Antibody-targeted photolysis: selective photodestruction of human T-cell leukemia cells using monoclonal antibody-chlorin e6 conjugates.

Abstract

Selective in vitro photodestruction of HPB-ALL human T-cell leukemia cells was accomplished using the photosensitizer chlorin e6 coupled through dextran molecules to an anti-T-cell monoclonal antibody (mAb), anti-Leu-1.Conjugates with mAb/chlorin molar ratios as high as 1:36 retained mAb binding activity, and the absorption spectrum and quantum efficiency for singlet oxygen production of bound chlorin (0.7 ± 0.2) were unchanged from that of the free photosensitizer. Phototoxicity, as measured by a clonogenic assay and by uptake of ethidium bromide, was dependent on the doses of both mAb-chlorin and 630-to 670-nm light, was enhanced by 2H20, and was observed only in target populations that bound the mAb. Similarly, free chlorin e6 in solution had no photodynamic effect in amounts 100 times more than that carried by the mAb. For this antibody-targeted system, approximately 1010 molecules of singlet oxygen were necessary to kill a cell.A principal objective in the development of chemotherapeutic strategies is to devise modalities that are capable of selectively acting on the offending cells while sparing normal tissues. Selectivity may be provided by the preferential action of a particular agent or by preferential localization of molecules that act nonspecifically. Monoclonal antibodies (mAbs) that bind to cell surface antigens have great promise for directing cytotoxic agents to appropriate sites; and a substantive body of literature has already accumulated on the use of monoclonal antibodies to enhance selectivity of cytotoxic drugs, radioisotopes, or toxins (1-4).The selectivity ofmAb-directed treatment may be reduced, however, by cross-reactivity due to shared antigens on normal tissues and by nonspecific tissue uptake (5, 6). These problems can be obviated by utilizing mAbs that carry light-activated molecules (i.e., photosensitizers, PSs), which are innocuous without illumination but which produce toxic or reactive species such as singlet oxygen (102) upon absorbing light. By confining illumination to areas containing specifically bound PSs, toxicity can be limited to target cells. Nontarget cells, which adventitiously bind the mAb-PS but which are outside the illuminated volumes, would be spared.The present report describes selective cytolysis of human T-cell leukemia cells in vitro using mAb conjugated to the PS chlorin e6, which has a molar extinction coefficient of about 59,000 M-1-cm-1 at 660 nm (7). Chlorin/mAb conjugation ratios of approximately 30:1 were achieved, and mAb and chlorin activities were retained. Selective destruction of target T cells was accomplished by irradiation of cells containing mAb-PS conjugates bound to their cell surface. MATERIALS AND METHODSChromophore Preparation. In brief, chlorin e6-monoethylenediamine monoamide (chlorin e6-A) was prepared as follows: A monoactivated species (mixed anhydride) was formed by sequential addition oftriethylamine and ethyl chloroformate to chlorin e6 dissolved in anhydrous N,N'-dimethylformamide. TLC analysis using C18-bonded silica pla...

Photodynamic targeting of human retinoblastoma cells using covalent low-density lipoprotein conjugates

“…Better methods of localization in which sensitizers are combined with carriers that recognize the target tissue may therefore be useful in certain instances in order to increase the selective accumulation of the PS within the tumour (Jori, 1990;Hasan, 1992;Hamblin and Newman, 1994). Such delivery systems include monoclonal antibodies (MAbs) (Mew et al, 1983;Oseroff et al, 1986;Jiang et al, 1991), liposomes (Jori, 1990;Jori and Reddi, 1990), low-density lipoproteins (LDLs) (Mosley et al, 1981;Jori et al, 1984;SchmidtErfurth et al, 1994) and microspheres (Bachor et al,199 la). MAbs exhibit the highest selectivity, but their uptake by the target tissue is limited by vascular barrier problems and rapid clearance by the reticuloendothelial system.…”

Antibody-targeted photolysis: in vitro studies with Sn(IV) chlorin e6 covalently bound to monoclonal antibodies using a modified dextran carrier.

“…For example, using direct attachment of radionuclide chelators to the carbohydrate of mAbs, Rodwell et al (16) reported chelator:mAb ratios of 5. To increase the ratio of active agent covalently coupled to mAb, polymeric carriers have been used as bridges between the active agent and the mAb (14,34,35 (Fig. 3) showed a sublethal damage region below 10 J/cm2 followed by logarithmic cell killing to nearly complete cell death at higher light doses.…”

“…By employing photosensitizer (PS) active agents, these potential drawbacks are minimized as a result of the double selectivity required (i.e., antigen binding coupled with light activation of the PS). Although extensive work with PS alone has been reported (11,12), comparatively few studies on mAb'PS immunoconjugates have been published (13,14).…”

PHOTOSENSITIZED PROCESSES in vivo: PROPOSED PHOTOTHERAPEUTIC APPLICATIONS*

“…Thus, hydrophobic photosensitizers may be incorporated into the lipid moiety of LDL and released inside the malignant cells at the level of the lysosomes (Barel et al, 1986). The potential of these novel procedures of photosensitizer delivery to increase the efficacy of PDT must (Oseroff et al, 1986) that different endotissular compartments of tumors may be labeled by selected photosensitizers through the concerted use of appropriate delivery systems.…”