Liposome formulations for treatment of cancers and drug resistance of cancers
Disclosed herein are liposomal pharmaceutical compositions encapsulating one or more anticancer agents and methods of using the liposomal pharmaceutical compositions for treating cancers, in which multiple anticancer agents can be administered to a cancer patient in a synergistic molar ratio through liposome delivery particles. Methods of preparing the liposomes encapsulating multiple anticancer agents and the pharmaceutical compositions thereof are also disclosed.
1 . A pharmaceutical composition comprising liposomes suspended in a liquid medium, wherein the liquid medium comprises water and a pH buffer agent; wherein the liposome comprises an interior compartment surrounded by an outer lipid bilayer membrane, wherein the interior compartment comprises a hydrophilic chemotherapeutic agent, a protein kinase inhibitor, and a trapping agent selected from ammonium or substituted ammonium salts of polyanionized sulfobutyl ether cyclodextrin, ammonium or substituted ammonium salts of polyanionized sulfated carbohydrates, ammonium or substituted ammonium salt of polyphosphate, transition metal salts, quaternary ammonium compounds, polyoxyethylene, and coconut amine, in an aqueous medium; wherein the lipid bilayer membrane comprises a hydrophilic inner surface forming the interior compartment, a lipophilic bilayer, and a hydrophilic outer surface in contact with the liquid medium of the composition; and wherein the hydrophilic chemotherapeutic agent and protein kinase inhibitor can be released from the liposomes in a synergistic mode.
2 . The pharmaceutical composition of claim 1 , wherein the lipid bilayer membrane comprises (a) at least 10 mol % of a phospholipid selected from the group consisting of phosphatidylcholine, phosphatidylglycerol, phosphatidylinositol, glyceroglycolipids, sphingoglycolipids, and combinations thereof; (b) 0-60 mol % cholesterol, or a derivative thereof; and (c) optionally a charged phospholipid derivatized to polyethylene glycol.
3 . The pharmaceutical composition of claim 1 , wherein the one or more lipids are independently selected from the group consisting of HSPC, DSPC, DPPC, DMPC, DSPG, mPEG-DSPE-2000, and cholesterol.
4 . The pharmaceutical composition of claim 1 , wherein the trapping agent is selected from ammonium or substituted ammonium salts of polyanionized sulfobutyl ether cyclodextrin, ammonium or substituted ammonium salts of polyanionized sulfated carbohydrates, ammonium or substituted ammonium salt of polyphosphate, transition metal salts, and quaternary ammonium compounds.
5 . The pharmaceutical composition of claim 4 , wherein the ammonium or substituted ammonium salts of polyanionized sulfobutyl ether cyclodextrin are selected from TEA-SBE-α-cyclodextrin, TEA-SBE-β-cyclodextrin, TEA-SBE-γ-cyclodextrin, Tris-SBE-α-cyclodextrin, Tris-SBE-β-cyclodextrin and Tris-SBE-γ-cyclodextrin; the ammonium or substituted ammonium salts of polyanionized sulfated carbohydrates are selected from TEA-SOS and Tris-SOS; the ammonium or substituted ammonium salt of polyphosphate is selected from triethylammonium inositol hexaphosphate and tris(hydroxymethyl) aminomethane inositol hexaphosphate; the transition metal salts are selected from salts of copper, zinc, manganese, nickel, cobalt, or the like, with halide, sulfate and gluconate; and the quaternary ammonium compounds are selected from benzalkonium chloride, benzethonium chloride, cetrimonium bromide, and stearyl dimethylbenzyl ammonium chloride.
6 . The pharmaceutical composition of claim 1 , wherein the outer surface of the lipid bilayer membrane comprises a surface negative charged lipid or a surface-modifying agent containing polyethylene glycol, wherein the molar ratio of the total lipid to the protein kinase inhibitor, or the total amount of the hydrophilic chemotherapeutic agent and protein kinase inhibitor combined when both present, is at least equivalent (1:1).
7 . The pharmaceutical composition of claim 1 , wherein the chemotherapeutic agent is selected from the group consisting of doxorubicin, cyclophosphamide, carboplatin, paclitaxel, daunorubicin, epirubicin, 5-fluorouracil, gemcitabine, eribulin, ixabepilone, methotrexate, mutamycin, mitoxantrone, vinorelbine, docetaxel, thiotepa, bleomycin, vincristine, dacarbazine, capecitabine, prednisone, camptothecin, topotecan, irinotecan, BCNU, carmustine, cis-platin, lenalidomide, and pemetrexed.
8 . The pharmaceutical composition of claim 1 , wherein the protein kinase inhibitor is selected from the group consisting of imatinib, sunitinib, afatinib, nintedanib, ponatinib, ruxolitinib, crizotinib, ibrutinib, acalabrutinib, abemaciclib, aflibercept, alectinib, avapritinib, axitinib, bosutinib, cabozantinib, capmatinib, ceritinib, cobimetinib, crizotinib, dabrafenib, dacomitinib, dasatinib, encorafenib, entrectinib, erdafitinib, erlotinib, everolimus, fedratinib, fostamatinib, gefitinib, gilteritinib, ibrutinib, lapatinib, larotrectinib, lenvatinib, lorlatinib, nintedanib, neratinib, nilotinib, netarsudil, osimertinib, pacritinib, pazopanib, pexidartinib, pemigatinib, palbociclib, ponatinib, pexidartinib, pralsetinib, quizartinib, regorafenib, ribociclib, ripretinib, selpercatinib, selumetinib, sorafenib, temsirolimus, tofacitinib, trametinib, tucatinib, upadacitinib, vandetanib, vemurafenib, zanubrutinib, and ziv-aflibercept.
9 . The pharmaceutical composition of claim 1 , wherein the interior compartment of the liposome comprises an anticancer agent or a combination selected from the group consisting of the following:
(b) doxorubicin and imatinib co-encapsulated;
(d) doxorubicin and sunitinib co-encapsulated;
(f) gemcitabine and sunitinib co-encapsulated;
(g) doxorubicin and imatinib in about 30:1 to about 1:30 molar ratio;
(h) doxorubicin and sunitinib in about 30:1 to about 1:30 molar ratio; and
(i) gemcitabine and sunitinib in about 30:1 to about 1:30 molar ratio.
10 . The pharmaceutical composition of claim 1 , wherein the liposomes have a mean particle size between 4.5 nm and 450 nm, inclusive.
11 . The pharmaceutical composition of claim 1 , wherein molar ratio of the chemotherapeutic agent and protein kinase inhibitor is such that when said ratio is provided to cancer cells relevant to said cancer in an in-vitro assay over the concentration range at which the fraction of affected cells is about 0.20 to 0.80, synergy is exhibited over at least 20% of said range.
12 . The pharmaceutical composition of claim 1 , wherein the liposome encapsulated with a chemotherapeutic agent and a protein kinase inhibitor, maintains for at least one hour of the said synergistic molar drug ratio in blood after in-vivo administration.
13 . A method of treating a cancer or drug resistance of a cancer, comprising administering to a subject in need of treatment a therapeutically effective amount of a pharmaceutical composition according to claim 1 , wherein the cancer is selected from breast cancer, melanoma, gastrointestinal cancer, lung cancer, colorectal cancer, Ewing sarcoma, pancreatic cancer, prostate cancer, bladder cancer, kidney cancer, thyroid cancer, uterine cancer, and gastrointestinal stromal tumors.
14 . The method of claim 13 , wherein the breast cancer is triple negative breast cancer, and wherein the lung cancer is caused by either a high level of phosphorylation of a wild-type EGFR or a mutation within an EGFR amino acid sequence.
15 . A method of preparing a liposomal pharmaceutical composition according to claim 1 , wherein the liposome is made by a process comprising the steps of:
(a) forming multilamellar liposome vesicles in a solution comprising water, lipid(s), and trapping agent(s) selected from ammonium or substituted ammonium salts of polyanionized sulfobutyl ether cyclodextrin, ammonium or substituted ammonium salts of polyanionized sulfated carbohydrates, ammonium or substituted ammonium salts of polyphosphate, transition metal salts, quaternary ammonium compounds, polyoxyethylene, and coconut amine;
(b) extruding the multilamellar liposome vesicles multiple times at an elevated temperature through a polycarbonate membrane to form unilamellar liposomes;
(c) substantially removing the trapping agent(s) that are outside of the liposomes by diafiltration or size exclusion chromatography, or other buffer exchanging methods; and
(d) heating the unloaded liposomes at an elevated temperature in an aqueous solution comprising a chemotherapeutic agent and a protein kinase inhibitor, thereby forming drug encapsulated liposomes.
16 . The method of claim 15 , wherein the lipid is selected from the group consisting of: (a) at least 10 mol % of a phospholipid selected from the group consisting of phosphatidylcholine, phosphatidylglycerol, phosphatidylinositol, glyceroglycolipids, sphingoglycolipids, and combinations thereof; (b) 0-60 mol % cholesterol, or a derivative thereof; and (c) optionally a charged phospholipid derivatized to polyethylene glycol.
17 . The method of claim 15 , wherein the ammonium or substituted ammonium salts of polyanionized sulfobutyl ether cyclodextrin are selected from TEA-SBE-α-cyclodextrin, TEA-SBE-β-cyclodextrin, TEA-SBE-γ-cyclodextrin, Tris-SBE-α-cyclodextrin, Tris-SBE-β-cyclodextrin and Tris-SBE-γ-cyclodextrin; the ammonium or substituted ammonium salts of polyanionized sulfated carbohydrates are selected from TEA-SOS and Tris-SOS; the ammonium or substituted ammonium salts of polyphosphate are selected from triethylammonium inositol hexaphosphate and tris(hydroxymethyl) aminomethane inositol hexaphosphate; the transition metal salts are selected from salts of copper, zinc, manganese, nickel, cobalt, or the like, with halide, sulfate, are gluconate; and the quaternary ammonium compounds are selected from benzalkonium chloride, benzethonium chloride, cetrimonium bromide, and stearyl dimethylbenzyl ammonium chloride.
18 . The method of claim 15 , comprising a process selected from: active loading, passive loading, and combination thereof; wherein the active or passive loading is selected from:
(a) pH gradient-based active loading method, which encapsulates the chemotherapeutic agent and protein kinase inhibitor based on a transmembrane pH gradient, wherein the pH value of the interior aqueous compartment of the liposome is lower than that outside the liposome;
(b) transition metal-based active loading method, which encapsulates the chemotherapeutic agent and protein kinase inhibitor by utilizing transition metal ions to drive the uptake of the chemotherapeutic agent and protein kinase inhibitor into liposomes via complexation;
(c) passive loading method, which encapsulates the chemotherapeutic agent and protein kinase inhibitor during the liposome formation; and
(d) passive loading method, which involves passive equilibration after the formation of liposomes.
19 . The method of claim 18 , wherein the pH gradient is formed by a concentration gradient of an ammonium ion or a concentration gradient of an organic compound having an ammonium derivative or substituted ammonium capable of being protonated.
20 . The method of claim 15 , wherein the chemotherapeutic agent is selected from the group consisting of doxorubicin, cyclophosphamide, carboplatin, paclitaxel, daunorubicin, epirubicin, 5-fluorouracil, gemcitabine, eribulin, ixabepilone, methotrexate, mitomycins, mitoxantrone, vinorelbine, docetaxel, thiotepa, bleomycin, vincristine, dacarbazine, capecitabine, prednisone, camptothecin, topotecan, irinotecan, carmustine, cis-platin, BCNU, lenalidomide, and pemetrexed; and wherein the protein kinase inhibitor is selected from the group consisting of imatinib, sunitinib, afatinib, nintedanib, ponatinib, ruxolitinib, crizotinib, ibrutinib, acalabrutinib, abemaciclib, aflibercept, alectinib, avapritinib, axitinib, bosutinib, cabozantinib, capmatinib, ceritinib, cobimetinib, crizotinib, dabrafenib, dacomitinib, dasatinib, encorafenib, entrectinib, erdafitinib, erlotinib, everolimus, fedratinib, fostamatinib, gefitinib, gilteritinib, ibrutinib, lapatinib, larotrectinib, lenvatinib, lorlatinib, nintedanib, neratinib, nilotinib, netarsudil, osimertinib, pacritinib, pazopanib, pexidartinib, pemigatinib, palbociclib, ponatinib, pexidartinib, pralsetinib, quizartinib, regorafenib, ribociclib, ripretinib, selpercatinib, selumetinib, sorafenib, temsirolimus, tofacitinib, trametinib, tucatinib, upadacitinib, vandetanib, vemurafenib, zanubrutinib, ziv-aflibercept, and combinations thereof.
21 . The method of claim 15 , where the liposomes have a mean particle size in the range of between 4.5 nm and 450 nm, inclusive.
22 . A treatment kit comprising a container and a plurality of the drug-loaded liposomes of claim 1 in the container, wherein the drug-loaded liposomes are or can be suspended in a sterile diluent solution ready for administration to a subject in need of treatment.