title: Chain-dependent photocytotoxicity of tricationic porphyrin conjugates and related mechanisms of cell death in proliferating human skin keratinocytes.
abstract: Photodynamic therapy (PDT) is a poor treatment option for nodular basal cell carcinomas and squamous cell carcinomas. As a result, the search for new photosensitizers with better effectiveness is of current interest. The photocytotoxicity of conjugates (P-R) of a water-soluble tri-cationic porphyrin (P-H) having similar efficiency of production of singlet oxygen, the PDT cytotoxin, has been assessed in vitro. Links between uptake, intracellular localization, photooxidative stress, photocytotoxicity and ability to induce programmed cell death are established. Conjugates bearing methyl (P-Me), Di-O-isopropylidene-(-d-galactopyranosyl (P-OGal) or N,N'-dicyclohexylureidooxycarbonyl (P-DDC) chains are efficiently taken-up by proliferating NCTC 2544 keratinocytes. The relative order of photocytotoxicity is P-OGal >P-DDC=P-Me≫P-H. The photocytotoxic potential of P-Me, P-OGal and P-DDC equals that of endogenous protoporphyrin IX induced by δ-aminolevulinic acid or its esters, the pro-drugs currently employed for PDT of skin lesions. Microfluorometry shows that P-Me, P-OGal, and P-DDC localize in endocytotic or pinocytotic vesicles but not in mitochondria or nucleus. Absence of annexin V binding, caspase activation or chromatin condensation suggests that cell photosensitization by P-R does not induce apoptosis. On the other hand, P-OGal photocytotoxicity correlates with appearance of multiple vesicles that have hallmarks of autophagy compartments, being decorated with the marker LC3 in cells transfected with an expression vector encoding GFP-LC3. p38 and JNK phosphorylation and inhibition of ERK1/2 phosphorylation suggest close relationship between mortality of NCTC 2544 keratinocytes and MAPK pathway impairment. Given their potentially easy formulation, water-soluble P-R are promising powerful photosensitizers for PDT of skin lesions.
authors.affiliation: Hospital de Santa Maria, Clinica Universitária de Dermatologia, Lisboa, Portugal.
authors.forename: João Nuno
authors.initials: JN
authors.lastname: Silva
authors.forename: Antoine
authors.initials: A
authors.lastname: Galmiche
authors.forename: João P C
authors.initials: JP
authors.lastname: Tomé
authors.forename: Agnès
authors.initials: A
authors.lastname: Boullier
authors.forename: Maria G P M S
authors.initials: MG
authors.lastname: Neves
authors.forename: Eduarda M P
authors.initials: EM
authors.lastname: Silva
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Capiod
authors.forename: José A S
authors.initials: JA
authors.lastname: Cavaleiro
authors.forename: René
authors.initials: R
authors.lastname: Santus
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Mazière
authors.forename: Paulo
authors.initials: P
authors.lastname: Filipe
authors.forename: Patrice
authors.initials: P
authors.lastname: Morlière
mesh_terms: D017304:Annexin A5 / Q000032:analysis; D017209:Apoptosis / Q000187:drug effects*; D001343:Autophagy / Q000187:drug effects; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D006801:Humans; D015603:Keratinocytes / Q000187:drug effects*; D008856:Microscopy, Fluorescence; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology; D018384:Oxidative Stress; D010778:Photochemotherapy; D011166:Porphyrins / Q000494:pharmacology*
chemical_list: D017304:Annexin A5; D011166:Porphyrins; D020928:Mitogen-Activated Protein Kinases; D020169:Caspases