Therapeutic method for treating patients with acute intermittent porphyria (AIP) and other porphyric diseases

Abstract

A method for treatment or prophylaxis of disease caused by deficiency, in a subject, of an enzyme belonging to the heme biosynthetic pathway, the method comprising administering, to the subject, an effective amount of a catalyst which is said enzyme or an enzymatically equivalent part or analogue thereof. The disease is selected from the group consisting of acute intermittent porphyria (AIP), ALA deficiency porphyria (ADP), Porphyria cutanea tarda (PCT), Hereditary coproporphyria (HCP), Harderoporphyria (HDP), Variegata porphyria (VP), Congenital erythropoetic porphyria (CEP), Erythropoietic protoporphyria (EPP), and Hepatoerythropoietic porphyria (HEP). The catalyst is one or more enzymes selected from the group consisting of delta-aminolevulininic acid synthetase, delta-aminolevulinic acid dehydratase (ALAD), porphobilinogen deaminase (PBGD), uroporphyrinogen III cosythetase, uroporphyrinogen decarboxylase, coproporphyrinogen oxidase, protoporphyrinogen oxidase, and ferrochelatase, or an enzymatically equivalent part or analogue thereof. In addition the invention relates to the use of PBGD, to human recombinant PBGD and to a method of gene therapy. The invention also relates to an expression plasmid pExp1-M2-BB (Seq. ID No. 1) and to use of a DNA fragment, the EcoR I-Hind III linear fragment (seq. ID No. 2), used for transformation in the hemC disruption strategy for production of rhPBGD expressed in E. coli.

1 . A method for treatment or prophylaxis of disease caused by deficiency, in a subject, of one or more enzymes belonging to the heme biosynthetic pathway, the method comprising administering, to the subject, an effective amount of one or more catalysts which is/are said enzyme(s), or an enzymatically equivalent part or analogue thereof.  
     
     
         2 . A method according to  claim 1 , wherein the disease is selected from the group consisting of 
 acute intermittent  porphyria  (AIP),    ALA deficiency  porphyria  (ADP),      Porphyria cutanea  tarda (PCT),    Hereditary  coproporphyria  (HCP),    Harderoporphyria (HDP),      Variegata porphyria  (VP),    Congenital  erythropoetic porphyria  (CEP),      Erythropoietic protoporphyria  (EPP), and      Hepatoerythropoietic porphyria  (HEP).    
     
     
         3 . A method according to  claim 1 , wherein the catalyst is one or more enzymes selected from the group consisting of 
 delta-aminolevulininic acid synthetase,    delta-aminolevulinic acid dehydratase (ALAD),    porphobilinogen deaminase (PBGD),    uroporphyrinogen III cosythetase,    uroporphyrinogen decarboxylase,    coproporphyrinogen oxidase,    protoporphyrinogen oxidase, and    ferrochelatase,    or an enzymatically equivalent part or analogue thereof.    
     
     
         4 . A method according to  claim 1 , wherein the disease is AIP and the enzyme is PBGD or an enzymatically equivalent part or analogue thereof.  
     
     
         5 . A method according to  claim 1 , wherein the catalyst is a recombinant form of the enzyme belonging to the heme biosynthetic pathway or of the enzymatically equivalent part or analogue thereof.  
     
     
         6 . A method according to  claim 1 , wherein the catalyst is administered by a route selected from the group consisting of the intravenous route, the intraarterial route, the intracutaneous route, the subcutaneous route, the oral route, the buccal route, the intramuscular route, the anal route, the transdermic route, the intradermal route, and the intratechal route.  
     
     
         7 . A method according to  claim 1 , wherein the catalyst is formulated in an isotonic solution.  
     
     
         8 . A method according to  claim 7 , wherein the catalyst is lyophilised.  
     
     
         9 . A method according to  claim 8 , wherein the catalyst is sterile filtered.  
     
     
         10 . A method according to  claim 1 , wherein the catalyst is formulated as lipid vesicles comprising 
 phosphatidylcholine or phosphatidylethanolamine or combinations thereof.    
     
     
         11 . A method according to  claim 1 , wherein the catalyst is incorporated into erythrocyte ghosts.  
     
     
         12 . A method according to  claim 1 , wherein the catalyst is formulated as a sustained release formulation involving biodegradable microspheres.  
     
     
         13 . A method according to  claim 1 , wherein the catalyst is lyophilized in a two-compartment cartridge, where the catalyst will be in the front compartment and water for reconstitution in the rear compartment.  
     
     
         14 . A method according to  claim 13 , wherein the two compartment cartridge is combined with an injection device to administer the catalyst either by a needle or by a needle-less (high pressure) device.  
     
     
         15 . A method according to  claim 1 , wherein the catalyst is formulated in a physiological buffer containing an enhancer for nasal administration.  
     
     
         16 . A method according to  claim 1 , wherein the catalyst is formulated as an oral formulation containing lipid vesicles, such as those comprising phospatidylcholine, phosphatidylethanolamine, or sphingomyeline, or dextrane microspheres.  
     
     
         17 . A method according to  claim 1 , wherein the catalyst is formulated so as to enhance the half-life thereof in the subject's bloodstream.  
     
     
         18 . A method according to  claim 17 , wherein the catalyst has a polyethylene glycol coating.  
     
     
         19 . A method according to  claim 17 , wherein the catalyst is complexed with a heavy metal.  
     
     
         20 . A method according to  claim 1 , wherein the catalyst is an enzymatically equivalent part or analogue of the enzyme and exerts at least part of its enzymatic activity intracellularly upon administration to the subject.  
     
     
         21 . A method according to  claim 20 , wherein the catalyst is a small artificial enzyme or an organic catalyst which can polymerize porphobilinogen to hydroxymethylbilane  
     
     
         22 . A method according to  claim 1 , wherein the catalyst is said enzyme formulated in such a manner that it exerts at least part of its enzymatic activity intracellularly upon administration to the subject.  
     
     
         23 . A method according to  claim 22 , wherein the catalyst is tagged with specific carbohydrates or other liver cell specific structures for specific liver uptake.  
     
     
         24 . A method according to  claim 1 , wherein the catalyst exerts substantially all its enzymatic activity extracellularly in the bloodstream.  
     
     
         25 . A method according to  claim 24 , wherein the enzymatic activity of the catalyst on its relevant heme precursor results in a metabolic product which 1) either moves into the intracellular compartment and is converted further via the remaining steps of the heme biosynthetic pathway or 2) is excreted from the subject via urine and/or faeces.  
     
     
         26 . A method according to  claim 1 , wherein the catalyst has been prepared by a method comprising 
 a) introducing, into a suitable vector, a nucleic acid fragment which includes a nucleic acid sequence encoding the catalyst;    b) transforming a compatible host cell with the vector;    c) culturing the transformed host cell under conditions facilitating expression of the nucleic acid sequence; and    d) recovering the expression product from the culture and optionally subjecting the expression product to post-translational processing, such as in vitro protein refolding, enzymatic removal of fusion partners, alkylation of amino acid residues, and deglycosylation, so as to obtain the catalyst.    
     
     
         27 . A method according to  claim 1 , wherein the catalyst has been prepared by liquid-phase or solid-phase peptide synthesis.  
     
     
         28 . A method according to  claim 26  wherein the catalyst is free from any other biological material of human origin.  
     
     
         29 . A method according to  claim 1 , wherein the catalyst is administered at least once a day.  
     
     
         30 . A method according to  claim 29  wherein the daily dosage is in the range of 0.01-1.0 mg/kg body weight per day.  
     
     
         31 . A method according to  claim 29 , wherein the daily dosage is about 0.1 mg per kg body weight per day.  
     
     
         32 . A method according to claims  1  wherein the catalyst is a recombinant form of the enzyme.  
     
     
         33 . A method according to  claim 32  wherein the catalyst is recombinant human PBGD based on any of Seq. ID NO 3 (clone PBGD 1.1) and Seq. ID NO 4 (non-erythro PBGD 1.1.1).  
     
     
         34 . A method for treating a patient having a mutation in the PBGD gene causing an enzyme defect, comprising the use of a human PBGD cDNA sequence of either non-erythropoietic form or erythropoietic form according to whether the tissue in which PBGD should be expressed is in cells of erythroid origin or in other cells, and transfection of the patient with the relevant cDNA.  
     
     
         35 . The method according to  claim 34 , wherein the enzyme defiency is selected from enzyme defiencies resulting in a disease selected from Acute Intermittent Porphyria, (AIP), ALA deficiency  porphyria  (ADP),  Porphyria cutanea  tarda (PCT), Hereditary  coproporphyria  (HCP), Harderoporphyria (HDP),  Variegata porphyria  (VP), Congenital  erythropoietic porphyria  (CEP),  Erythropoietic protoporphyria  (EPP), and  Hepatoerythropoietic porphyria  (HEP).  
     
     
         36 . The method according to  claim 35  wherein the disease is Acute Intermittent Porphyria, (AIP).  
     
     
         37 . The method according to  claim 34 , wherein the human PBGD cDNA sequence is selected from Seq. ID NO 3 (clone PBGD 1.1) and Seq. ID NO 4 (non-erythro PBGD 1.1.1).  
     
     
         38 . The method according to  claim 34  wherein the transfection is by use of a vector selected from the group consisting of adenovirus, retrovirus and associated adenovirus vectors.  
     
     
         39 . The method according to  claim 34  wherein the PBGD transfection of the patient (erythropoeitic and/or non-erythropoietic cells) results in substantially normal PBGD activity measured as a normalisation in urinary and/or serum levels of delta-aminolevulinic acid (ALA) and porphobilonogen (PBG) compared to the levels before treatment or to a reduction in the frequency of attack of symptons.  
     
     
         40 . A method of gene therapy treatment of patients with Acute Intermittent Porphyria (AIP) by a correction of one of the specific point mutations identified causing AIP by use of chimeraplasty gene repair.  
     
     
         41 . The method according to claim  40 -comprising a delivery system for transfection which is by use of non-viral vectors formulated in a vehicle preparation comprisng one or more components selected from the group consisting of cationic phospholipids, phospholipids, phospholipids mixed with neutral lipids, lictosylated PEI, liposomes liposomes comprising mixtures of natural phopholipids and neutral lipids.  
     
     
         42 . A method according to  claim 40  wherein the mutation is selected from Table A.  
     
     
         43 . A pharmaceutically acceptable composition comprising a therapeutically or prophylactically effective amount of catalyst which is an enzyme of the heme biosynthetic pathway or an enzymatically equivalent part or analogue thereof.  
     
     
         44 . A composition according to  claim 43 , wherein the catalyst is recombinant human PBGD based on any of Seq. ID NO 3 (clone PBGD 1.1) and Seq. ID NO 4 (non-erythro PBGD 1.1.1).