Receptor‐Mediated Targeting of Phthalocyanines to Macrophages Via Covalent Coupling to Native or Maleylated Bovine Serum Albumin

Abstract

Targeted delivery of aluminum tetrasulfophthalocyanine (AIPcS,) to the scavenger receptor of macrophages, via coupling to maleylated bovine serum albumin (mal-BSA), was explored as a means to improve photodynamic efficacy. The AIPcS4 was covalently coupled to BSA (9:l molar ratio) via one or two sulfonamide-hexanoic-amide spacer chains, followed by treatment with maleic anhydride to yield the mal-BSA-phthalocyanine conjugates. The latter were tested for singlet oxygen production, receptor-mediated cell uptake and phototoxicity toward 5774 cells of macrophage origin and nonphagocytic EMT-6 cells. Cell uptake of 1251-mal-BSA showed specific binding for 5774 cells but not for EMT-6 cells. Competition studies of the conjugates with 1251-mal-BSA showed that coupling of AIPcS4 to BSA resulted in recognition of the conjugate by the scavenger receptor, whereas coupling to mal-BSA further enhanced its binding affinity. This suggests that affinity for the scavenger receptor is related to the overall negative charge of the protein. Phototoxicity of the conjugates toward 5774 cells paralleled their relative affinity, with mal-BSA-A1PcS4 coupled via two spacer chains showing the highest activity. The conjugates were less phototoxic toward the EMT-6 cell line. The activities in both cell lines of all conjugated AlPcS4 preparations were, however, lower than that of the free disulfonated AlPcS,. Possible implications for the in vivo use of protein-photosensitizer conjugates to target selectively various macrophage-associated disorders is discussed.

Scavenger-Receptor Targeted Photodynamic Therapy¶

“…The use of protein-PS conjugates to obtain cell type-specific targeting may constitute an improvement in PS delivery in PDT. One way of achieving this goal is the covalent attachment of PS to ligands of the macrophage scavenger receptor, this approach has previously been reported by us (12,13) and others (20,22). In this report we have shown that purification of the albumin conjugates in order to remove unbound lipophilic c e6 species allows specific targeting of receptor-positive J774 macrophages, with essentially no phototoxicity toward the receptor-negative OVCAR-5 cancer cells.…”

“…In a recent publication Brasseur et al (20) described the preparation and phototoxicity of series of conjugates between BSA or mal-BSA and the PS aluminum phthalocyanine tetrasulfonate. They used J774 cells and a non-scavenger receptor-expressing murine cancer cell line EMT-6.…”

“…Scavenger receptors are high capacity, lead to rapid endocytosis and subsequent routing to endosomes and lysosomes (17). The J774 murine macrophage cell line has been widely studied by workers investigating expression of scavenger receptors and targeting various drugs by conjugation to scavenger receptor ligands (18)(19)(20). Other workers have subsequently described the use of PS conjugates with maleylated serum albumin (a well-known scavenger receptor ligand [21]) to target phthalocyanines to J774 cells †Abbreviations: BSA, bovine serum albumin; c e6 , chlorin e6 ; DMSO, dimethyl sulfoxide; EDTA, ethylenediamine-tetraacetic acid; FCS, fetal calf serum; HEPES, N-2-hydroxythylpiperazine-NЈ-2ethane-sulfonic acid; mal, maleylated; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; NHS, N-hydroxysuccinimide; PBS, phosphate-buffered saline; PDT, photodynamic therapy; PS, photosensitizer; R123, rhodamine 123; SDS, sodium dodecyl sulfate; SDS-PAGE; SDS polyacrylamide gel electrophoresis; TAM, tumor-associated macrophage; TLC, thin-layer chromatography.…”

“…Other workers have subsequently described the use of PS conjugates with maleylated serum albumin (a well-known scavenger receptor ligand [21]) to target phthalocyanines to J774 cells †Abbreviations: BSA, bovine serum albumin; c e6 , chlorin e6 ; DMSO, dimethyl sulfoxide; EDTA, ethylenediamine-tetraacetic acid; FCS, fetal calf serum; HEPES, N-2-hydroxythylpiperazine-NЈ-2ethane-sulfonic acid; mal, maleylated; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; NHS, N-hydroxysuccinimide; PBS, phosphate-buffered saline; PDT, photodynamic therapy; PS, photosensitizer; R123, rhodamine 123; SDS, sodium dodecyl sulfate; SDS-PAGE; SDS polyacrylamide gel electrophoresis; TAM, tumor-associated macrophage; TLC, thin-layer chromatography. (20), and there is also a report (22) of a maleylated serum albumin conjugate being used to target the photosensitizer chlorin e6 (c e6 ) to experimental intimal hyperplasia in vivo. Tumor-associated macrophages (TAM) have been shown to play a major role in cancer growth (23).…”

Drug Delivery Systems for Phthalocyanines for Photodynamic Therapy

“…When one of these is administrated, redistribution of MPCs between albumin and LDL is expected. Covalent binding to albumin was described as well (124).…”