Delta-aminolevulinate-induced host-parasite porphyric disparity for selective photolysis of transgenic Leishmania in the phagolysosomes of mononuclear phagocytes: a potential novel platform for vaccine delivery.

Abstract

Leishmania double transfectants (DTs) expressing the 2nd and 3rd enzymes in the heme biosynthetic pathway were previously reported to show neogenesis of uroporphyrin I (URO) when induced with delta-aminolevulinate (ALA), the product of the 1st enzyme in the pathway. The ensuing accumulation of URO in DT promastigotes rendered them light excitable to produce reactive oxygen species (ROS), resulting in their cytolysis. Evidence is presented showing that the DTs retained wild-type infectivity to their host cells and that the intraphagolysosomal/parasitophorous vacuolar (PV) DTs remained ALA inducible for uroporphyrinogenesis/photolysis. Exposure of DT-infected cells to ALA was noted by fluorescence microscopy to result in host-parasite differential porphyrinogenesis: porphyrin fluorescence emerged first in the host cells and then in the intra-PV amastigotes. DT-infected and control cells differed qualitatively and quantitatively in their porphyrin species, consistent with the expected multi- and monoporphyrinogenic specificities of the host cells and the DTs, respectively. After ALA removal, the neogenic porphyrins were rapidly lost from the host cells but persisted as URO in the intra-PV DTs. These DTs were thus extremely light sensitive and were lysed selectively by illumination under nonstringent conditions in the relatively ROS-resistant phagolysosomes. Photolysis of the intra-PV DTs returned the distribution of major histocompatibility complex (MHC) class II molecules and the global gene expression profiles of host cells to their preinfection patterns and, when transfected with ovalbumin, released this antigen for copresentation with MHC class I molecules. These Leishmania mutants thus have considerable potential as a novel model of a universal vaccine carrier for photodynamic immunotherapy/immunoprophylaxis.