1、雙特異性T細胞銜接器抗體 Bispecific T-cell Engager (BiTE) Antibody1. 背景介紹1.1 腫瘤免疫治療的熱點1.2 雙特異性抗體的結構1.3 無Fc段雙特異性抗體的發展1.4 雙特異性抗體的作用1.5 Fc段的作用1.6 雙特異性抗體的臨床應用2. 作用機理2.1 BiTE的作用機理2.2 BiTE的抗原表位擴展效應3. 臨床研究3.1 Blinatumomab簡介3.2 Blinatumomab的臨床試驗結果4. 前景展望4.1 BiTE的優點和缺點4.2 BiTE的改進4.3 用于BiTE的腫瘤相關抗原4.4 雙特異性NK細胞銜接器(BiKE)4.5

2、 BiTE與免疫檢查點抑制劑的聯合4.6 溶瘤病毒表達BiTE4.7 自體T細胞表達BiTE腫瘤免疫治療的熱點單克隆抗體免疫檢驗點單抗PD1/PDL1，CTLA4，CD47雙特異性抗體(BsAb)Triomab，BiTE，TandAb，DART過繼細胞療法腫瘤浸潤淋巴細胞療法(TIL)T細胞受體療法(TCR)嵌合抗原受體修飾的T細胞療法(CAR)腫瘤疫苗樹突狀細胞(DC)疫苗蛋白/多肽/DNA疫苗定義：通過調動宿主的免疫防御機制或給予某些生物活性物質以取得或者增強腫瘤免疫效應的治療方法。IgG抗體結構雙特異性抗體的結構Spiess C, Zhai Q, Carter P J. Alternat

3、ive molecular formats and therapeutic applications for bispecific antibodiesJ. Molecular immunology, 2015.目前已報道的雙特異性抗體結構雙特異性抗體的結構目前已進入臨床試驗的雙特異性抗體Kontermann R E, Brinkmann U. Bispecific antibodiesJ. Drug discovery today, 2015.無Fc段雙特異性抗體的發展Spiess C, Zhai Q, Carter P J. Alternative molecular formats an

4、d therapeutic applications for bispecific antibodiesJ. Molecular immunology, 2015.Geering B, Fussenegger M. Synthetic immunology: Modulating the human immune systemJ. Trends Biotechnol, 2014.無Fc段雙特異性抗體的發展（*代表已進入臨床試驗）已進入臨床試驗的無Fc段雙特異性抗體Diabody：VHa-VLb和VHb-VLascDiabody：VHa-VLb-VHb-VLaDART: VHa-VLb和VHb-

5、VLa 二硫鍵(VHa=VHb)TandAb: VHa-VLb-VHb-VLa 二聚化BiTE: VLa-VHa-VLb-VHbNanobody: VH雙特異性抗體的作用Kontermann R E, Brinkmann U. Bispecific antibodiesJ. Drug discovery today, 2015.重定向T細胞或NK細胞殺傷腫瘤細胞結合兩種細胞受體或配體，抑制信號轉導催化兩種蛋白相互作用雙特異性抗體的三種作用機制Fc段的作用Grandjenette C, Dicato M, Diederich M. Bispecific Antibodies: An Innova

6、tive Arsenal to Hunt, Grab and Destroy Cancer CellsJ. Current Pharmaceutical Biotechnology, 2015, 16(8): 670-683.具有Fc段的雙抗(如Triomab)發揮作用無Fc段的雙抗(如BiTE)發揮作用特點：容易提純，穩定性高，半衰期長（FcRn介導再回收）功能：結合輔佐細胞（NK，DC，M、粒），抗體依賴的細胞毒性（ADCC），補體依賴的細胞毒性（CDC）特點：穿透能力強，半衰期短（降解、腎清除）分子靶點結構研發公司適應癥臨床試驗狀態注冊號Catumaxomab(Removab)EpCAM

7、CD3TrioMabFresenius Biotech(Trion)EpCAM陽性腫瘤的惡性腹水2009EUEpCAM陽性腫瘤的惡性腹水II/III完成NCT00836654胃癌、胃腺癌、食管胃連接部腺癌、腹膜轉移癌、卵巢癌、上皮卵巢癌、惡性腹水、輸卵管腫瘤、腹膜腫瘤II/III12項試驗Ertumaxomab(Rexomun)Her2CD3TrioMabFresenius Biotech(Trion)Her2/Neu陽性晚期固體瘤I/IINCT01569412Blinatumomab(Blincyto, MT103, AMG103)CD19CD3BiTEAmgen(Micromet)復發/難

8、治性費城染色體陰性前體B細胞急性淋巴細胞白血病2014USA急性淋巴細胞白血病IINCT01466179非霍奇金淋巴瘤、急性淋巴細胞白血病、彌漫大B細胞淋巴瘤I/II/III16項試驗MT110(AMG110)EpCAMCD3BiTEAmgen(Micromet)固體瘤I完成NCT00635596MT111(MEDI-565)CEACD3BiTEAmgen(Micromet)胃腸道腺癌I完成NCT01284231MT112(BAY2010112)PSMACD3BiTEBayer(Micromet)前列腺腫瘤INCT01723475AMG330CD33CD3BiTEAmgen(Micromet)

9、復發/難治性急性髓細胞白血病INCT02520427JNJ-64052781CD19CD3DARTB細胞慢性淋巴細胞白血病、彌漫大B細胞淋巴瘤、濾泡性淋巴瘤、套細胞淋巴瘤、急性淋巴細胞白血病INCT02454270MGD006CD123CD3DARTMacroGenicsand Sevier急性髓細胞白血病INCT02152956MGD007gpA33CD3DARTMacroGenicsand Sevier結直腸癌INCT02248805MGD009B7-H3CD3DARTMacroGenicsand Sevier間皮瘤、膀胱癌、黑色素瘤、頭頸部鱗狀細胞癌、卵巢癌、甲狀腺癌、乳腺癌、胰腺癌、前

10、列腺癌、結腸癌、軟組織肉瘤INCT02628535AFM13CD30CD16aTandAbAffimed霍奇金淋巴瘤I完成NCT01221571霍奇金淋巴瘤IINCT02321592AFM11CD19CD3TandAbAffimed復發/難治性B細胞非霍奇金淋巴瘤INCT02106091rM28HMW-MAACD28Tandem scFvUniversity HospitalTubingen惡性黑色素瘤I/II完成NCT00204594From by December 25, 2015目前已進入臨床試驗的用于連接效應細胞與腫瘤細胞的雙特異性抗體BiTE的作用機理Oak E, Bartlett

11、N L. Blinatumomab for the treatment of B-cell lymphomaJ. Expert opinion on investigational drugs, 2015, 24(5): 715-724.Zimmerman Z, Maniar T, Nagorsen D. Unleashing the clinical power of T cells: CD19/CD3 bi-specific T cell engager (BiTE) antibody construct blinatumomab as a potential therapyJ. Inte

12、rnational immunology, 2014: dxu089.BiTE的結構BiTE的作用機理BiTE與CD3和TAA結合，TCR-CD3復合物激活T細胞T細胞增殖，分泌細胞因子，CD25、CD69表達上調溶細胞突觸形成，釋放穿孔素和顆粒酶B穿孔素依賴Ca2+聚合形成跨膜通道，顆粒酶B進入腫瘤細胞顆粒酶B激活Caspase通路，誘導腫瘤細胞凋亡ESK1-BiTEESK1：模擬TCR單克隆抗體，識別通過HLA-A*02:01呈遞的胞內腫瘤蛋白WT1Dao T, Pankov D, Scott A, et al. Therapeutic bispecific T-cell engager

13、antibody targeting the intracellular oncoprotein WT1J. Nature biotechnology, 2015, 33(10): 1079-1086.BiTE的抗原表位擴展效應WT1-RMF和HER2/Neu陽性卵巢癌患者的外周血單個核細胞(PMBC)給予ESK1-BiTE并接受自體腫瘤細胞刺激和再刺激后抗原表位特異性應答Blinatumomab簡介商標名Blincyto有效成分Blinatumomab生產商Amgen Inc.FDA批準日期2014年12月適應癥復發/難治性費城染色體隱性前體B細胞急性淋巴細胞白血?。≧/R Ph- B-AL

14、L）半衰期2.1小時毒副作用細胞因子釋放綜合癥（CRS），神經毒性等單抗種類鼠源化BiTE結構VLCD19-VHCD19-VHCD3-VLCD3靶點CD19CD3給藥方式靜脈費用兩個療程共17.8萬美元Przepiorka D, Ko C W, Deisseroth A, et al. FDA approval: blinatumomabJ. Clinical Cancer Research, 2015, 21(18): 4035-4039.Rogala B, Freyer C W, Ontiveros E P, et al. Blinatumomab: enlisting serial ki

15、ller T-cells in the war against hematologic malignanciesJ. Expert opinion on biological therapy, 2015, 15(6): 895-908.Blinatumomab的作用機理Blinatumomab的臨床試驗結果注冊號研究人群樣本數平均年齡CR/CRhMRD陰性率無復發生存期總生存期NCT01466179II: R/R B-ALL18939(18-79)43%(36-51)82%(72-90)5.9 months(4.8-8.3)6.1 months(4.2-7.5)NCT00560794II: M

16、RD B-ALL2050(20-77)80%(56-94)NCT01207388II: MRD B-ALL11645(18-76)78%(69-85)NCT01209286II: R/R B-ALL3640(18-79)70%(52-84)69%(52-84)7.7 months(5.8-9.6)9.9 months(8.5-15.0)I: 兒童R/R B-ALL411832%77%8.3 months(3.0-16.0)5.7 months(3.3-9.7)I/II: 兒童R/R B-ALL399(2-16)31%(17%-48%)42%5.6 months(2.6-12.1)4.3 mon

17、ths(3.6-8.1)首次復發完全緩解(CR)：30-45%總生存期(OS)：5-9月原發難治/首次緩解小于12月/造血干細胞移植(HSCT)后復發/多線治療無效完全緩解(CR)：20-30%總生存期(OS)：3-6月Topp M S, Gkbuget N, Stein A S, et al. Safety and activity of blinatumomab for adult patients with relapsed or refractory B-precursor acute lymphoblastic leukaemia: a multicentre, single-arm

18、, phase 2 studyJ. The Lancet Oncology, 2015, 16(1): 57-66.Blinatumomab的I/II期臨床試驗結果From by December 25, 2015BiTE的優點和缺點優點CD3TAA，重定向CD4+和CD8+T細胞（CD8為主，記憶T為主，Treg？）不依賴于TCR和MHCI呈遞，不需要共刺激腫瘤特異性半衰期短（隨時停止應對突發）組織穿透能力強，調動固體瘤內TIL和Treg清除微小殘留疾病（MRD）療效好，相比單抗劑量低，低E:T比不易出現耐藥無基因操作、細胞移植風險，易獲得缺點半衰期短，難維持血清水平（持續靜脈給藥，結合PE

19、G或白蛋白）Laszlo G S, Gudgeon C J, Harrington K H, et al. T-cell ligands modulate the cytolytic activity of the CD33/CD3 BiTE antibody construct, AMG 330J. Blood cancer journal, 2015, 5(8): e340.Kontermann R E. Strategies for extended serum half-life of protein therapeuticsJ. Current opinion in biotechn

20、ology, 2011, 22(6): 868-876.Koristka S, Cartellieri M, Arndt C, et al. Tregs activated by bispecific antibodies: Killers or suppressors?J. OncoImmunology, 2015, 4(3): e994441.BiTE的改進延長半衰期改變配體腫瘤相關抗原多種CD3CD8？，CD16，CD28抗原結合位點多個與其他腫瘤療法結合Rituximab單抗免疫檢查點抑制劑體內持續穩定表達溶瘤病毒T細胞間充質干細胞應用于BiTE的腫瘤相關抗原靶點名稱適應癥臨床試驗CD

21、19Blinatumomab/MT103/AMG103/MEDI-538急性淋巴細胞白血病非霍奇金淋巴瘤I/II/IIIEpCAMMT110固體瘤ICEAMT111/MEDI-565胃腸道腺癌IPSMAMT112/BAY2010112前列腺癌ICD33急性髓細胞白血病IEGFR結腸癌臨床前Her2臨床前EphA2bscEphA2CD3固體瘤臨床前MCSPMCSP-BiTE黑色素瘤臨床前ADAM17A300E前列腺癌臨床前PSCA前列腺癌臨床前17-1A臨床前NKG2D配體固體瘤、白血病臨床前CD133膠質瘤臨床前WT1ESK1-BiTE固體瘤、白血病臨床前臨床和臨床前用于BiTE的腫瘤抗原靶點

22、Huehls A M, Coupet T A, Sentman C L. Bispecific T-cell engagers for cancer immunotherapyJ. Immunology and cell biology, 2014.雙特異性NK細胞銜接器(BiKE)CD16CD33BiKE對健康供者髓系抑制性細胞(MDSC)具有顯著殺傷作用骨髓增生異常綜合征(MDS)NK細胞 CD16 髓系抑制性細胞(MDSC)CD16CD33激活NK細胞，殺傷CD33+MDSCGleason M K, Ross J A, Warlick E D, et al. CD16xCD33 bisp

23、ecific killer cell engager (BiKE) activates NK cells against primary MDS and MDSC CD33+ targetsJ. Blood, 2014, 123(19): 3016-3026.BiTE與免疫檢查點抑制劑的聯合Khnke T, Krupka C, Tischer J, et al. Increase of PD-L1 expressing B-precursor ALL cells in a patient resistant to the CD19/CD3-bispecific T cell engager antibody blinatumomabJ. Journal of hematology & oncology, 2015, 8(1): 1-5.Osada T, Patel S P, Hammond S A, et al. CEA/CD3-bispecific T cell-engaging (BiTE) antibody-mediated T lymphocyte cytotoxicity maximized by inhibition of