Degree of substitution of chlorin e_{6} on
charged poly- L-lysine chains affects their cellular uptake, localization and
phototoxicity towards macrophages and cancer cells
Issue title: Special Issue on Biophotonics: Sensing and
treatment with ionizing and non-ionizing radiation
Affiliations: Wellman Laboratories of Photomedicine, WEL224,
Massachusetts General Hospital, Boston, MA 02114, USA | Department of Dermatology, Harvard Medical School, MA.
02115, USA
Note: [] Corresponding author and Author to whom proofs and reprint
requests should be addressed. Fax: +1 617 726 8566; E-mail:
[email protected]
Abstract: Macromolecular photosensitizer conjugates are under investigation as
improved delivery vehicles for dyes used in photodynamic therapy. We have
previously described the use of conjugates between photosensitizers such as
chlorin_{e6} (c_{e6}) and poly-L-lysine
(pL) chains which are versatile molecular species because the size of the chain
can be varied, and the overall charge can be altered from cationic through
neutral to anionic. We now report on a series of
pL-c_{e6}conjugates in their cationic (native), neutral
(acetylated) and anionic (succinylated) forms, where the number of
c_{e6} molecules attached to each chain was varied (pL:
c_{e6} ratios, 1:4, 1:8, 1:12, and 1:16). The fluorescence
emissions were measured in both saline and a disaggregating solvent. We studied
two cell lines (an epithelial ovarian cancer, OVCAR-5 and a mouse macrophage,
J774) and measured cellular uptake, subcellular localization (by confocal
fluorescence microscopy) and phototoxicity. The cellular uptake of the
conjugates with four substitution ratios all delivered at 2 μM
c_{e6}equivalent concentration showed a maximum at 12
c_{e6} per chain for both cationic and anionic conjugates,
but the uptake of the neutral conjugate was proportional to the substitution
ratio. The macrophages took up several times more c_{e6}
than the ovarian cancer cells. Confocal fluorescence micrographs showed more
cellular fluorescence with the lower substitution ratios, and more lysosomal
localization with the cationic conjugates. The phototoxicity was much higher
for the neutral conjugates. For the cationic and neutral conjugates the 12
c_{e6} per chain was the most effective at killing cells,
while for the anionic conjugate it was the 16 c_{e6} per
chain. The anionic conjugate was better at killing OVCAR-5 cells, while the
cationic was better for J774 cells, and the neutral was approximately the same.
These data will help to optimize the parameters to be used in preparing
polymeric-photosensitizer conjugates for photodynamic therapy.
Keywords: photodynamic therapy, photosensitizer, polymeric drug conjugate, confocal fluorescence microscopy, macrophage, cancer
