6-9 장, 11 장. 재조합단백질 바이오의약품의실제 단백질의약품 미생물면역학전공 (youhee@cha.ac.kr)
3 의약품품목별매출순위 순위품목 2011 2010 % 순위품목 2011 2010 % 1 Lipitor 12.5 12.7-3.3 11 Singulair 6.1 5.5 10.3 2 Plavix 9.3 8.8 3.7 12 Zyprexa 5.7 5.7-3.1 3 Seretide 8.7 8.6 0.04 13 Rituxan 5.7 5.1 8.6 4 Crestor 8.0 6.8 14.4 14 Lantus 5.5 4.7 15.2 5 Nexium 79 7.9 84 8.4-6.2 62 15 Avastin 54 5.4 56 5.6-7.5 75 6 Seroquel 7.6 6.8 9.5 16 Herceptin 4.8 4.3 7.9 7 Humira 73 7.3 60 6.0 17.8 17 Cymbalta 47 4.7 39 3.9 19.2 8 Enbrel 6.8 6.2 6.7 18 Spiriva 4.7 4.0 13.6 9 Remicade 6.8 6.1 8.4 19 Neulasta 4.2 3.8 9.4 10 Abilify 6.3 5.4 14.3 20 Glivec 4.1 3.8 6.5 ( 단위: 10 억달러 ) 바이오의약품의현황
4 공부할내용 중요의약품명상품명단백질기능적응증 Etanercept Enbrel TNFR/IgG1(Fc) 융합단백질류마티스관절염 Insulin glargine Lantus 인슐린 ( 지속성 ) 당뇨병 EPO Epogen/Procrit EPO 빈혈 Pegfilgrastim Neulasta Pegylated G CSF 호중구감소증 Interferon beta 1A Avonex/Rebif IFN β 다발성경화증 (MS) Botulinum toxin (BoNT) Botox/Dysport 보툴리눔독소사시,. 경련, 주름개선 Coagulation Factor VIII Advate/Kogenate Factor VIII 혈우병 Interferon alpha 2A Pegasys IFN α B 형,C 형간염, 간경화 Insulin aspart NovoLog/NovoRapid 인슐린 ( 속효성 ) 당뇨병 Darbepoetin Aranesp/NESP EPO 빈혈 Insulin lispro Humalog 인슐린 ( 속효성 ) 당뇨병 암기자료
5 공부할내용 중요의약품명상품명단백질기능적응증 Epoetin beta rch NeoRecormon EPO (CHO 생산 ) 빈혈 Insulin detemir Levemir 인슐린 ( 지속성 ) 당뇨병 Coagulation Factor VIIa NovoSeven Factor VIIa 혈우병 Interferon beta 1B Betaferon/Extavia t IFN β 다발성경화증 (MS) Insulin aspart (biphasic) NovoMix 인슐린 ( 속효성 ) 당뇨병 Filgrastim Neupogen G CSF 호중구감소증 Insulin Humulin 인슐린 ( 표준 ) 당뇨병 Denileukin diftitox Ontak IL 2/Diphtheria toxin 융합단백질피부 T 세포림프종 Abatacept Orencia CTLA 4/IgG 융합단백질류마티스관절염 Lutropin alpha Luveris LH 불임 (IVF) Desirudin Iprivask 히루딘혈전, 심근경색 Interferon alpha 2B Intron A IFN α B 형,C 형간염 Anakinra Kineret IL 1R 길항제 (IL 1 활성저해 ) 류마티스관절염 Rasburicase Elitek Urate oxidase 통풍, 종양용해증후군 암기자료
6 공부할내용 중요의약품명상품명단백질기능적응증 Nepidermin Regen D EGF 피부재생, 창상 Agalsidase alpha Replagel Alpha galactosidase 파브리병 Idursulfase Elaprase ($375,000/yr) iduronate 2 sulfatase 헌터증후군 Mecasermin Increlex IGF 1 유전적성장장애 (IGF 1 결핍 ) Follitropin alpha Gonal F FSH 불임 ( 과배란유도 ) hgh Protropin GH 성장호르몬 재조합단백질의약품의종류재조합싸이토카인 : IFN α, IFN β / 저해제 : Etanercept, Abatacept, Anakinra 재조합저해제 / 길항제 : IL2 DT, BoNT, Factors VIII & VII, DTIs, Enfuvirtide 재조합호르몬 : Insulins, hgh, calcitonin 재조합성장인자 : G CSF, GM CSF, EPO, TPO, EGF, FGF, IGF 1, PDGF, VEGF 재조합효소 : UO, ADA, Iduronate sulfatase, galactosidase, tpa, asparaginase 암기자료
7 Cytokines 미생물학 2. 면역반응의조절 : 싸이토카인 ( 선천면역 )
8 Cytokines 미생물학 2. 면역반응의조절 : 싸이토카인 ( 획득면역 )
9 Cytokines 미생물학 2. 면역반응의조절 : 싸이토카인 ( 조혈 )
10 Immunosuppressants Purine biosynthesis de novo pathway ( 신생경로 ): 분열중인세포에서활발 ( 억제타겟 ) salvage pathway ( 회피경로 ) Fig. 40.1 책에서다루는면역억제제 면역억제제의분류 ( 분자세포약물학 )
11 Steps in T cell activation Signal 1 Signal 2 Signal 3 미생물학 2. 면역반응의조절 : TCR, T- 세포의분화, 증식, 활성화
12 Interleukin 2 (IL-2) secreted by Th cells stimulates T cell proliferatioin and differentiation stimulates NK cells as well as plasma B cells : impacts on both CMI and humoral immunity and on tolerance as well thus, the one of the key soluble regulators in immune responses binds to IL 2R (cf. IL 2Rγ = CD25) activates mtor and CDK pathways cell proliferation cf. Clinical application of IL 2 Immune activator per se (Proleukin) The major target for immunosuppression Zenapax, Simulect (transplantation); Ontak (CTCL) Cyclosporin, FK 506 주요싸이토카인
13 Interferons (IFN-α, IFN-β) secreted by various cells including virus infected fibroblasts stimulates T and NK cells proliferatioin and differentiation cf. increases MHC I and IFN γ expression : impacts on anti viral responses (mostly, CMI) thus, the one of the key soluble regulators in CMI and antiviral immunity binds to IFNAR (cf. IL 2Rγ = CD25) activates JAK1/TYK2 pathway (and STAT2) cf. INF γ 주요싸이토카인
14 Tumor necrosis factor (TNF-α) aka. cachectin/cachexin in acute phase reaction secreted by T cells, NK cells, and macrophages induces fever, apoptotic cell death, cachexia, and inflammation : inhibits tumorigenesis and viral replication implicated in diseases due to its dysregulation (Alzheimer s disease, IBD etc) thus, the one of the key cytokines in (acute) immune responses binds to TNFR (= death receptor) activates caspase pathways etc. cell death etc. cf. Clinical application of TNF α Immune/apoptotic activator per se The major target for inflammation 주요싸이토카인
15 Denileukin diftitox Ontak an antineoplatic agent combining IL 2 anddt binds to IL 2R, introducing DT into IL 2R expressing cells To treat some leukemias and lymphomas with malignant cells expressing IL 2R FDA apporved (1999) for CTCL such as Mycosis Fungoides ( 균상식육종 ) treatment cf. CTCL 환자의 60% 가 IL 2R 을발현 Patients 3 (a, b) and 4 (c, d) before therapy and after 2 cycles of denileukin diftitox plus total skin electron beam radiation. 재조합단백질의약품 : 싸이토카인 / 독소융합단백질
16 IFN-α Pegasys (IFN α2a), Intron A/Peg intron (IFN α2b) Interferon therapy (in combination with chemotherapy) to treat cancers and viral infections (including HIV coinfection, cirrhosis) Both HBV and HCV are treated (over age of 18), highly effective toward HCV genotype I (60 80% success rate) by ribavirin combination. Sometimes used for treatment of HPV (condyloma) 재조합싸이토카인
17 IFN-β Avonex/Rebif (IFN β1a), Betaferon/Extavia (IFN β1b) Avonex was apporved in 1996 (USA) and in 1997 (EU) and is the leading MS (multiple sclerosis) therapy in the US. www.avonex.com 재조합싸이토카인
18 Etanercept Enbrel a recombinant protein composed of human TNFR2 and the end (CH2 and CH3) of human IgG1 Enbrel, Humira, Remicade has been approved to treat RA, and now used to treat autoimmune diseases as well. 재조합저해제 / 길항제
19 Abatacept Orencia a fusion protein of the extracellular domain of CTLA 4 and the Fc region of IgG1 binds to and sequester B7 (the ligand for CD28 and CTLA 4) with increased affinity (~30X), resulting in the loss of T cell co stimulation to treat RA (rheumatoid arthritis), in case of inadequate response to anti TNF α therapy 재조합저해제 / 길항제
20 Anakinra Kineret a recombinant, non glycosylated version of human IL 1R agonist prepared from Escherichia coli. to treat RA (rheumatoid arthritis) may cause neutropenia 재조합저해제 / 길항제
21 Botulinum toxin BoNT/A (Botox, Dysport and Xeomin) and BoNT/B (MyoBloc) Botulinum toxin (BoNT) is a neurotoxin produced by Clostridium botulinum the most acutely lethal toxin known: LD 50 of 1.3~2.1 ng/kg (iv/im) 10~13 ng/kg (inhaled) cf. the mean concentration in BoNT/A products Botox: 0.73 ng/100 unit (i.e. 137 units/ng) Dysport: 0.65 ng/100 unit (i.e. 154 units/ng) Xeomin: 0.44 ng/100 unit (i.e. 227 units/ng) 재조합저해제 / 길항제
22 Coagulation factors VIII and VII Advate/Kogenate and NovoSeven Factor VIII (aka. anti hemophilic factor). Defects in the F8 gene results in hemophilia A, a recessive X linked coagulation disorder. Factor VII (prothrombin conversion accelerator; proconvertin). used for uncontrolled bleeding in hemophilia patients. It is sometimes used unlicensed in severe uncontrollable bleeding, although there have been safety concerns. A biosimilar (AryoSeven) is also available, but does not play any considerable role in the market. 재조합단백질의약품
23 Coagulation factors VIII and VII 재조합단백질의약품
24 Direct thrombin inhibitors (DTIs) A class of anticoagulant drugs targeting thrombin (i.e. factor IIa) since 1990s, which can be used to prevent and treat thromboses ( 혈전 )andembolisms ( 색전 ) due to blood clots caused by various diseases. cf. heparin and warfarin Based on hirudin (from medicinal leeches such as Hirudo medicinalis) Desirudin (Iprivask), Lepirudine (Refludan) 재조합변형거머리트롬빈저해제히루딘 Bivalirudin (Angiomax) 히루딘활성부위만을갖는 20 aa 합성펩타이드 cf. 저분자 DTIs ( 합성의약품 ) Argatroban: 히루딘활성부위에대한 peptidomimetic design 재조합저해제 / 길항제
25 Enfuvirtide Fuzeon the first of a novel class of antiretroviral drugs used in combination therapy for the treatment of HIV 1 a biomimetic peptide p designed to mimic the HIV 1 fusion machinery component. Binding to gp41, a viral transmembrane protein, then undergoes a conformational change that assists in the fusion of the viral membrane to the host membrane, preventing the creation of an entry pore for the capsid, keeping it out of the cell. Too expensive (~US$25,000 per year) and inconvenient Ac-Tyr-Thr-Ser-Leu-Ile-His-Ser-Leu-Ile-Glu-Glu-Ser-Gln-Asn-Gln-Gln-Glu-Lys-Asn-Glu-Gln-Glu-Leu-Leu-Glu-Leu-Asp-Lys-Trp-Ala-Ser-Leu-Trp-Asn-Trp-Phe-NH 2 항바이러스바이오의약품
26 Hormones A class of signaling molecules produced by glands, which are transported to circulatory system to target distant organs to regulate physiology and behavior Amines (amino acids), peptide/proteins, steroids Hormones as biopharmaceuticals from animals (only for those with the same structures) from dead humans from human excretions such as urine (LH, FSH etc) from engineered biosystems : Humulin is the first biopharmaceutical (1982) 재조합호르몬
27 Insulins Insulin? a peptide hormone produced by beta cells of the islets of Langerhans in the pancreas undergoes extensive posttranslational modifications and deposited as hexamers (requring Zn2+) cf. 시판인슐린제제는페놀성첨가제함유 ( 화학적안정성높임 ) Diabetes mellitus ( 당뇨병 ) Type 1 DM ( 인슐린의존성 ): 인슐린결핍 ( 자가면역질환 ) Type 2 DM ( 인슐린저항성 ): 인슐린수용체결핍 ( 인슐린반응없음 ) A B folded mature 재조합호르몬
Representative Insulin Derivatives 28 Insulin Detemir myristic acid Insulin Glargine Insulin Aspart Insulin lispro 재조합호르몬
29 당뇨병및저혈당증 재조합호르몬
인슐린제제의종류 30 해리및지속시간에따른구분 : 표준, 속효성, 중간형, 지속성, 혼합형 제형 Onset ( 해리 ) Peak ( 최대 ) Duration ( 지속 ) 표준인슐린제제 (Humulin) 30~60 min 2~3 h 6~8 h 속효성인슐린제제 (NovoLog/NovoRapid) <15 min ~1 h 3~6 h 중간형인슐린제제 1~3 h 6~12 h 18~24 h 지속성인슐린제제 (Lantus, Levemir) 8~20 h 24~28 h 표준 (regular or short acting) 인슐린제제 아연 (0.4%) 과페놀성보존제첨가에의해화학적으로안정화되어있음 세포막으로흡수되기전에이량체나단량체로해리되어야함 (0.5 1 h) 속효성 (fast acting) 인슐린제제 식후포도당농도증가에보다자연스럽게대응하기위해개발 돌연변이를통해단량체전환속도증가및단량체안정화 (<15 min) Insulin aspart (B28 Pro is changed to Asp) Insulin lispro (B28 29 Pro Lys are changed to Lys Pro) ) 재조합호르몬
인슐린제제의종류 31 중간형 (intermediate acting) 인슐린제제 해리시간을지연시켜 (1 3h) 활성지속시간을연장시킴 (18 24 h. 1 일 1 회투약 ) 프로타민첨가, 아연함량등에따라달라짐 : NPH insulin, Lente insulin NPH (neutral protamine Hagedorn) insulin 인슐린과프로타민을복합결정화하여만들어진중성결정상현탁액프로타민에대한면역원성이발생할수있음 Lente insulin, Ultralente insulin 육량체아연인슐린결정과무정형인슐린입자의혼합제제 Ultralente insulin 활성이 36 h 까지지속 ( 지속성인슐린?) (NPH보다지속성높음 ) Discontinued in the US by Eli Lilly (2005) due to declining use by physicians 지속성 (long acting) 인슐린제제 서서히용출 (20 2626 h). 일부는 peakless profile 을보임 Insulin glargine (A21 Asn to Gly and two Arg addition at B31 32: more neutral pi) Insulin detemir (myristic acid is attached to the B29 Lys for albumin binding) 재조합호르몬
인슐린제형및투여 32 제형 주사제제 직사광선을피해 2 8 C 냉장보관 투여 매우가는주사바늘사용 ( 주사시통증경감목적 ) 펜주입기 (pen injector), cartridge형주입기정확하고재현성있는용량조절간편한휴대다양한투여용량 재조합호르몬
33 Toujeo vs Lantus Toujeo was approved by KFDA in August, enabling the drug s domestic sales to begin within the year, according to the company. It has been more than 10 years since Lantus was introduced, d revolutionizing i i diabetes treatment methods around the world by enabling out of hospital insulin injections, said Sanofi Aventis Korea general manager Bae Kyung eun. Sanofi has since continued its product innovation to launch htoujeo, a next generation ti basal insulin that has focused on maximizing the patient s comfort and safety, she said. 재조합호르몬
34 Growth Hormone (GH) aka. somatotrpin (somatropin) A peptide hormone (191 aa) that stimulates growth, cell proliferation and regeneration 성장과발육에필수적인단백질로지질분해, 단백질합성촉진및대사억제등을포함한다양한생체대사에영향을줌 ( 연골 / 뼈발달, 지방분해, 글리코겐분해 ) 뇌하수체전엽의신체친화성세포에서분비 시상하부에서조절 GHRH: 분비촉진 GHRIH (somatostatin) : 분비억제 20세부터매년 10~15% 씩분비량감소 : 유아기, 청소년기에가장많이분비 GH 수용체및결합단백질 GHR: 대부분의조직, 간에가장많이존재 GHBP : 혈중 GH와결합,GH소거 (40~45%) 재조합호르몬
35 Growth Hormone (GH) cf. Clinical application of hgh GH 결핍시나타나는증상 ( 소아 ): 성장장애, 만성신장질환 GH 투여로개선되는증상 ( 어른 ): 유즙분비증가, 노화억제, 비만치료 / 체지방감소, 근육강도및운동능력향상, 골밀도증가, 배란유도 GH 투여방식 : 피하주사 ( 매일 ) 수용체결핍에의한성장호르몬저항성에도효과 Turner 증후군개선성염색체하나가소실된여성질환성장감소, 난소미숙등이차성징불완전 GH + oxandrolone 과함께투여 Examples Prototropin, Nutropin, Humatrope, Genotropin, Norditropin, Tev tropin, Bop tropin, p Zomacton, Brook Christa Shields (May 31. 1965) Serostim, Saizen, Zorbtive, Somavert Princeton Alumnus 재조합호르몬
36 Gonadotropins 생식선을자극하는호르몬으로 2 차성징과생식기능을조절 생식선자극호르몬합성부위생리작용 난포자극호르몬 (FSH) 여성 : 난포세포성장자극뇌하수체전엽 ( 임상적으로사용 ) 남성 : 정자생산항진 황체형성호르몬 (LH) 뇌하수체전엽 여성 : 배란유도남성 : testosterone 합성 융모막성생식선자극호르몬 (CG) 뇌하수체전엽임산부 : 황체유지, 임신유지 Examples Follitropin α (Gonal F), Follitropin β (Follistim) Lutropin α (Luveris): used in combination with Gonal F Oxytocin (Pitocin) 재조합호르몬
37 Thyroid hormone and Parathormone 갑상선호르몬 & 부갑상선호르몬 Examples Calcitonin (Fortical): salmon calcitonin (human calcitonin의 30배효과 ) Teriparatide (Forteo), Preotact (Preos) 재조합호르몬
38 Other Growth Factors A class of naturally occurring molecules capable of stimulating cellular growth, proliferation, healing, and differentiation. They include cytokines and hormones Examples G CSF ( 과립구콜로니촉진인자 ): 호중구생성촉진 ( 호중구감소증치료 ) Filgrastim, pegfilgrastim ( 대장균 ), Lenograstim (CHO) 부작용이적어재조합 GM CSF 보다많이사용됨 GM CSF ( 과립구 대식세포콜로니촉진인자 ): 호중구생성촉진 Sargramostim ( 대장균생산 ), Morgramostim ( 효모 ), Regramostim (CHO) EPO ( 에리쓰로포이에틴 ): 적혈구생성과조절에관여 각종빈혈치료 ( 만성신장기능이상, 류마티스성관절염, 항암화학요법, AIDS) 90% 이상신장에서생산, 산소분압변화에반응하여 EPO 생산을조절 ( 조혈모세포가적혈구로분화하는데 EPO, IL 3, GM CSF가관여 ) N 결합당쇄, O 결합당쇄를모두가진당단백질로, 당화가어려운점이특징임 Epoetin α (Epogen, Procrit, Eprex), Epoetin β (NeoRecormon) TPO ( 트롬보포이에틴 ): 거핵세포및혈소판생성과조절에관여 재조합성장인자
39 Pegfilgrastim and Filgrastim Neulasta and Neupogen Filgrastim is a G CSF dervied from E. coli : stimuates bone marrow to increase the numbers of HSCs and neutrophils. Pegfilgrastim is a peggylated version of Filgrastim to increase the human half life (15 18 h vs 3 4 h). to treat neutropenia, which can be caused by chemotherapy and BM transplantation. 재조합싸이토카인 / 성장인자
40 Erythropoietin (EPO) Epogen/Procrit/Eprex (Epoietin α) NeoRecormon (Epoietin β) Aranesp/NESP (Darbepoetin α) contains two more N linked oligosaccharides with increased half life (~3 times) They are produced in cell culture system (CHO) to treat anemia, commonly associated with chronic renal failure and cancer therapy A potential drug that can be abused by athletes EPO treatment may increase the risk of cardiovascular problems. 재조합싸이토카인 / 성장인자
41 조직세포성장인자 조직손상 ( 상처 ) 이치유되는과정중여러세포에서생산되는다양한성장인자 Collagen, elastin 등의기질물질을생산하는섬유아세포를포함하여상피세포, 혈관내피세포등을활성화하여성장을촉진함 Examples FGF ( 섬유아세포성장인자 ) IGF 1, IGF 2 ( 인슐린양성장인자 )(somatomedin): Mescarsemin (Increlex) EGF ( 상피세포성장인자 ): Nepidermin (Regen D) PDGF ( 혈소판유래성장인자 ): Becaplermin(Regramex Gel) 외용제 VEGF ( 혈관내피세포성장인자 ) 재조합성장인자
42 효소의약품 효소질환재조합효소 Urate oxidase 고요산혈증 Rasburicase (Elitek) Adenosine deaminase SCID Pegadamase bovine (Adagen) Iduronate 2 sulfatase Hunter 증후군 Aldursulfase (Elaprase) α galactosidase Fabry 병 Agalsidase α (Replagel), Agalsidase β (Fabrazyme) α glucosidase Pompe 병 Aglucosidase α (Myozyme) α L iduronidase Hurler 증후군 Laronidase (Aldurazyme) β glucocerebrosidae Gaucher 병 Aglucerase(Ceredase), Imiglucerase(Cerezyme) tpa 심근경색, 뇌졸증 Alteplase (Activase) L asparaginase 급성림프구성백혈병 Pegaspagase (Oncaspar) 재조합효소