title: Multifunctional 5-aminolevulinic acid prodrugs activating diverse cell-death pathways.
abstract: Herein we describe a series of multifunctional 5-aminolevulinic-acid (ALA) prodrugs for photodynamic dependent and independent cancer therapy (PDT). We studied the cell-death mechanisms in glioblastoma U251 cells treated with four ALA-prodrugs: (1) AlaAcBu, that releases ALA, acetaldehyde, and butyric acid; (2) AlaFaBu, that releases ALA, formaldehyde, and butyric acid; (3) AlaFaPi, that releases ALA, formaldehyde and pivalic acid (4) AlaAcPi that releases ALA, acetaldehyde and pivalic acid. We examined the light-activated and dark cell-death mechanisms of the active metabolites released from the prodrugs by unspecific cellular hydrolases. The active moieties accelerated biosynthesis of protoporphyrin IX (PpIX) due to upregulated porphobilinogen deaminase (PBGD) activity. AlaAcBu was found to be the superior prodrug for PDT due to its ability to induce the highest PpIX synthesis. Photo-irradiation of AlaAcBu-treated cells led to dissipation of the mitochondrial membrane potential and reduction in the mitochondria metabolic activities; apoptosis and necrosis. Electron microscopy analyses of these cells revealed mitochondrial and endoplasmic reticulum swelling, membrane blebbing, apoptotic bodies and necrotic cell rupture. The formaldehyde-releasing prodrugs AlaFaBu and AlaFaPi induced low PDT efficacy, moreover sequestering the formaldehyde with semicarbazide resulted in high PpIX synthesis, suggesting that formaldehyde inhibited its synthesis. ALA and AlaAcBu phototherapy resulted in a dramatic accumulation of ubiquitinated proteins due to reduced proteasome activity and expression. In conclusion, the PDT potency of the prodrugs was in the order: AlaAcBu, AlaAcPi > AlaFaBu ≥ ALA > AlaFaPi, and the superiority of AlaAcBu stems from lower molar concentrations of AlaAcBu and lower light intensity needed to activate cell death following PDT.
authors.affiliation: The Mina and Everard Goodman Life Sciences Faculty, Bar Ilan University, Ramat Gan, 52900, Israel.
authors.forename: Gili
authors.initials: G
authors.lastname: Berkovitch-Luria
authors.forename: Michal
authors.initials: M
authors.lastname: Weitman
authors.forename: Abraham
authors.initials: A
authors.lastname: Nudelman
authors.forename: Ada
authors.initials: A
authors.lastname: Rephaeli
authors.forename: Zvi
authors.initials: Z
authors.lastname: Malik
mesh_terms: D000622:Aminolevulinic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects*; D045744:Cell Line, Tumor; D005909:Glioblastoma / Q000188:drug therapy* / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D011163:Hydroxymethylbilane Synthase / Q000378:metabolism; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D010778:Photochemotherapy; D011355:Prodrugs / Q000494:pharmacology*; D011524:Protoporphyrins / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D011355:Prodrugs; D011524:Protoporphyrins; D017382:Reactive Oxygen Species; D000622:Aminolevulinic Acid; C028025:protoporphyrin IX; D011163:Hydroxymethylbilane Synthase