Hybridosomes, compositions comprising the same, processes for their production and uses thereof
The present invention provides a hybrid biocompatible carrier (hybridosome) which comprises structural and bioactive elements originating from at least one biocompatible delivery module (BDM) and at least one engineered drug encapsulation module (EDEM) comprising at least one tunable fusogenic moiety. The invention further provides pharmaceutical compositions comprising said hybridosomes, processes for their manufacture, as well as pharmaceutical uses and pharmaceutical methods based thereon.
1. A process for manufacturing a hybrid biocompatible carrier (hybridosome), said process comprising contacting a first vesicle with a second vesicle, thereby uniting said first vesicle with said second vesicle and producing said hybridosome,
wherein said first vesicle has been produced in vitro, and said first vesicle comprises a fusogenic, ionizable cationic lipid, and wherein said first vesicle encapsulates a bioactive agent;
wherein said second vesicle comprises a lipid bilayer, and wherein said second vesicle has been produced in vivo and is released into the extracellular environment; and
wherein the contacting step is performed at a pH at which the fusogenic, ionizable cationic lipid is predominantly in a charged, cationic form.
2. The process of claim 1 , wherein the contacting is performed in at least one of:
a. in a buffer having a pH between 4 and 6; and
b. at a reaction temperature of about 37° C.
3. The process of claim 1 , wherein said first vesicle comprises an ionizable cationic lipid at a molar concentration of at least 30% of total lipid of the first vesicle.
4. The process of claim 1 , wherein the bioactive agent is a therapeutic agent.
5. The process of claim 1 , wherein the bioactive agent is:
a. a drug or a pharmaceutically acceptable salt or derivative thereof;
b. an antibody-based therapeutic agent; or
c. a peptide, a protein, or a nucleic acid.
6. The process of claim 1 , wherein:
a. said first vesicle is selected from the group consisting of lipid-based nanoparticles (LNPs), liposomes, polymer-stabilized LNPs, cerasomes, sphingosomes, polymersomes, synthetic-nanoparticle stabilized LNPs, natural membrane-derived LNPs, and natural membrane-coated LNPs, and
b. said second vesicle is selected from the group consisting of exosomes, ectosomes, microvesicles and apoptotic bodies.
7. The process of claim 1 , wherein said ionizable cationic lipid is selected from the group consisting of 1,2-Dilinoleyloxy-N,N-dimethylaminopropane (DLinDMA), 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1,3]-dioxolane (DLin-KC2-DMA), heptatriaconta-6,9,28,31-tetraen-19-yl4-(dimethylamino)butanoate (DLin-MC3-DMA), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), N-(4-carboxybenzyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOBAQ), YSKO5, 4-(((2,3-bis(oleoyloxy)propyl)-(methyl)amino)methyl)benzoic acid (DOBAT), N-(4-carboxybenzyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOBAQ), 3-((2,3-bis(oleoyloxy)propyl)(methyl)amino)propanoic acid (DOPAT), N-(2-carboxypropyl)-N,N-dimethyl-2,3-bis-(oleoyloxy)-propan-1-aminium (DOMPAQ), N-(carboxymethyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOAAQ), Alny-100, 3-(dimethylamino)-propyl(12Z, 15Z)-3-[(9Z,12Z)-octadeca-9,12-dien-1-yl]-henicosa-12,15-dienoate (DMAP-BLP), and derivatives of ionizable amino-lipids.
8. The process of claim 1 , wherein the process further comprises terminating said uniting process by increasing the pH of the environment of said contacting step.
9. A hybridosome, wherein said hybridosome is manufactured by a process comprising contacting a first vesicle with a second vesicle, thereby uniting said first vesicle with said second vesicle and producing said hybridosome,
wherein said first vesicle has been produced in vitro, and said first vesicle comprises a fusogenic, ionizable cationic lipid at a molar concentration of at least 30% of total lipid of the first vesicle,
wherein said first vesicle encapsulates a bioactive agent; and
wherein said second vesicle comprises a lipid bilayer, and wherein said second vesicle has been produced in vivo and is released into the extracellular environment.
10. The hybridosome of claim 9 , wherein the contacting is performed in at least one of:
a. in a buffer having a pH between 4 and 6; and
b. at a reaction temperature of about 37° C.
11. The hybridosome of claim 9 , wherein the bioactive agent is a therapeutic agent.
12. The hybridosome of claim 9 , wherein the bioactive agent is:
a. a drug or a pharmaceutically acceptable salt or derivative thereof;
b. an antibody-based therapeutic agent; or
c. a peptide, a protein, or a nucleic acid.
13. The hybridosome of claim 9 , wherein:
a. said first vesicle is selected from the group consisting of lipid-based nanoparticles (LNPs), liposomes, polymer-stabilized LNPs, cerasomes, sphingosomes, polymersomes, synthetic-nanoparticle stabilized LNPs, natural membrane-derived LNPs, and natural membrane-coated LNPs, and
b. said second vesicle is selected from the group consisting of exosomes, ectosomes, microvesicles and apoptotic bodies.
14. The hybridosome of claim 9 , wherein said ionizable cationic lipid is selected from the group consisting of 1,2-Dilinoleyloxy-N,N-dimethylaminopropane (DLinDMA), 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1,3]-dioxolane (DLin-KC2-DMA), heptatriaconta-6,9,28,31-tetraen-19-yl4-(dimethylamino)butanoate (DLin-MC3-DMA), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), N-(4-carboxybenzyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOBAQ), YSKO5, 4-(((2,3-bis(oleoyloxy)propyl)-(methyl)amino)methyl)benzoic acid (DOBAT), N-(4-carboxybenzyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOBAQ), 3-((2,3-bis(oleoyloxy)propyl)(methyl)amino)propanoic acid (DOPAT), N-(2-carboxypropyl)-N,N-dimethyl-2,3-bis-(oleoyloxy)-propan-1-aminium (DOMPAQ), N-(carboxymethyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOAAQ), Alny-100, 3-(dimethylamino)-propyl(12Z, 15Z)-3-[(9Z,12Z)-octadeca-9,12-dien-1-yl]-henicosa-12,15-dienoate (DMAP-BLP), and derivatives of ionizable amino-lipids.
15. A pharmaceutical composition comprising the hybridosome of claim 9 .
16. A method for delivering a bioactive agent into a cell, wherein said method comprises contacting a hybridosome comprising the bioactive agent with the cell, and wherein said hybridosome is manufactured by a process comprising contacting a first vesicle with a second vesicle, thereby uniting said first vesicle with said second vesicle and producing said hybridosome,
wherein said first vesicle has been produced in vitro, and said first vesicle comprises a fusogenic, ionizable cationic lipid at a molar concentration of at least 30% of total lipid of the first vesicle,
wherein said first vesicle encapsulates the bioactive agent; and
wherein said second vesicle comprises a lipid bilayer, and wherein said second vesicle has been produced in vivo and is released into the extracellular environment.
17. The method of claim 16 , wherein the contacting of said first and second vesicles is performed in at least one of:
a. in a buffer having a pH between 4 and 6; and
b. at a reaction temperature of about 37° C.
18. The method of claim 16 , wherein the bioactive agent is a therapeutic agent.
19. The method of claim 16 , wherein the bioactive agent is
a. a drug or a pharmaceutically acceptable salt or derivative thereof;
b. an antibody-based therapeutic agent; or
c. a peptide, a protein, or a nucleic acid.
20. The method of claim 16 , wherein the cell is a leukocyte, a glial cell or a cell during an ex vivo expansion.
21. The method of claim 16 , wherein:
a. said first vesicle is selected from the group consisting of lipid-based nanoparticles (LNPs), liposomes, polymer-stabilized LNPs, cerasomes, sphingosomes, polymersomes, synthetic-nanoparticle stabilized LNPs, natural membrane-derived LNPs, and natural membrane-coated LNPs, and
b. said second vesicle is selected from the group consisting of exosomes, ectosomes, microvesicles and apoptotic bodies.
22. The method of claim 16 , wherein said ionizable cationic lipid is selected from the group consisting of 1,2-Dilinoleyloxy-N,N-dimethylaminopropane (DLinDMA), 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1,3]-dioxolane (DLin-KC2-DMA), heptatriaconta-6,9,28,31-tetraen-19-yl4-(dimethylamino)butanoate (DLin-MC3-DMA), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), N-(4-carboxybenzyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOBAQ), YSKO5, 4-(((2,3-bis(oleoyloxy)propyl)-(methyl)amino)methyl)benzoic acid (DOBAT), N-(4-carboxybenzyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOBAQ), 3-((2,3-bis(oleoyloxy)propyl)(methyl)amino)propanoic acid (DOPAT), N-(2-carboxypropyl)-N,N-dimethyl-2,3-bis-(oleoyloxy)-propan-1-aminium (DOMPAQ), N-(carboxymethyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOAAQ), Alny-100, 3-(dimethylamino)-propyl(12Z, 15Z)-3-[(9Z,12Z)-octadeca-9,12-dien-1-yl]-henicosa-12,15-dienoate (DMAP-BLP), and derivatives of ionizable amino-lipids.
23. The method of claim 16 , wherein said first vesicle further comprises a targeting moiety, wherein the targeting moiety is selected from the group consisting of antibodies or fragments thereof, antibody-like molecules, peptides, proteins, aptamers, oligonucleotides and polysaccharides.
24. The method of claim 16 , wherein said first vesicle further comprises a PEG-modified lipid selected from the group consisting of a PEG-phospholipid, PEG-modified phosphatidylethanolamine (PEG-PE), PEG-modified ceramides, PEG-modified dialkylamines, PEG-modified diacylglycerols, polyethylene glycol dipalmitoylglycerol (PEG-DPG), PEG-modified dialkylglycerols, (methoxy polyethylene glycol)-dimyristolglycerol (PEG-s-DMG), a PEG-dialkyloxypropyl (DAA),R-3-[(w-methoxy-poly(ethyleneglycol)2000)carbamoyl)]-1,2-dimyristyloxypropyl-3-amine (PEG-c-DOMG), and N-acetylgalactosamine-((R)-2,3-bis(octadecyloxy)propyl-1-(methoxy-poly(ethylene glycol)2000)propylcarbamate)) (GaINAc-PEG-DSG).
25. The method of claim 16 , wherein said second vesicle is derived from:
a. a tumor cell of a cancer or pre-cancer patient, or is derived from a tumor or cancer cell line;
b. a glioblastoma cells or a mantle cell lymphoma cell;
c. a cell selected from the group consisting of B-cells, antigen presenting cells, lymphocytes, thrombocytes, neutrophils, activated polymorphonuclear neutrophils and leukocytes;
d. a bacterial pathogen, amoebic pathogen, parasitic pathogen or fungal pathogen; or
e. a pathogen infected cell.
26. The method of claim 16 , wherein said first vesicle further comprises a nucleic acid selected from the group consisting of small interfering RNA (siRNA), antisense RNA, micro RNA (miRNA), small or short hairpin RNA (shRNA), guide RNA (gRNA), clustered regularly interspaced short palindromic repeat RNA (crRNA), trans-activating clustered regularly interspaced short palindromic repeat RNA (tracrRNA), immune-stimulating oligonucleotides, plasmids, antisense nucleic acids and ribozymes.
27. The method of claim 16 wherein:
a. said first vesicle comprises at least one of a modified nucleic acid molecule and mRNA which encodes at least one antigen; or
b. said second vesicle comprises a disease-associated antigen selected from the group consisting of a tumor-associated antigen and a pathogen-associated antigen.
28. The method of claim 16 , wherein said first vesicle comprises an agent selected from the group consisting of:
a. a radioisotope selected from 225Ac, 72As, 211At, 11B, 128Ba, 212Bi, 75Br, 77Br, 14C, 109Cd, 62Cu, 64Cu, 67Cu, 18F, 67Ga, 68Ga, 3H, 123I, 125I, 130I, 131I, 111In, 177Lu, 13N, 15O, 32P, 33P, 212Pb, 186Re, 188Re, 47Sc, 153Sm, 89Sr, 99mTc, 88Y, and 90Y; and
b. a quantum dot, and a metal nanoparticle selected from a gold, a silver, an iron oxide and an iron nanoparticle.
29. A hybridosome comprising
a therapeutic agent encapsulated by a membrane of the hybridosome; wherein the membrane of the hybridosome comprises:
a. elements of a membrane of a first vesicle, wherein the elements of a membrane of the first vesicle comprise a fusogenic, ionizable cationic lipid, wherein said fusogenic, ionizable cationic lipid has at least one protonatable or deprotonatable group, such that the lipid is positively charged at a pH below physiological pH and neutral above physiological pH, and wherein said first vesicle has been produced in vitro such that the fusogenic, ionizable cationic lipid is present at a molar concentration of at least 30% of total lipid of the first vesicle; and
b. elements of a lipid bilayer of a second vesicle, wherein said second vesicle has been produced in vivo and is released into the extracellular environment.
30. The hybridosome of claim 29 , wherein
a. the first vesicle is an exosome, ectosome, microvesicle, or apoptotic body; and
b. the second vesicle is a lipid-based nanoparticle (LNP), liposome, polymer-stabilized LNP, cerasome, sphingosome, polymersome, synthetic-nanoparticle stabilized LNP, natural membrane-derived LNP, or natural membrane-coated LNP.
31. The hybridosome of claim 29 , wherein
a. the first vesicle is an exosome; or
b. the second vesicle is an LNP.
32. The hybridosome of claim 29 , wherein
a. the first vesicle is an exosome; and
b. the second vesicle is an LNP.
33. The hybridosome of claim 29 wherein the hybridosome comprises a lipid of an exosome, ectosome, microvesicle, or apoptotic body.
34. The hybridosome of claim 33 , wherein the exosome, ectosome, microvesicle, or apoptotic body is a naturally secreted exosome, ectosome, microvesicle or apoptotic body.
35. The hybridosome of claim 29 wherein the hybridosome further comprises a targeting moiety selected from the group consisting of antibodies or fragments thereof, antibody-like molecules, peptides, proteins, aptamers, oligonucleotides, sugars, polysaccharides and vitamins.
36. The hybridosome of claim 35 , wherein said targeting moiety binds to a moiety on the cell surface of a target cell.
37. The hybridosome of claim 29 wherein the hybridosome further comprises a fusion peptide anchored to its surface, wherein said fusion peptide is selected from soluble N-ethyl maleimide sensitive factor attachment protein receptors (SNARE proteins) and synthetics mimics thereof.
38. The hybridosome of claim 29 wherein the hybridosome further comprises a PEG-modified lipid.
39. The hybridosome of claim 29 , wherein the therapeutic agent is:
a. a drug or a pharmaceutically acceptable salt or derivative thereof;
b. an antibody-based therapeutic agent; or
c. a peptide, a protein, or a nucleic acid.