BT9 7BL, UK. p.mccarron@qub.ac.uk
Donnelly
Ryan F
RF
Gilmore
Brendan F
BF
Woolfson
A David
AD
McClelland
Raymond
R
Zawislak
Agnieszka
A
Price
John H
JH
D000622:Aminolevulinic Acid / Q000008:administration & dosage / Q000633:toxicity*; D016923:Cell Death / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D017484:Dermatitis, Phototoxic / Q000473:pathology*; D015194:Diffusion Chambers, Culture; D004848:Epithelium / Q000473:pathology; D005260:Female; D005831:Genital Diseases, Female / Q000188:drug therapy* / Q000473:pathology; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D007553:Isotope Labeling; D008027:Light; D010778:Photochemotherapy; D011506:Proteins / Q000378:metabolism; D011524:Protoporphyrins / Q000378:metabolism; D014014:Tissue Adhesives; D014621:Vagina / Q000378:metabolism
D011506:Proteins; D011524:Protoporphyrins; D014014:Tissue Adhesives; D000622:Aminolevulinic Acid; C028025:protoporphyrin IX