Synergism of photodynamic and electropermeation effects on cell vitality as a novel cytotoxic agent

Abstract

The present invention is based on the discovery that electroporation of a photosensitive agent in a cell and subsequent activation of the agent provides more effective killing of the electroporated cell than cells exposed to a photosensitive agent alone. The invention provides a method and apparatus for inhibiting cell growth or enhancing cell death. The method includes providing a photosensitive agent to a cell; applying an electric pulse to the cell of a sufficient strength and duration to electroporate the cell with the photosensitive agent; and applying light of an activatable wavelength to the cell thereby activating the agent and inhibiting cell growth or enhancing cell death.

1 . A method for inhibiting cell growth or enhancing cell death, comprising: 
 (a) providing to a cell a pharmaceutically acceptable formulation that comprises a photosensitive agent and a pharmaceutically acceptable carrier, provided that the photosensitive agent is not encapsulated in liposomes;    (b) applying an electric pulse of a sufficient strength and duration to electroporate the cell to facilitate entry of the photosensitive agent into the cell; and    (c) applying light of a photoactivating wavelength to the cell, thereby activating the photosensitive agent and inhibiting cell growth or enhancing cell death.    
   
   
       2 . A method according to  claim 1 , wherein multiple electric pulses are applied.  
   
   
       3 . A method according to  claim 1 , wherein the electric pulse is characterized by at least one of: 
 an amplitude of from about 0.1 kV/cm to about 6.0 kV/cm and a duration of from about 0.1 to about 10 milliseconds.    
   
   
       4 . A method according to  claim 1 , wherein the pulse is applied using at least two electrodes, optionally at least two meander electrodes.  
   
   
       5 . A method according to  claim 1 , wherein the light is applied prior to, simultaneously with, or following the electric pulse.  
   
   
       6 . A method according to  claim 1 , wherein the light is applied by a light source selected from the group consisting of a laser, a tungsten lamp, and a near ultraviolet lamp, and wherein the light optionally (i) has a wavelength of about 300 to about 950 nm and (ii) is applied in an amount about 50 to about 1000 J/cm 2 .  
   
   
       7 . A method according to  claim 1 , further comprising applying heat to the cell, optionally in the range of about 36° C. to about 42° C.  
   
   
       8 . A method according to  claim 1 , wherein the photosensitive agent is a photooxidizing agent, optionally selected from the group consisting of (i) a cytostatic agent, optionally selected from the group consisting of daunomycin, adriamycin and actinomycin, and (ii) thiopyronin, acridine orange, Zn-phthalocyanine-sulfonate, benzoporphyrin, protoporphyrin, protoporphyrin IX, PHOTOFRIN I, PHOTOFRIN II, ANTRIN, and porphycene.  
   
   
       9 . A method according to  claim 1 , further comprising providing at least one of a sensitizing agent, optionally aevuline acid, and a visualizing agent.  
   
   
       10 . A method according to  claim 1 , wherein the subject is performed in a subject, optionally a human subject.  
   
   
       11 . A method for treating a cell proliferative disorder in a subject comprising:. 
 (a) administering to a subject, optionally a human, having or suspected of having a proliferative disorder a pharmaceutically acceptable formulation comprising a pharmaceutically acceptable carrier and (i) a photosensitive agent, provided that the photosensitive agent is not encapsulated in liposomes, or (ii) a plurality of different photosensitive agent species, wherein at least one of the photosensitive agent species is a photooxidizing agent;    (b) applying an electric pulse of a sufficient strength and duration to electroporate cells of the subject to facilitate entry of the agent(s) into said cells, wherein the electric pulse is applied using at least two electrodes, optionally at least two meander electrodes;    (c) prior to, simultaneously with, or following the electric pulse, applying light of a photoactivating wavelength to said cells to activate the agent(s) to effect treatment of the cell proliferative disorder; and, optionally, at least one of (i) a sensitizing agent, optionally aevuline acid, and (ii) a visualizing agent.    
   
   
       12 . A method according to  claim 11 , wherein the cell proliferative disorder is benign or malignant, and wherein the photosensitive agent is a photooxidizing agent, optionally selected from the group consisting of (i) a cytostatic agent, optionally selected from the group consisting of daunomycin, adriamycin, and actinomycin, and (ii) thiopyronin, acridine orange, Zn-phthalocyanine-sulfonate, benzoporphyrin, protoporphyrin, protoporphyrin IX, PHOTOFRIN I, PHOTOFRIN II, ANTRIN, and porphycene.  
   
   
       13 . A method according to  claim 11 , wherein the cell proliferative disorder is a cancer, optionally a cancer selected from the group consisting of skin cancer, a solid tumor, a metastasizing cancer and hematopoietic cancer, wherein when the cancer is a hematopoietic cancer, it optionally is histiocytic lymphoma.  
   
   
       14 . A method according to  claim 11 , wherein the electric pulse is characterized by at least one of: an amplitude of from about 0.1 kV/cm to about 6.0 kV/cm and a duration of from about 0.1 to about 10 milliseconds, and the light is applied by a light source selected from the group consisting of a laser, a tungsten lamp, and a near ultraviolet lamp, and wherein the light optionally (i) has a wavelength of about 300 to about 950 nm and.(ii) is applied in an amount about 50 to about 1000 J/cm  
   
   
       15 . A method according to  claim 1   1 , further comprising applying heat to the cell, optionally in the range of about 36° C. to about 42° C.  
   
   
       16 . A method according to  claim 11 , wherein at least one light conductor is combined with an electrode.  
   
   
       17 . A method, comprising: 
 (a) providing to a cell a photooxidizing agent and at least one of a sensitizing agent and a visualizing agent;    (b) prior to, simultaneously with, or following provision of the agents to the cell, applying an electric pulse to the cell of a sufficient strength and duration to electroporate the cell with the agents; and    (c) applying light of a photoactivating wavelength to the cell, thereby activating the photooxidizing agent to inhibit cell growth or enhance cell death, wherein the method is used for (i) inhibiting cell growth or enhancing cell death or (ii) treating a cell proliferative disorder.    
   
   
       18 . A method according to  claim 17 , wherein the photosensitive agent is a cytostatic agent, optionally selected from the group consisting of daunomycin, adriamycin, and actinomycin.  
   
   
       19 . A method according to  claim 17  used to (a) inhibit cell growth or enhance cell death or (b) to treat a cell proliferative disorder in a subject, optionally a human, wherein the cell proliferative disorder is benign or malignant, and wherein the photooxidizing agent is optionally selected from the group consisting of (i) a cytostatic agent, optionally selected from the group consisting of daunomycin, adriamycin, and actinomycin, and (ii) thiopyronin, acridine orange, Zn-phthalocyanine-sulfonate, benzoporphyrin, protoporphyrin, protoporphyrin IX, PHOTOFRIN I, PHOTOFRIN II, ANTRIN, and porphycene.  
   
   
       20 . A method according to  claim 11 , wherein the electric pulse is characterized by at least one of: an amplitude of from about 0.1 kV/cm to about 6.0 kV/cm and a duration of from about 0.1 to about 10 milliseconds, and the light is applied by a light source selected from the group consisting of a laser, a tungsten lamp, and a near ultraviolet lamp, and wherein the light optionally (i) has a wavelength of about 300 to about 950 nm and (ii) is applied in an amount about 50 to about 1000 J/cm 2 , and wherein the method further optionally comprises applying heat to the cell, optionally in the range of about 36° C. to about 42° C.