The administration route of immune agonist
阅读说明：本技术 免疫激动剂的施用路径 (The administration route of immune agonist ) 是由 G·阿库纳 E·R·科斯 J·沙罗 M·莱赫曼 M·斯特恩 于 2018-05-31 设计创作，主要内容包括：本文中公开的是包含免疫激动性抗体的液体药学制剂的一种新用途,其在至少维持相同水平的所述免疫激动剂的生物学活性的同时导致改善的耐受性/不利事件概况。(Disclosed herein is a kind of new application of the Liquid pharmaceutical preparation comprising immune agonistic antibody, leads to improved tolerance/adverse events overview while at least maintaining the biological activity of the immune agonist of phase same level.)
1. for the Liquid pharmaceutical preparation comprising immune agonist used in treating cancer or infectious disease, wherein the Liquid pharmaceutical Preparation subcutaneous administration.
2. for the Liquid pharmaceutical preparation used according to claim 1, it is characterised in that it includes aqueous solution, which includes:
(a) agonist is about immunized described in 1-100mg/ml；
(b) about 10-30mM is at least one provides the buffer of pH 5 to 6；With
(c) about 0.01 to 0.1% (w/v) nonionic surface active agent.
3. for the Liquid pharmaceutical preparation used according to claims 1 or 2, it is characterised in that it includes aqueous solution, the aqueous solution packet Contain:
(a) agonist is about immunized described in 10-80mg/ml；
(b) about 20mM is at least one provides the buffer of pH about 5.5；With
(c) about 0.02% (w/v) nonionic surface active agent.
4. further including about 180-300mM sucrose for the Liquid pharmaceutical preparation used according to any one of claims 1 to 3 About 0-20mM methionine.
5. further including about 240mM sucrose and about 0-10mM first sulphur for the Liquid pharmaceutical preparation used according to claim 4 Propylhomoserin.
6. for the Liquid pharmaceutical preparation used according to any one of claim 2 to 5, wherein the buffer is acetate or amber Hydrochlorate buffer, such as sodium acetate or sodium succinate or combinations thereof optionally further include about 140mM sodium chloride.
7. for the Liquid pharmaceutical preparation used according to any one of claim 2 to 6, wherein the nonionic surface active agent is PLURONICS F87 or polysorbate 20 or polyoxyethylene sorbitan monoleate.
8. wherein the immune agonist is excitability CD40 for the Liquid pharmaceutical preparation used according to any one of claim 1 to 7 Antibody.
9. for the Liquid pharmaceutical preparation used according to any one of claim 1 to 8, it is characterised in that it is containing about 10mg/ml Excitability anti-CD 40 antibodies；About 20mM sodium succinate and about 0.02% (w/v) are selected from polysorbate 20, polyoxyethylene sorbitan monoleate or pool Lip river The surfactant of husky nurse 188, the aqueous solution in about 5.5 pH.
10. wherein the immune agonist is comprising SEQ for the Liquid pharmaceutical preparation used according to any one of claim 1 to 9 The excitability anti-CD 40 antibodies of the heavy chain variable domain (VH) of the light-chain variable domain (VL) and SEQ ID NO:3 of ID NO:2.
11. wherein the excitability anti-CD 40 antibodies are with ATCC guarantor for the Liquid pharmaceutical preparation used according to claim 10 The antibody of hiding PTA-3605 or the antibody with INN Selick Shandong monoclonal antibody (selicrelumab).
12. for the Liquid pharmaceutical preparation used according to any one of claim 1 to 11, it is characterised in that the use is treatment cancer Disease, specifically solid tumor, more specific is the gland cancer of colon and rectum, non-small cell lung cancer, the squamous cell carcinoma of head and neck.
13. for the Liquid pharmaceutical preparation used according to any one of claim 1 to 11, it is characterised in that the use is that treatment passes It catches an illness, especially hepatitis type B virus (HBV), Hepatitis C Virus (HCV) and/or human immunodeficiency virus (HIV).
14. a kind of injection device is used to apply the Liquid pharmaceutical preparation for using according to any one of claim 1 to 13.
15. a kind of kit, it includes one or more liquid dosages equipped with for using according to any one of claim 1 to 13 The phial of agent and for by the preparaton subcutaneous administration in the injection device according to claim 14 of patient.
16. the Liquid pharmaceutical preparation according to any one of claim 1 to 11 is used to treat the patient's for suffering from cancer or infectious disease Purposes, it is characterised in that the Liquid pharmaceutical preparation subcutaneous administration.
17. being used to manufacture the drug for the treatment of cancer or infectious disease according to the Liquid pharmaceutical preparation of any one of claim 1 to 11 Purposes, it is characterised in that the Liquid pharmaceutical preparation subcutaneous administration.
18. the purposes of the Liquid pharmaceutical preparation according to claim 17, it is characterised in that carry out skin using the Liquid pharmaceutical preparation The immune agonist of lower application 14,15,16,17 or 18mg dosage, preferably with the antibody of INN Selick Shandong monoclonal antibody.
19. being used to manufacture the purposes of the drug for the treatment of cancer according to the Liquid pharmaceutical preparation of any one of claim 17 or 18.
20. further including according to the purposes of claim 19 comprising selected from Aunar pearl monoclonal antibody (atezolizumab) or shellfish Cut down the drug of the antibody of pearl monoclonal antibody (bevacizumab) as second composition, for combining together with the monoclonal antibody of Selick Shandong, simultaneously Or sequence is applied.
Invention field
The present invention relates to the fields of parenteral antibody formulations, especially play the antibody or antibody piece of immune agonist effect Subcutaneous (sc) pharmaceutical formulations of section.
Background of invention
Immunomodulatory antibody, which provides, a kind of treat method and can be used for directly reinforcing anti-tumor immune response or be used as anti- The adjuvant (Melero, I et al., Nat Rev Cancer 2007,7:95-106) of cancer vaccine.Immune agonist, especially It is that excitability anti-CD 40 antibodies constitute one of most effective classification of these reagents.CD40 is at antigen presenting cell (APC), such as One cell surface member of the tumor necrosis factor superfamily expressed on dendritic cells, B cell and macrophage.Anti- CD40 swashs The preclinical study of dynamic agent prompts alternative under normal circumstances with cross-linking antibody triggering CD40 on antigen presenting cell (APC) Via CD40 Ligand provide cd4 t cell auxiliary, and push CD8 effector T cell activation and expansion (Li, F et al., Science 2011,333:1030-1034).In addition, the macrophage of CD40 activation, which can also play, directly kills tumour function (Beatty,GL et al.,Science 2011,331:1612-1616；Vonderheide,RH et al., Oncoimmunology 2013,2:e23033).CD40 agonist is disclosed in WO2003/040170.
The conventional immune agonist of intravenous (iv) application is usually related with bad adverse events or tolerance overview, can Cause the therapeutic efficiency of the immune agonist lower than best or even interruption treatment (see, for example, Vonderheide, RH et al.,Clin Cancer Res 2013,19(5):1035-1043).Subcutaneous (sc) administration of antibodies can be used to overcome to vein Some consequences that interior antibody preparaton is seen.Know some subcutaneous preparaton (Lundin, J the et al., Blood of antibody 2002,2001,100(3):768-773；Davies, A et al., Lancet Oncol 2014,15 (3): 343-352), have Some commercial prods can obtain.However these most of preparatons include antagonistic antibodies, and usually need specific figuration Desired effect is realized in agent, such as hyaluronidase.New there is still a need for search, alternative exempts from suitable for subcutaneous administration The Pharmaceutical formulation of epidemic disease agonist.
The pharmaceutical compositions of specific CD40 agonist disclosed herein are disclosed in such as WO2003/040170.
Summary of the invention
The present invention includes the Liquid pharmaceutical preparation comprising immune agonist for treating the patient for suffering from cancer or infectious disease Purposes, wherein the Liquid pharmaceutical preparation subcutaneous administration and the wherein subcutaneous administration provide other stomaches with the same preparation Parenteral administration route compares improved tolerance and at least equal effect.
The present invention further comprises purposes described above, and wherein the immune agonist is to specifically bind and activate people The antibody of CD40 or its antigen-binding portion thereof.
The present invention further comprises for the liquid medicine length of schooling comprising immune agonist used in treating cancer or infectious disease Agent, it is characterised in that its subcutaneous administration.
The present invention further comprises a kind of injection device, and it includes Liquid pharmaceutical preparatons as defined herein.
The present invention further comprises a kind of kit, and it includes the phials of the Liquid pharmaceutical preparation, optionally with for according to According to subcutaneous administration of the present invention in patient injection device together.
The present invention further comprises that a kind of treat suffers from infectious disease or proliferative diseases, the method for the patient of such as cancer, The method includes the Liquid pharmaceutical preparation that subcutaneous administration includes immunoregulation agent, especially anti-CD 40 antibodies.
Brief description
Fig. 1: according to intravenous and subcutaneous administration PK data (C of the invention in monkeymax) comparison.
Fig. 2: subcutaneous administration maintains at least identical biological activity compared with intravenous application, shows themselves in that (a) and vein Interior application is compared to the B cell activation after subcutaneous administration；
(b) t cell activation compared with intravenous application after subcutaneous administration.
Fig. 3: influence of the Selick Shandong monoclonal antibody dosage to the activation of the T cell of expression CD69.
Sequence table
SEQ ID NO 1: people CD40.
SEQ ID NO 2: according to the light-chain variable domain (VL) of CD40 antibody of the invention.
SEQ ID NO 3: according to the heavy chain variable domain (VH) of CD40 antibody of the invention.
Detailed description of the invention
With for the activity molecule on immunocyte, such as CD3 (Topp, MS et al., Lancet Oncol 2015,16(1):57-66),CD28(Suntharalingam,G et al.,N Engl J Med 2006,355(10): The monoclonal antibody of exciting function 1018-28) can lead to cytokine release after intravenous application, this may limit agent Amount.
Via it in antigen presenting cell (APC), including bone-marrow-derived lymphocyte, dendritic cells (DC), and the table on monocyte It reaches, CD40, a member of Tumor Necrosis Factor Receptors (TNFR) superfamily, is a kind of crucial adjusting of anti-tumor immune response Object is (see, for example, Grewal, IS et al., Ann Rev Immunol 1998,16:111-35；Van Kooten,C et al.,J Leukoc Biol 2000,67:2-17；Or O'Sullivan, B et al., Crit Rev Immunol 2003,23 (12):83-107).The DC up-regulation antigen processing of CD40 stimulation and presentation approach are simultaneously migrated to lymph node to activate Naive T cells. Excitability CD40 antibody shows the function of substitution CD4+ lymphocyte, causes cytotoxic T lymphocyte (CTL) to expand, energy Established lymthoma is (see, for example, Sotomayor, EM et al., Nature Medicine enough in removing mouse model 1999,5(7):780-7；Gladue,RP et al.,Cancer Immunol Immunother 2011,60(7):1009- 17).CD40 agonist triggers immunostimulation by activation host APC, and then driving is for the t cell response of tumour (referring to example Such as Vonderheide, RH, Clin Cancer Res 2007,13:1083-8).
The inventors found that being previously used for a kind of instant pharmaceutical formulations intravenously applied can not repair Subcutaneous administration is used in the case where changing.Subcutaneous use of the preparation is at least tieed up in the entire biological activity that agonist is immunized Cause in patients while holding or even improve improved tolerance and/or adverse events overview (NCT02665416, NCT02304393).The maintenance of the entire biological activity of the immune agonist comes for example, by the ability of immune cell activated Prove, this cause to be proliferated and activate and (be based respectively on Ki67 in these cells, such as cd8 t cell and CD69 expression measurement) and/ Or these cells (such as CD20 or CD19 expressivity B cell) migrate other compartments into body from peripheral blood.And according to Of the invention subcutaneously uses the immune agonist for leading to that higher doses is applied in a treatment cycle, therefore causes each The higher effect for the treatment of cycle and improved tolerance.In another aspect of the invention, so, inventor also identifies and is used for According to the specific dosage or dosage range of present invention application CD40 antibody.
In one embodiment, therefore, the present invention includes being applied to the trouble for suffering from cancer or infectious disease for subcutaneous (sc) Liquid pharmaceutical preparation comprising immune agonist and pharmaceutically acceptable carrier used in person, wherein use be characterized in that and Other parenteral administration paths of the identical immune agonist in identical pharmaceutically acceptable carrier, especially intravenous (iv) Injection is compared to improvement adverse events overview.Subcutaneous administration includes that the Liquid pharmaceutical preparation of the immune agonist maintains at least Identical or substantive identical biological activity, just as the Liquid pharmaceutical comprising the immune agonist can be injected intravenously Preparation is such.In certain embodiments of the invention, the biological activity of the immune agonist is according to of the invention subcutaneous Even improve compared with intravenously applying identical pharmaceutical formulations after application.
As used in this article, the receptor in " immune agonist " combination cell and starting and the native ligand by this receptor The similar or identical reaction or activity started.
As used in this article, " CD40 agonist " includes any or all but is not limited to following response: B cell proliferation And/or differentiation；Via such as ICAM-1, e-selectin, VC AM waits molecules to raise intercellular adhesion；Secrete pro-inflammatory cellular The factor, such as IL-1, IL-6, IL-8, IL-12, TNF, etc.；By such as TRAF (such as TRAF2 and/or TRAF3), MAP swashs Enzyme, such as NIK (NF-kB inductivity kinases), I- Kappa B kinases (IKK/ beta), transcription factor NF-kB, Ras and MEK/ERK Approach, PI3K AKT approach, P38MAPK approach, etc. approach the signal transduction via CD40 receptor；By such as XIAP, Mcl-1, bcl-x, the transduction of equimolecular anti-apoptotic signal；B and/or T cell memory generate；B cell antibody generates；B cell MHC II class and the cell surface expression of CD80/86 are raised in isotype conversion, etc..
Agonist activity means using identical reading, than by the mean value effect of irrelevant isotype control antibody induction up to The agonist activity of few 3 standard deviations is (thin by such as cell Proliferation or induction costimulation molecule such as CD40 expressivity The parameters such as CD80 or CD86 on born of the same parents' bone-marrow-derived lymphocyte are read).
As used in this article, " CD40 agonist " (or " CD40 antibody ") includes any excitement CD40/CD40L phase interaction Module.Typically, these modules can be excitability CD40 antibody or excitability CD40L polypeptide.For example, these antibody Human antibody including specific agonism CD40/CD40L binding interactions, chimeric antibody, humanized antibody, bispecific are anti- Body, scFv, and antibody fragment.In one embodiment, excitability CD40 antibody can be comprising chimeric, complete people or humanization CD40 antibody.In a further preferred embodiment, excitability CD40 antibody can include complete people CD40 antibody.
In another embodiment, the piece that agonist is CD40 agonist or CD40 agonist is immunized as mentioned above Section.It is to belong to Tumor Necrosis Factor Receptors (TNF-R) family by people's CD40 antigen according to CD40 agonist targeting of the invention A kind of 50kDa cell surface glycoprotein (Stamenkovic et al., EMBO J 1989,8:1403-10).It is also referred to " A member of the TNF receptor family 5 ".Alternative names include B cell surface antigen 40, Bp50, CD40L receptor, CDw40, CDW40, MGC9013, p50 or TNFRSF5.It is with such as UniProt registration number P25942 registration.In an embodiment party In case, people's CD40 antigen has the sequence (being shown in Table 1) according to SEQ ID NO:1.
Table 1: the protein sequence of people's CD40 antigen
In one embodiment, people CD40 has the sequence according to SEQ ID NO:1.As used in this article, " in conjunction with People CD40 " or " specific binding people CD40 " or " in conjunction with people CD40's " or " anti-CD 40 antibodies " refer to KDValue is 1.0x 10- 8Mol/l or less, in one embodiment KDValue is 1.0x 10-9Mol/l or the specific binding of less binding affinity The antibody of people's CD40 antigen.With standard binding assay, such as surface plasmon resonance technology (GE- Healthcare Uppsala, Sweden) measurement binding affinity.As used in this article, so, " in conjunction with and activate people The antibody of CD40 " refers to KD 1.0x 10-8Mol/l or less (10 1.0x in one embodiment-8Mol/l to 1.0x 10-13), mol/l K in one embodimentD 1.0x 10-9Mol/l or less (10 1.0x in one embodiment-9mol/l To 1.0x 10-13Mol/l the antibody of binding affinity specific binding people CD40 antigen).In another embodiment, according to According to anti-CD 40 antibodies of the invention with 4x 10-10M or less KDIn conjunction with people CD40.
In one embodiment of the invention, the antibody in conjunction with people CD40 is agonist (" CD40 agonist ").
In one embodiment, the CD40 antibody is the fully human antibodies of IgG2 subclass.In also another implementation In scheme, the antibody is such as any anti-CD 40 antibodies specifically disclosed in WO2003/040170.Still there is another embodiment party In case, it is selected from according to CD40 agonist of the invention and is referred to as 3.1.1,7.1.2,10.8.3 according to WO2003/040170, 15.1.1,21.4.1,21.2.1,22.1.1,23.5.1,23.25.1,23.29.1 the group with the antibody of 24.2.1.Secrete those The hybridoma of antibody is according to budapest treaty preservation.Deposit number can be in the paragraph [0250] of WO2003/040170 It finds.In another embodiment, it include antibody 21.4.1 (ATCC deposit number PTA- according to CD40 antibody of the invention 3605) heavy chain and light chain variable domain amino acid sequence.In yet another embodiment, according to CD40 antibody of the invention It is made of the heavy chain and light-chain amino acid sequence of antibody 21.4.1 (ATCC deposit number PTA-3605).
In one embodiment, it include amino acid sequence SEQ ID according to people's excitability anti-CD 40 antibodies of the invention The light-chain variable domain of NO:2 and the heavy chain variable domain (table 2) of amino acid sequence SEQ ID NO:3.
Table 2: according to light (VL) and the amino acid sequence of (VH) chain variable domain again of CD40 agonist of the invention
In another embodiment, CD40 agonist is with 4x 10-10M or less KDIgG2 in conjunction with people CD40 is sub- The fully human antibodies of class；Or the antibody of the VH of the VL and SEQ ID NO:4 comprising SEQ ID NO:3；Or include antibody 21.4.1 The heavy chain of (ATCC deposit number PTA-3605) and the antibody of light chain variable domain amino acid sequence.
In another embodiment, it is with INN Selick Shandong according to CD40 antibody or CD40 agonist of the invention The antibody of monoclonal antibody.
As used in this article, term " improved tolerance " means other parenteral with same liquid pharmaceutical formulations Administration route, such as intravenous application is compared to less adverse events in patient.Term " adverse events " or " adverse events Overview " means direct or indirect consequence caused by the agonism as the CD40 on cell, non-shows typically but only To discharge cell factor by CD40 expressivity immunocyte.This overview include clinical symptoms (such as feel cold, generate heat, low blood pressure, Deng) and serum parameters for monitoring organ dysfunction, the variation of such as liver functional test and coagulation parameters.
As used in this article, term " subcutaneous administration (sc) " means exciting via subcutaneous tissue application CD40 is injected into Agent pushes absorption (such as hyaluronidase) with or without adding ingredient.Subcutaneous administration includes splitting dosage and in same administration It is applied to the option of multiple regions of anatomy day, but also including both single and repeat administration (interval 3-4 weeks).
As used in this article, " Liquid pharmaceutical preparation " has the phase mixing in the form of aqueous solution or freeze-dried formulation The immune agonist of prestige degree and one or more optional " pharmaceutically acceptable carrier " (Remington's Pharmaceutical Sciences 16th edition, Osol, A.Ed. (1980)) it prepares.As used in this article, " pharmaceutically acceptable carrier " is usually nontoxic to recipient in used dosage and concentration, and includes but is not limited to: slow Electuary, such as phosphate, citrate, and other organic acids；Antioxidant, including ascorbic acid and methionine；Preservative (such as octadecyl dimethyl benzyl ammonium chloride；Hexamethonium chloride；Benzalkonium chloride；Benzethonium chloride；Phenol, butanol or benzyl alcohol； Alkyl paraben, such as methyl p-hydroxybenzoate or propyl ester；Catechol；Resorcinol；Hexamethylene；3- penta Alcohol；And metacresol)；Low molecular weight (less than about 10 residues) polypeptide；Protein, such as serum albumin, gelatin, or immune ball Albumen；Hydrophilic polymer, such as polyvinylpyrrolidone；Amino acid, such as glycine, glutamine, asparagine organize ammonia Acid, arginine, or lysine；Monosaccharide, disaccharides, and other carbohydrate, including glucose, mannose, or dextrin；Chelating Agent, such as EDTA；Sugar, such as sucrose, mannitol, trehalose or sorbierite；At salt gegenion, such as sodium；Metal composite (such as Zn- protein complex)；And/or nonionic surface active agent, such as polyethylene glycol (PEG).Illustration herein The pharmaceutically acceptable carrier of property includes interstitial drug dispersing agent again, such as soluble neutral reactive transparent matter acid enzyme glycoprotein Such as human soluble PH-20 hyaluronidase glycoprotein, (sHASEGP), such as rHuPH20 (Baxter International,Inc.).Certain illustrative sHASEGP and application method, including rHuPH20 are described in U.S. Patent Publication Text No.2005/0260186 and 2006/0104968.In one aspect, sHASEGP and one or more other osamines are poly- The combination of sugar, such as chondroitinase.
Illustrative lyophilized antibodies preparaton is described in United States Patent (USP) No.6,267,958.Aqueous antibody preparaton includes the U.S. Described in patent No.6,171,586 and WO2006/044908 those, the latter's preparaton includes histidine-acetate buffer Liquid.
It is usually sterile for being used for the preparaton of subcutaneous administration.Aseptic can realize easily, for example, by across Sterilised membrane filter filtering.
In one embodiment, the present invention provides the CD40 antibody 21.4.1 (ATCC as disclosed in WO2003/040170 Deposit number PTA-3605) as immune agonist, by 20mM sodium acetate, 140mM sodium chloride, 0.02% polyoxyethylene sorbitan monoleate group At with the concentration preparation of 10mg/ml in the buffer solution of pH 5.5.The liquid adjustments provide in 2mL phial, so with The amount of 20mg/ phial contains the CD40 antibody.This preparaton is also disclosed in embodiment 1 herein.
It can change the actual dose level that agonist, especially CD40 agonist are immunized in pharmaceutical compositions of the invention Desired treatment is effectively realized to particular patient, composition, and administration mode in the case where not having toxicity to patient to obtain Property response amount (effective quantity) active component.Selected dosage level can depend on a variety of pharmacokinetics factors, including be used Specific immune agonist of the invention activity, or age of patient treated, gender, weight, situation, general health and First medical history, and similar factor well known to medical domain.In one embodiment, according to CD40 agonist of the invention Dosage be about 1 to 100mg/ml, or about 10 to 80mg/ml, or about 10 to 40mg/ml, or about 10 to 30mg/ml, or about 5 to 55mg/ml.In another embodiment, it is about 5mg/ml, or about 10mg/ according to the dosage of CD40 agonist of the invention Ml, or about 15mg/ml, or about 20mg/ml, or about 25mg/ml, or about 30mg/ml, or about 35mg/ml, or about 40mg/ml, or About 45mg/ml, or about 50mg/ml, or about 55mg/ml.
In another embodiment, to be selected from about 8mg to 48mg, preferably from about 12mg to each treatment cycle of about 32mg Flat dosage；Or to be selected from 14,15,16,17, or the dosage of each treatment cycle of 18mg applies CD40 antibody.
In another embodiment, CD40 antibody is the antibody with INN Selick Shandong monoclonal antibody, and each with 16mg The flat dosage for the treatment of cycle is applied.
It can change between 5.0 and 6.0 according to the pH of composition of the invention.It can be used any physiology acceptable Buffer obtains such pH.In one embodiment, buffer is selected from sodium succinate or sodium acetate, with or without sodium chloride.In In another embodiment, selected buffer is with about 10 to 30mM, and the amount of especially about 20mM exists, and pH is about 5.5.
Surfactant can be contained according to composition of the invention.In one embodiment, surfactant right and wrong from Subtype surfactant.In another embodiment, surfactant is selected from the group of poloxamer or polysorbate.Another In a embodiment, surfactant is poloxamer-188, or Tween-20, or polyoxyethylene sorbitan monoleate.According to the present invention, Surfactant exists with the concentration of about 0.02 to 0.1% (w/v).In one embodiment, surfactant is with 0.02, 0.03,0.04,0.05,0.06,0.07,0.08,0.09 or the concentration of 0.1% (w/v) exists.
In yet another embodiment of the invention, it can optionally contain cryoprotector according to composition of the invention And/or antioxidant.In one embodiment, cryoprotector is sugar, and especially sucrose and antioxidant are methionines. Cryoprotector is with about 180-300nM, or the amount of about 240nM exists.Antioxidant is with about 0 to 20mM, or about 0-10mM, or about The amount of 10mM exists.
As used in this article, term " cancer " preferably means that solid tumor, such as lung cancer, non-small cell lung (NSCL) cancer, bronchioloalveolar cell lung cancer, osteocarcinoma, cancer of pancreas, cutaneum carcinoma, head or neck cancer, skin or intraocular melanoma, son Palace cancer, oophoroma, the carcinoma of the rectum, anal region cancer, gastric cancer, the cancer of stomach, colon cancer, breast cancer, uterine cancer, carcinoma of fallopian tube, intrauterine Film cancer, cervical carcinoma, carcinoma of vagina, carcinoma of vulva, Huo Qijin (Hodgkin) family name disease, cancer of the esophagus, carcinoma of small intestine, endocrine system cancers, first Shape gland cancer, parathyroid carcinoma, adrenal, soft tissue sarcoma, carcinoma of urethra, carcinoma of penis, prostate cancer, bladder cancer, kidney or urine output Pipe cancer, clear-cell carcinoma, carcinoma of renal pelvis, celiothelioma, hepatocellular carcinoma, cholangiocarcinoma, central nervous system (CNS) neoplasm, spinal cord axis are swollen Tumor, brain stem glioma, glioblastoma multiforme, astrocytoma, schwannoma, ependymoma, medulloblast Tumor, meningioma, squamous cell carcinoma, pituitary adenoma, lymthoma, lymphocytic leukemia, including any of above of obstinate pattern Cancer, or the combination of one or more above-mentioned cancers.In a preferred embodiment, such cancer is breast cancer, and colon is straight Intestinal cancer, melanoma, head and neck cancer, lung cancer or prostate cancer.In one embodiment, such cancer is selected from breast cancer, lung Cancer, colon cancer, oophoroma, melanoma, bladder cancer, the cancer of kidney, kidney, liver cancer, head and neck cancer, colorectal cancer, cancer of pancreas, stomach The solid tumor of cancer, cancer of the esophagus, celiothelioma or prostate cancer.In another embodiment, such cancer is haematological tumours, all Such as such as leukaemia (such as AML, CLL), lymthoma, myeloma.In still having another embodiment, cancer is breast cancer, Lung cancer, colon cancer, colorectal cancer, cancer of pancreas, gastric cancer or prostate cancer.
As used in this article, term " infectious disease " means by such as bacterium, virus, the organisms such as fungi or parasitic animal and plant Caused illness.The specific example of infectious disease including but not limited to hepatitis type B virus (HBV), Hepatitis C Virus (HCV) and/ Or human immunodeficiency virus (HIV).
In one embodiment, according to subcutaneous administration of the invention for preventing or treating transfer.
In one embodiment, according to subcutaneous administration of the invention for treating immune correlated disease, such as tumour is exempted from Epidemic disease postpones its progress.
In one embodiment, according to subcutaneous administration of the invention for stimulating immune response or function, such as T cell Activity.
As used in this article, term " variable domain " (light-chain variable domain VL, heavy chain variable domain VH) expression directly involves anti- Each of a pair of light and heavy domain of body combination antigen.It can lighten and heavy chain domain general structure having the same and every A domain includes four frame (FR) areas, and sequence is guarded extensively, is connected by three " hypervariable regions " (or complementary determining region, CDR).Frame Frame area takes β-sheet conformation and CDR can form connection β-lamellar structure ring.CDR in every chain keeps it by framework region Three-dimensional structure and be formed together antigen binding site with the CDR from another chain.The weight of antibody and the area light chain CDR3 exist It is of the invention another according to playing a particularly important role in binding specificity/affinity of antibody of the invention and therefore providing A purpose.
As used herein, term " antigen-binding portion thereof of antibody " refers to the amino acid for being responsible for antigen binding in antibody Residue.The antigen-binding portion thereof of antibody includes the amino acid residue from " complementary determining region " or " CDR "." frame " or " FR " area It is the variable domain region those of other than some hypervariable region residues as defined herein.Therefore, the light and heavy chain variable domain of antibody It include domain FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4 from N to C-terminal.Especially, the CDR3 of heavy chain is to antigen binding Contribution is maximum and limits the region of the characteristic of antibody.The area CDR and FR is according to Kabat et al., Sequences of Proteins of Immunological Interest,5th ed.,Public Health Service,National The standard of Institutes of Health, Bethesda, MD (1991) define to determine and/or be those from " hypervariable loop " Residue.
In one embodiment of the invention, specifically bind and activate " antigen-binding portion thereof " of the antibody of people CD40 CDR1, CDR2 and the CDR3 of weight and light-chain variable domain comprising antibody 21.4.1 (ATCC deposit number PTA-3605).
As used in this article, term " nucleic acid " or " nucleic acid molecules " are intended to include DNA molecular and RNA molecule.Nucleic acid point Son can be it is single-stranded or double-strand, it is preferred that being double-stranded DNA.
As used in the application, term " amino acid " indicates the natural group that carboxyl a-amino acid occurs, and includes alanine (three-letter codes: ala, single letter code: A), arginine (arg, R), asparagine (asn, N), aspartic acid (asp, D), Cysteine (cys, C), glutamine (gln, Q), glutamic acid (glu, E), glycine (gly, G), histidine (his, H) are different Leucine (ile, I), leucine (leu, L), lysine (lys, K), methionine (met, M), phenylalanine (phe, F), dried meat Propylhomoserin (pro, P), serine (ser, S), threonine (thr, T), tryptophan (trp, W), tyrosine (tyr, Y), and valine (val,V)。
" part Fc " of antibody does not involve antibodies bind antigen directly, but shows various effector functions." antibody The part Fc " is the term of the known and papain cutting definition based on antibody of those of skill in the art.According to their heavy chain Constant region amino acid sequence, antibody or immunoglobulin be divided into class: IgA, IgD, IgE, IgG and IgM, and in these It is several to be further separated into subclass (isotype), such as IgG1, IgG2, IgG3, and IgG4, IgA1, and IgA2.According to heavy chain perseverance Determine area, inhomogeneous immunoglobulin is referred to as α, δ, ε, γ, and μ.The part Fc of antibody directly involves ADCC (antibody-dependant Property cell-mediated cytotoxicity) and CDC (complement-dependent cytotoxicity), be based on complement activation, C1q is combined and Fc receptor In conjunction with.Complement activation (CDC) is started by complement factor C1q in conjunction with the part Fc of most of IgG antibody subclass.Although antibody Influence to complement system depends on certain conditions, but is caused by the binding site that limits in the part Fc in conjunction with C1q.This Class binding site is known in the art and is described in such as Boackle, R.J.et al., Nature (1979) 282:742- 743；Lukas,T.J.et al.,J.Immunol.(1981)127:2555-2560；Brunhouse,R.and Cebra, J.J.,Mol.Immunol.(1979)16:907-917；Burton,D.R.et al.,Nature(1980)288:338-344； Thommesen,J.E.et al.,Mol.Immunol.(2000)37:995-1004；Idusogie,E.E.et al., J.Immunol.(2000)164:4178-4184；Hezareh,M.et al.,J.Virology(2001)75:12161- 12168；Morgan,A.et al.,Immunology(1995)86:319-324；EP 0 307 434.Such binding site is Such as (numbering is seen below according to Kabat, E.A. by L234, L235, D270, N297, E318, K320, K322, P331 and P329 The EU index of text).The antibody of subclass IgG1, IgG2 and IgG3 usually show that complement activation and C1q and C3 are combined, and IgG4 does not live Change complement system and does not combine C1q and C3.
In one embodiment, according to immune agonist of the invention, preferably CD40 agonist is monoclonal antibody.In In another embodiment, according to immune agonist of the invention, preferably CD40 agonist be human IgG class (i.e. IgG1, or IgG2, or IgG3 or IgG4 subclass).
In one embodiment, immune agonist described herein, preferably CD40 agonist are characterized in that constant chain It is that people originates from.Such constant chain is known in the art and by such as Kabat, E.A. description (see, for example, Johnson, G.and Wu,T.T.,Nucleic Acids Res.(2000)28:214-218)。
Immune agonist described herein, preferably CD40 agonist are preferably generated by recombinant means.Such method is The prior art is widely known and is included in protokaryon and eukaryocyte and expresses protein and be subsequently isolated antibody polypeptides and usually It is purified to pharmaceutically acceptable purity.For protein expression, by standard method by encoded light and heavy chain or the nucleic acid of its segment It is inserted into expression vector.In suitable protokaryon or eukaryotic host cell, such as Chinese hamster ovary celI, NS0 cell, SP2/0 cell, HEK293 Cell, COS cell are implemented expression in yeast, or Bacillus coli cells, and are returned from cell (from supernatant or after cell lysis) Receive antibody.
The recombination generation of antibody is known in the art and is described in such as review article Makrides, S.C., Protein Expr.Purif.(1999)17:183-202；Geisse,S.et al.,Protein Expr.Purif.(1996) 8:271-282；Kaufman,R.J.,Mol.Biotechnol.(2000)16:151-161；Werner,R.G.,Drug Res. (1998)48:870-880。
Antibody can be in intact cell, in cell lysates, or with partially purified, or substantive pure form is deposited In.In order to remove other cell components or other pollutants, for example, other cellular nucleic acids or protein and implement to purify, pass through Standard technique carries out, including alkali/SDS processing, CsCl are chromatographed at band, column, agarose gel electrophoresis, and well known in the art other Technology.Referring to Ausubel, F.et al., ed.Current Protocols in Molecular Biology, Greene Publishing and Wiley Interscience,New York(1987)。
Expression in NS0 cell is described in such as Barnes, L.M.et al., Cytotechnology (2000) 32: 109-123；Barnes,L.M.et al.,Biotech.Bioeng.(2001)73:261-270.Transient expression is described in for example Durocher,Y.et al.,Nucl.Acids.Res.(2002)30:E9.The clone of variable domain is described in Orlandi, R.et al.,Proc.Natl.Acad.Sci.USA(1989)86:3833-3837；Carter,P.et al., Proc.Natl.Acad.Sci.USA(1992)89:4285-4289；Norderhaug,L.et al., J.Immunol.Methods(1997)204:77-87.A kind of preferred transient expression system (HEK 293) is described in Schlaeger, E.-J.and Christensen, K., in Cytotechnology (1999) 30:71-83 and Schlaeger, E.-J.,in J.Immunol.Methods(1996)194:191-199。
It will be according to weight and light-chain variable domain of the invention and promoter, translation initiation, constant region, 3' non-translational region, poly- gland Nucleotide, and the combined sequence of tanscription termination is to form expression vector establishment object.Weight and light chain expression constructs can be combined At single carrier, cotransfection is continuous to transfect, or is separately transfected into host cell, merges host cell then to be formed and expresses this The single host cell of two chains.
Control sequence suitable for prokaryotes includes such as promoter, optional operator sequence, and ribosomes Binding site.Known eukaryocyte utilizes promoter, enhancer and polyadenylation signal.
When nucleic acid is positioned to be in functional relationship with another nucleic acid sequence, it is " being operatively connected ".Such as such as Fruit presequence (presequence) secretes the preceding protein that leading DNA is expressed as the secretion of participation polypeptide (preprotein), then the DNA of it and polypeptide is operatively connected；If promoter or enhancer influence turning for coded sequence Record, then it is operatively connected with the sequence；Or if the positioning of ribosome bind site pushes translation, then it and coding Sequence is operatively connected.Usually, " being operatively connected " means that connected DNA sequence dna adjoins, and leading secreting In the case where adjoin and with frame.However enhancer needs not be and adjoins.Connection passes through the company at convenient restriction site Fetch realization.If there is no such site, then using synthetic oligonucleotide adapter or connector according to conventional practice.
It purifies regulation by conventional immune globulins appropriately to separate monoclonal antibody with culture solution, such as albumin A- Sepharose, hydroxyapatite, gel electrophoresis, dialysis, or affinity chromatography.The DNA and RNA of coding monoclonal antibody are easy to It is separated and is sequenced using routine protocols.Hybridoma may act as the source of such DNA and RNA.Once separation, so that it may to incite somebody to action DNA is inserted into expression vector, then expression vector is transfected into the host for not generating immunoglobulin protein in other cases Cell, such as HEK293 cell, Chinese hamster ovary celI, or myeloma cell, to obtain the conjunction of recombinant monoclonal antibodies in host cell At.
As used in this article, " cell " is stated, " cell line ", and " cell culture " are used interchangeably and all such Title includes offspring.So, word " transformant " and " inverted cell " include primary theme cell and culture from derived from it Object, regardless of the number of passage.It is further appreciated that, due to intentional or unintentional mutation, all offsprings can not in terms of DNA content It is accurate identical.Including have with after the variant of the identical function or biological activity screened in initial conversion cell Generation.
Method for generating the specific CD40 agonistic antibody used according to the present invention is also disclosed in WO2003/ 040170。
The present invention further comprises a kind of method for treating the patient with cancer, it is characterised in that according to the present invention Immune agonist, preferably the Liquid pharmaceutical preparation of CD40 agonist are included to patient's subcutaneous administration therapeutically effective amount.
The present invention further comprises a kind of kit, and it includes one or more to match equipped with any liquid disclosed herein The phial of preparation and for by preparaton subcutaneous administration in the injection device of patient.In this embodiment, according to implementation The liquid adjustments of example 1 are preferred.
The present invention further comprise for another therapeutic agent, preferably anticancer agent combine, simultaneously or sequentially subcutaneously (s.c.) pharmaceutical formulations and dosage of CD40 antibody or agonist as defined herein are applied.In one embodiment, institute It states other therapeutic agent and is selected from the antibody with INN Aunar pearl monoclonal antibody or Avastin.Can according to well known to technical staff and Such as such as productWithProduct information described in method apply these antibody.
The present invention can thus be summarized by following more specifical embodiment, wherein all variables and term have The range and/or definition previously herein provided:
1. for the Liquid pharmaceutical preparation comprising immune agonist used in treating cancer or infectious disease, wherein the liquid Pharmaceutical formulations subcutaneous administration.
2. for the Liquid pharmaceutical preparation used according to claim 1, it is characterised in that it includes aqueous solution, the aqueous solution packet Contain:
(a) agonist is about immunized described in 1-100mg/ml；
(b) about 10-30mM is at least one provides the buffer of pH 5 to 6；With
(c) about 0.01 to 0.1% (w/v) nonionic surface active agent.
3. for the Liquid pharmaceutical preparation used according to claims 1 or 2, it is characterised in that it includes aqueous solution, this is water-soluble Liquid includes:
(a) agonist is about immunized described in 10-80mg/ml；
(b) about 20mM is at least one provides the buffer of pH about 5.5；With
(c) about 0.02% (w/v) nonionic surface active agent.
4. further including about 180-300mM for the Liquid pharmaceutical preparation used according to any one of claims 1 to 3 Sucrose and about 0-20mM methionine.
5. further including about 240mM sucrose and about 0-10mM for the Liquid pharmaceutical preparation used according to claim 4 Methionine.
6. for the Liquid pharmaceutical preparation used according to any one of claim 2 to 5, wherein the buffer be acetate or Succinate buffers, such as sodium acetate or sodium succinate or combinations thereof optionally further include about 140mM sodium chloride.
7. for the Liquid pharmaceutical preparation used according to any one of claim 2 to 6, wherein the non-ionic surfactant Agent is PLURONICS F87 or polysorbate 20 or polyoxyethylene sorbitan monoleate.
8. wherein the immune agonist is excitability for the Liquid pharmaceutical preparation used according to any one of claim 1 to 7 CD40 antibody.
9. for the Liquid pharmaceutical preparation used according to any one of claim 1 to 8, it is characterised in that it is containing about 10mg/ml excitability anti-CD 40 antibodies；About 20mM sodium succinate and about 0.02% (w/v) are selected from polysorbate 20, polysorbate 80 or PLURONICS F87 surfactant, the aqueous solution in about 5.5 pH.
10. wherein the immune agonist is to include for the Liquid pharmaceutical preparation used according to any one of claim 1 to 9 The excitability anti-CD 40 antibodies of the heavy chain variable domain (VH) of the light-chain variable domain (VL) and SEQ ID NO:3 of SEQ ID NO:2.
11. wherein the excitability anti-CD 40 antibodies are that have for the Liquid pharmaceutical preparation used according to claim 10 The antibody of ATCC deposit number PTA-3605 or antibody with INN Selick Shandong monoclonal antibody.
12. for the Liquid pharmaceutical preparation used according to any one of claim 1 to 11, it is characterised in that the use comprising Treating cancer, specifically solid tumor, more specific is the gland cancer of colon and rectum, non-small cell lung cancer, and the squamous of head and neck is thin Born of the same parents' cancer.
13. for the Liquid pharmaceutical preparation used according to any one of claim 1 to 11, it is characterised in that the use comprising Treat infectious disease, especially hepatitis type B virus (HBV), Hepatitis C Virus (HCV) and/or human immunodeficiency virus (HIV).
14. a kind of injection device is used to apply the liquid medicine length of schooling for using according to any one of claim 1 to 13 Agent.
15. a kind of kit, it includes one or more liquid equipped with for using according to any one of claim 1 to 13 The phial of preparaton and for by the preparaton subcutaneous administration in the injection device according to claim 14 of patient.
16. being used to treat the trouble for suffering from cancer or infectious disease according to the Liquid pharmaceutical preparation of any one of claim 1 to 11 The purposes of person, it is characterised in that the Liquid pharmaceutical preparation subcutaneous administration.
17. being used to manufacture the medicine for the treatment of cancer or infectious disease according to the Liquid pharmaceutical preparation of any one of claim 1 to 11 The purposes of object, it is characterised in that the Liquid pharmaceutical preparation subcutaneous administration.
18. the purposes of the Liquid pharmaceutical preparation according to claim 17, it is characterised in that use the Liquid pharmaceutical preparation Carry out the immune agonist of subcutaneous administration 14,15,16,17 or 18mg dosage, it is preferably anti-with INN Selick Shandong monoclonal antibody Body.
19. being used to manufacture the use of the drug for the treatment of cancer according to the Liquid pharmaceutical preparation of any one of claim 17 or 18 On the way.
20. further including includes selected from Aunar pearl monoclonal antibody or Avastin according to the purposes of claim 19 The drug of antibody is administered simultaneously or sequentially as second composition for combining together with the monoclonal antibody of Selick Shandong.
Embodiment
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