TWI490231B - 可用作誘發抗原專一性ｔ細胞反應之免疫原性增強劑之融合蛋白 - Google Patents

可用作誘發抗原專一性ｔ細胞反應之免疫原性增強劑之融合蛋白 Download PDF

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TWI490231B
TWI490231B TW102144581A TW102144581A TWI490231B TW I490231 B TWI490231 B TW I490231B TW 102144581 A TW102144581 A TW 102144581A TW 102144581 A TW102144581 A TW 102144581A TW I490231 B TWI490231 B TW I490231B
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fusion protein
amino acid
peptide
antigen
binding site
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TW102144581A
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TW201428001A (zh
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Hsiu Kang Chang
Wei I Chou
Jiun Ming Wu
Chia Mao Wu
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Thevax Genetics Vaccine Co Ltd
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Description

可用作誘發抗原專一性T細胞反應之免疫原性增強劑之融合蛋 白
本發明大致係關於融合蛋白及免疫。
分子生物學使吾人得以生產次單元疫苗，其中，次單元疫苗之免疫原為母蛋白或母複合物之一片段或次單元。最好能開發出一種穩定疫苗，其一方面可誘發T細胞致敏反應，一方面又具有足夠之靈活度，俾與各品系感染源之序列結合。
本發明之一態樣係關於一種融合蛋白，其包含：(a)一抗原呈現細胞(APC)結合部位或一CD91受體結合部位，其位於融合蛋白之N端；(b)一移位胜肽，其就長度而言具有34-112個胺基酸殘基，且包含一與序列編號第4、2、3或6號至少90%相同之胺基酸序列，移位胜肽位於APC結合部位或CD91受體結合部位之C端；(c)一病原體的一抗原，其位於移位胜肽之C端；(d)一核輸出信號，其包含序列編號第13號之胺基酸序列；及(e)一內質網(ER)滯留序列，其位於融合蛋白之C端。
在本發明之一實施例中，APC結合部位或CD91受體結合部位係一多胜肽，且多胜肽包含一與下列各項所組成之群組中選出之序列至少90%相同之胺基酸序列：序列編號第1及8-11號。
在本發明之另一實施例中，核輸出信號包含序列編號第14號之胺基酸序列。
在本發明之另一實施例中，內質網滯留序列包含序列編號第15號之胺基酸序列。
在本發明之另一實施例中，核輸出信號係位於移位胜肽與抗原之間。
在本發明之另一實施例中，核輸出信號係位於抗原與內質網滯留序列之間。
在本發明之另一實施例中，核輸出信號及ER滯留序列形成一融合胜肽，融合胜肽包含一與序列編號第12號至少90%相同之胺基酸序列。
在本發明之另一實施例中，移位胜肽就長度而言具有34-61個胺基酸殘基。
在本發明之另一實施例中，移位胜肽就長度而言具有34-46個胺基酸殘基。
在本發明之另一實施例中，APC結合部位或CD91受體結合部位完全不含綠膿桿菌外毒素A(PE)結合部位I之胺基酸序列。
在本發明之另一實施例中，APC結合部位或CD91受體結合部位包含序列編號第8號之胺基酸序列。
在本發明之另一實施例中，APC結合部位或CD91受體結合部位之胺基酸序列為序列編號第1號。
在本發明之另一實施例中，抗原係一病原體之兩個以上抗原胜肽之融合抗原。
在本發明之另一實施例中，ER滯留序列就長度而言具有多於4個胺基酸殘基。
在本發明之另一實施例中，移位胜肽包含一與序列編號第4、2、3或6號至少95%相同之胺基酸序列。
在本發明之另一實施例中，APC結合部位或CD91受體結合部位之特徵在於可辨識出並結合於一抗原呈現細胞(APC)上之一受體，其中抗原呈現細胞係由下列各項所組成之群組中選出：樹狀細胞、單核球、B細胞及淋巴球。
在本發明之另一實施例中，病原體係由下列各項所組成之群組中選出：PRRSV、PCV、FMDV、CSFV、NDV、豬傳染性胃腸炎病毒(TGEV)、豬流行性下痢病毒(PEDV)、流行感冒病毒、偽狂犬病病毒、小 病毒、豬水疱病毒(SVDV)、痘病毒、輪狀病毒、黴漿菌肺炎、疱疹病毒，禽類傳染性支氣管炎及傳染性華氏囊症病毒。
本發明之另一態樣實質上係由下列各項組成，或係由下列各項組成：(a)一抗原呈現細胞(APC)結合部位或一CD91受體結合部位，其位於融合蛋白之N端；(b)一移位胜肽，其就長度而言具有34-112個胺基酸殘基，且包含一與序列編號第4、2、3或6號至少90%相同之胺基酸序列，移位胜肽係位於APC結合部位或CD91受體結合部位之C端；(c)一病原體之一抗原，其位於移位胜肽之C端；(d)一核輸出信號，其包含序列編號第13號之胺基酸序列；及(e)一內質網滯留序列，其位於融合蛋白之C端。
本發明之又一態樣係關於一種疫苗組合物，其包含前述融合蛋白及一佐劑。
本發明之再一態樣係關於一種用以誘發並增強病原體抗原專一性T細胞反應之方法，方法包含下列步驟：對有需要之受治者施用一疫苗組合物，其中疫苗組合物包含一治療有效量之前述融合蛋白；及藉此誘發並增強對病原體抗原具有專一性之T細胞反應。
本發明亦關於將前述融合蛋白或疫苗組合物用於誘發並增強病原體抗原專一性T細胞反應，或將前述融合蛋白用於製造一可誘發並增強病原體抗原專一性T細胞反應之藥物。
第1A圖為全長綠膿桿菌外毒素A(PE)及其部分片段之示意圖。
第1B、1C圖為載體圖。
第2至5圖分別繪示本發明融合蛋白誘發並增強CD8+ /IFN-γ+ T細胞(第2A至5A圖)及CD4+ /IFN-γ+ T細胞(第2B至5B圖)媒介免疫原性之效果。
第6圖顯示動物組別、為動物接種之疫苗與劑量，以及接種時程。
第7、8圖分別繪示接種不同融合蛋白或安慰劑之動物組別之腫瘤大小 曲線及無腫瘤小鼠百分比。
本節將透過以下範例詳細說明本發明之內容，唯該等範例僅供例示之用，熟習此項技術者當可輕易思及多種修改及變化之方式。以下將詳述本發明之多種實施例。請參閱圖式，圖中相同標號均代表相同元件。在本說明書及後附之申請專利範圍中，除非上下文另外載明，否則「一」及「該」亦可解釋為複數。此外，在本說明書及後附之申請專利範圍中，除非上下文另外載明，否則「中」及「內」均包括「位於其中」及「位於其上」。再者，說明書中可能附有標題及小標題以方便閱讀，但該等標題並不影響本發明之範圍。以下將詳細定義本說明書所用之若干術語。
定義
本說明書所使用之術語在本發明範圍內及各術語之特定上下文中大致具有其相關領域中之通常涵義。本說明書中用以描述本發明之特定用語將在下文中或在本說明書之他處加以說明，以利業界人士瞭解本發明之相關敘述。為便於閱讀，某些術語可能會以斜體及/或引號等格式加以凸顯，但使用此些凸顯格式並不影響術語之範圍與意義。同一術語在相同上下文中之範圍與意義皆相同，此與是否採用凸顯格式無關。此外。同一件事之表達方式不止一種。因此，本文所討論之術語或可以替代用語及同義詞取代，且一術語是否在本文中經詳細說明或討論並無任何特別意義。本文提供某些術語之同義詞，但使用某一或多個同義詞並不表示排除其他同義詞。本說明書所提供之範例(包括此處所討論之術語範例)僅供說明之用，而非用於侷限本發明或任何例示術語之範圍與意義。同樣，本發明並不限於本說明書所列舉之各種實施例。
除非另有定義，否則本文中所有技術與科學術語之意涵均與熟習本發明相關技術者所普遍瞭解者相同。若有相互衝突之處，則以本說明書及其所提供之定義為解釋依據。
「抗原呈現細胞(APC)或輔助細胞」一詞係指一細胞可透過其表面上之主要組織相容性複合物(MHC)與外來抗原複合，進而呈現外來抗原，使T細胞得以經由T細胞受體(TCR)辨識出MHC與抗原之複合物。此種細胞可處理抗原，並對T細胞呈現抗原。專業抗原呈現細胞主要包 括以下幾類：樹狀細胞(DC)、巨噬細胞(其亦為CD4+，故亦可能感染HIV)、單核球及若干B細胞。
「抗原呈現細胞(APC)結合部位」一詞係指一可結合至一抗原呈現細胞(APC)之部位(其為一多胜肽)。此APC結合部位可為一多胜肽，其中多胜肽包含一與下列各項所組成之群組中選出之序列至少90%相同之胺基酸序列：序列編號第1及8-11號。APC結合部位係一配位子，其可辨識出並結合於APC上之受體。
分化群91(CD91)係一可於細胞膜中形成受體且參與受體媒介胞吞作用之蛋白。
「PEt 」一詞係指一就長度而言具有34-112個胺基酸殘基之移位胜肽(或移位部位)。PEt 可包含一與序列編號第2-4及6號至少90%相同之胺基酸序列。例如，PEt 之胺基酸序列可為PE之a.a.280-a.a.313片段(序列編號第4號)、a.a.268-a.a.313片段(序列編號第3號)、a.a.253-a.a.313片段(序列編號第2號)或a.a.253-a.a.364片段(序列編號第6號)。換言之，PEt 之胺基酸序列可含有PE部位II(a.a.253至a.a.364；序列編號第6號)之任一區，只要其包含a.a.280-a.a.313(序列編號第4號)此一必要序列(亦即必要片段)即可。
PE407 (序列編號第7號)在先前專利(美國專利第7,335,361B2號)中係稱為PE(△III)。
「最小移位胜肽」一詞係指PE253-313 (序列編號第2號)，其可將一抗原移位至一目標細胞之細胞質內。
「內質網(ER)滯留序列」一詞係指一胜肽之作用在於協助一抗原從細胞質移位至ER，並將其留在ER之內腔中。一ER滯留序列包含序列Lys Asp Glu Leu(KDEL；序列編號第15號)或RDEL。一ER滯留序列可包含序列KDEL、RDEL、KDELKDELKDEL(K3；序列編號第16號)、KKDLRDELKDEL(K3；序列編號第17號)、KKDELRDELKDEL(K3；序列編號第18號)或KKDELRVELKDEL(K3；序列編號第19號)。
一核輸出信號(NES)係指一蛋白中具有4個疏水性殘基之短胺基酸序列，其中胺基酸序列可利用核傳輸技術，透過核孔複合體將蛋白從細胞核輸出至細胞質。輸出接受器(exportin)可辨識出NES並加以拘 束。該等疏水性殘基最常見之間隔方式為Lxx KLxx Lx Lx (序列編號第13號)，其中「L」為白胺酸，「K」為離胺酸，「x」為任一種天然胺基酸。例如，一人造NES可包含序列Leu Gln Lys Lys Leu Glu Glu Leu Glu Leu Ala(LQKKLEELELA；序列編號第14號)。
「NESK」一詞係指一NES與一ER滯留信號之融合胜肽(亦即將NES融合至ER滯留信號)。NESK為一人造胜肽，其兼具有核輸出信號(NES)及ER滯留序列之功能。因此，NESK可透過核孔複合體將一抗原從細胞核輸出至細胞質，並協助一抗原從細胞質移位至ER，進而將抗原留在ER之內腔中。例如，NESK之胺基酸序列可為LQKKLEELELAKDEL(序列編號第12號)。
抗原可為一病原體蛋白、多胜肽或胜肽，其可導致由病原體所引發之疾病，或可於病原體所感染之宿主體內誘發一免疫反應；或為與腫瘤相關之抗原(TAA)，其為一專門表現於腫瘤細胞內之多胜肽。所述抗原可選自病原體或癌症細胞，其包括但不限於人類乳突病毒(HPV)、豬繁殖及呼吸症候群病毒(PRRSV)、人類免疫不全症病毒-1(HIV-1)、登革熱病毒、C型肝炎病毒(HCV)、B型肝炎病毒(HBV)、豬環狀病毒2(PCV2)、典型豬瘟病毒(CSFV)、口蹄疫病毒(FMDV)、新城雞瘟病毒(NDV)、豬傳染性胃腸炎病毒(TGEV)、豬流行性下痢病毒(PEDV)、流行感冒病毒、偽狂犬病病毒、小病毒、豬水疱病毒(SVDV)、痘病毒、輪狀病毒、黴漿菌肺炎、疱疹病毒、禽類傳染性支氣管炎、或傳染性華氏囊症病毒、非小細胞肺癌、乳癌、黑色素瘤、淋巴瘤、結腸癌、肝細胞癌，及上列各項之任一組合。例如，在此選用HPV E7蛋白(E7)、HCV核心蛋白(HCV核心)、HBV X蛋白(HBx)做為開發疫苗之抗原。所述抗原可為兩種以上之抗原(可選自一或多種病原體蛋白)融合而成之融合抗原。例如，可將PRRSV ORF6與ORF5片段融合為融合抗原，或將PRRSV與PCV2病原體之抗原蛋白加以融合。
「治療」一詞係指對已罹患癌症、或已遭受感染、或已出現疾病症狀、或具有罹病傾向並且有需要之受治者施用一有效量之融合蛋白，藉以治癒、減輕、解除、改善、緩解或預防疾病、或其症狀、或罹患疾病之傾向。一健康照護專業人士可根據任何適當診斷方法之結果判別出 此種受治者。
「有效量」一詞係指一活性化合物對受治者產生治療效果所需之劑量。熟習此項技術者應知，有效量會隨施用途徑、賦形劑之使用以及是否併用其他治療手段而有所變化。
本發明係關於一種可強化抗原傳送並調控細胞媒介免疫反應之融合蛋白。融合蛋白包含：(a)一抗原呈現細胞(APC)結合部位或一CD91受體結合部位，其位於融合蛋白之N端；(b)一移位胜肽，其就長度而言具有34-112個胺基酸殘基，且包含一與序列編號第2至4及6號至少90%相同之胺基酸序列，移位胜肽係位於APC結合部位或CD91受體結合部位之C端；(c)一病原體之一抗原，其位於移位胜肽之C端；(d)一核輸出信號(NES)；及(e)一內質網(ER)滯留序列，ER滯留序列係位於融合蛋白之C端，其中NES包含序列編號第13號之胺基酸序列。
以融合蛋白PE313 -ORF2-NESK為例，其策略在於以本發明之融合蛋白刺激特定T細胞之製造與活化，其中特定T細胞係指可辨識出第2型豬環狀病毒(PCV2)之病毒殼蛋白ORF2(抗原)者。融合蛋白自N端至C端依序包含：一PE部位I(APC結合部位)；一移位胜肽，其就長度而言具有34-112個胺基酸殘基(例如PE部位II之a.a.253-313)、一截短PCV2 ORF2蛋白(去除N端之核定位信號)；一NES信號；及一ER滯留序列(KDEL)。以PE313 -ORF2-NESK誘發並增強ORF2專一性T細胞免疫反應之基本機構涉及下列步驟：a)結合至樹狀細胞(或抗原呈現細胞)之表面受體(CD91)；b)藉由胞吞作用內部化；c)傳輸至ER，並於移位胜肽前方以弗林進行蛋白質水解；d)加以處理，並透過MHC I複合物呈現；及e)活化對抗原具有專一性之CD4+及CD8+ T細胞。CD4+ Th1細胞可有效率地刺激並增強胞毒CD8+ T細胞免疫反應。結合適應性免疫系統之以上兩種技術，吾人便能以具有專一性之方式殺死PCV2及遭PCV2感染之細胞。
本發明之融合蛋白PE313 -ORF2-NESK在許多方面均與Liao於美國專利第7,335,361號中所揭露之融合蛋白疫苗PE407 -Ag-K3不同。首先，PE313 (序列編號第5號)之長度為94個胺基酸殘基，較PE407 (序列編號第7號)為短，其優點在於可消除由額外PE片段所誘發之體液反應。其次，ER滯留序列亦隨之縮短。僅需要一段KDER或RDER而不需使用K3(即3段 KDER)。再者，僅細胞溶質抗原可被處理並經由MHC第I型通道呈現，因此，將NES信號加入融合蛋白中可提高抗原移位進入胞質液之機會，從而增強對病原體抗原具有專一性之T細胞反應。一病原體之抗原可輸入至細胞核內。結合NES信號後，輸入細胞核內之抗原便可藉由融合蛋白之NES信號而輸出至細胞質。
範例
以下所述僅為根據本發明實施例所提供之例示用設備、器材、方法與相關結果，不應視為本發明之限制。請注意，各範例之標題或子標題僅為方便閱讀之用，亦不應視為本發明之限制。此外，只要本發明係據其本身實施，不論係基於任何特定學說或實行方案，均不受在此所敘及之特定學說之限制，且與所述學說正確與否無涉。
範例1
表現載體之建構
如第1A圖所示，PE含有3個部位(I、II及III)。PE407 係PE從a.a.1至a.a 407之區域。PE407 不含胞毒部位III，僅含有部位I及II。PE313 係PE從a.a.1至a.a.313之區域，因此，PE313 僅含有PE之部位Ia及部位II之部分N端區域。
第1B、1C圖繪示多種表現載體之構造，各構造均包含一抗原呈現細胞(APC)結合部位、一移位胜肽、一抗原(下圖具有核輸出信號(NES)，上圖則無)，及一內質網(ER)滯留序列(上圖中之K3或下圖中之K)，其中ER滯留序列係位於融合蛋白之C端。質體pTac-2-PE313 -NESK、pTac-2-PE407 -K3、pTac-2-RAP1-PE268-313 -NESK及pTac-2-RAP1-PE268-313 -K3之產生方式如下：NdeI PE313 -(EcoRI,XhoI) -NESKXhoI 、NdeI PE407 -(EcoRI,XhoI) -K3XhoI 、NdeI RAP1-(EcoRI) -PE268-313 -(EcoRI,XhoI) -NESKXhoI 及NdeI RAP1-(EcoRI) -PE268-313 -(EcoRI,XhoI) -K3XhoI 片段以聚合酶連鎖反應法(PCR)合成後，利用康黴素抗藥基因將其綁入一pUC18骨幹中以形成各質體。
之後再將一目標DNA(其可為一目標病原體之一抗原或融合抗原編碼)插入前述質體中，藉以產生可表現一融合蛋白之表現載體。例如，可為第2型豬環狀病毒(PCV2)ORF2(序列編號第20號)、典型豬瘟病毒(CSFV)E2(序列編號第21號)、口蹄疫病毒(FMDV)VP1-3A(序列 編號第24號)及新城雞瘟病毒(NDV)FHN(序列編號第27號)等抗原編碼之DNA片段係於合成後分別插入質體pTac-2-PE313 -NESK與pTac-2-PE407 -K3中，藉以產生下列表現載體：(1)PE313 -ORF2-NESK；(2)PE407 -ORF2-K3；(3)PE313 -E2-NESK；(4)PE407 -E2-K3；(5)PE313 -VP1-3A-NESK；(6)PE407 -VP1-3A-K3；(7)PE313 -FHN-NESK及(8)PE407 -FHN-K3。可為第16型人類乳突病毒E7(序列編號第28號)抗原編碼之DNA片段則於合成後分別插入質體pTac-2-PE407 -K3、pTac-2-RAP1-PE268-313 -NESK及pTac-2-RAP1-PE268-313 -K3中以產生下列表現載體：(9)PE407 -E7-K3；(10)RAP1-PE268-313 -E7-NESK及(11)RAP1-PE268-313 -E7-K3。
範例2
蛋白表現
帶有可表現下列融合蛋白之質體之大腸桿菌(E.coli )BL21細胞係於37℃之溫度下，以內含25ppm康黴素之Luria Bertani培養液培養：(1)PE313 -ORF2-NESK；(2)PE407 -ORF2-K3；(3)PE313 -E2-NESK；(4)PE407 -E2-K3；(5)PE313 -VP1-3A-NESK；(6)PE407 -VP1-3A-K3；(7)PE313 -FHN-NESK；(8)PE407 -FHN-K3；(9)PE407 -E7-K3；(10)RAP1-PE268-313 -E7-NESK及(11)RAP1-PE268-313 -E7-K3。當培養基到達早期對數生長期(A600=0.1至0.4)時，加入異丙基-β-D-硫代半乳糖苷(IPTG)，使其最終濃度為0.5至2mM以達誘發之目的。誘發4小時後收集細胞，隨即以-70℃之溫度儲存。融合蛋白以尿素萃取法純化，一如前述(Liao等人，1995，Appl.Microbiol.Biotechnol.43：498-507)，之後再於4℃之溫度下，以50倍體積之TNE緩衝液(50mM Tris、50mM NaCl及1mM EDTA)進行隔夜透析，以完成融合蛋白之再摺疊。再摺疊後之蛋白接受SDS-PAGE分析，並以Bradford蛋白分析套組(Pierce)進行量化分析。分析結果顯示，大部分之再摺疊蛋白於非還原條件下為單體，意即融合蛋白可輕易再摺疊且未聚集成一體。
範例3
PCV2次單元疫苗免疫原性分析
小鼠以每周一次、連續3周之頻率，經皮下注射接種0.1ml PCV2次單元疫苗，疫苗含有40μg PE313 -ORF2-NESK或PE407 -ORF2-K3，以 及磷酸鋁(一可緩慢釋出融合蛋白之蛋白質吸收劑；10% v/v)與10μg皂素(一萃取自石鹼木(Quillaja saponaria)之佐劑)。控制組(安慰劑組)則注射不含融合蛋白之佐劑。所有小鼠均在最後一次接種後14天犧牲，並取其脾臟。脾細胞經分離後，於37℃及1μg/ml GolgiPlug(BD Pharmingen,San Diego,CA)存在之條件下，於6孔培養板中培養(108個細胞/2ml/孔)16小時，培養液中分別含有或不含有重組ORF2蛋白。刺激過後之脾細胞以FACScan緩衝液清洗，並以藻紅素共軛單株大鼠抗小鼠CD8a抗體及AF700共軛單株大鼠抗小鼠CD4抗體染色。接著再以Cytofix/Cytoperm套組進行細胞內細胞介素之染色，其操作方式悉依製造商(BD Pharmingen)之指示。細胞內之IFN-γ係以AF488共軛單株大鼠抗小鼠IFN-γ染色，以便量測免疫反應及細胞介素之含量。另以Gallios流動式細胞量測儀及Kaluza分析軟體(Beckman Coulter)進行流動式細胞量測分析。
第2A、2B圖分別顯示接種安慰劑(不含融合蛋白之佐劑)或融合蛋白之小鼠其脾細胞中CD8+及CD4+ IFN-γ T細胞之數量。脾細胞以ORF2刺激後，其CD4+及CD8+ T細胞所製造之IFN-γ係透過細胞內染色而以流動式細胞量測法加以偵測。柱狀圖顯示各組對ORF2具有專一性之IFN-γ+ CD4+ T細胞數(第2B圖)及IFN-γ+ CD8+ T細胞數(第2A圖)，其中灰色柱代表曾以ORF2胜肽刺激，黑色柱則代表未曾以ORF2胜肽刺激。結果顯示，曾接種PE313 -ORF2-NESK之小鼠其經由ORF2胜肽刺激而產生之ORF2專一性CD4+ IFN-γ+及CD8+ IFN-γ+ T細胞數均高於曾接種PE407 -ORF2-K3之小鼠組。
範例4
CSFV次單元疫苗免疫原性分析
在此使用相同之接種時程及劑量為小鼠接種CSFV次單元疫苗，其中次單元疫苗含有PE313 -E2-NESK或PE407 -E2-K3。脾細胞係依前述方式分離、培養，並以流動式細胞量測法分析，唯過程中另加入重組E2蛋白以刺激培養基中之脾細胞。
第3A、3B圖分別顯示接種安慰劑(不含融合蛋白之佐劑)或融合蛋白之小鼠其脾細胞中CD8+及CD4+ IFN-γ T細胞之數量。脾細胞以E2刺激後，其CD4+及CD8+ T細胞所製造之IFN-γ係透過細胞內染色而以流 動式細胞量測法加以偵測。柱狀圖顯示各組對E2具有專一性之IFN-γ+ CD4+ T細胞數(第3B圖)及IFN-γ+ CD8+ T細胞數(第3A圖)，其中灰色柱代表曾以E2胜肽刺激，黑色柱則代表未曾以E2胜肽刺激。結果顯示，曾接種PE313 -E2-NESK之小鼠其經由E2胜肽刺激而產生之E2專一性CD4+ IFN-γ+及CD8+ IFN-γ+ T細胞數均高於曾接種PE407 -E2-K3之小鼠組。
範例5
FMDV次單元疫苗免疫原性分析
在此使用相同之接種時程及劑量為小鼠接種FMDV次單元疫苗，其中次單元疫苗含有PE313 -VP1-3A-NESK或PE407 -VP1-3A-K3。脾細胞係依前述方式分離、培養，並以流動式細胞量測法分析，唯過程中另加入重組VP1-3A蛋白以刺激培養基中之脾細胞。
第4A、4B圖顯示接種安慰劑或融合蛋白之小鼠其脾細胞中CD8+及CD4+ IFN-γ T細胞之數量。脾細胞以VP1-3A刺激後，其CD4+及CD8+ T細胞所製造之IFN-γ係透過細胞內染色而以流動式細胞量測法加以偵測。柱狀圖顯示各組對VP1-3A具有專一性之IFN-γ+ CD4+ T細胞數(第4B圖)及IFN-γ+ CD8+ T細胞數(第4A圖)，其中灰色柱代表曾以VP1-3A胜肽刺激，黑色柱則代表未曾以VP1-3A胜肽刺激。結果顯示，曾接種PE313 -VP1-3A-NESK之小鼠其經由VP1-3A胜肽刺激而產生之VP1-3A專一性CD4+ IFN-γ+及CD8+ IFN-γ+ T細胞數均高於曾接種PE407 -VP1-3A-K3之小鼠組。
範例6
NDV次單元疫苗免疫原性分析
在此使用相同之接種時程及劑量為小鼠接種NDV次單元疫苗，其中次單元疫苗含有PE313 -FHN-NESK或PE407 -FHN-K3。脾細胞係依前述方式分離、培養，並以流動式細胞量測法分析，唯過程中另加入重組FHN蛋白以刺激培養基中之脾細胞。
第5A、5B圖顯示接種安慰劑或融合蛋白之小鼠其脾細胞中CD8+及CD4+ IFN-γ T細胞之數量。脾細胞以FHN刺激後，其CD4+及CD8+ T細胞所製造之IFN-γ係透過細胞內染色而以流動式細胞量測法加以偵測。柱狀圖顯示各組對FHN具有專一性之IFN-γ+ CD4+ T細胞數(第5B圖)及 IFN-γ+ CD8+ T細胞數(第5A圖)，其中灰色柱代表曾以FHN胜肽刺激，黑色柱則代表未曾以FHN胜肽刺激。結果顯示，曾接種PE313 -FHN-NESK之小鼠其經由FHN胜肽刺激而產生之FHN專一性CD4+ IFN-γ +及CD8+ IFN-γ + T細胞數均高於曾接種PE407 -FHN-K3之小鼠組。
範例7
加強抑制第16型人類乳突病毒E7蛋白所引發之腫瘤生長
融合蛋白PE407 -E7-K3、RAP1-PE268-313 -E7-K3及RAP1-PE268-313 -E7-NESK係以類似前述之方式表現及再摺疊。小鼠係以皮下注射之方式，以2 x 103 個TC-01細胞(其為帶有第16型HPV E7基因之小鼠肺部上皮細胞)攻毒，藉以誘發第16型HPV上皮癌。以TC-01細胞攻毒十二天後，小鼠以每周一次、連續3周之頻率，經皮下注射分別接種安慰劑(PBS)、PE407 -E7-K3(100mg/劑)、RAP1-PE268-313 -E7-K3(100μg/劑)或RAP1-PE268-313 -E7-NESK(100μg/劑)(均以AS04C(GlaxoSmithKline)為佐劑)(第6圖)。AS04C係一胞毒T淋巴球增強佐劑，包含MPL(單磷酯脂質A，一免疫增效劑)及磷酸鋁(一用於傳送抗原之蛋白質吸收劑)。「K3」一詞係指一包含KDEL之ER滯留序列。例如，K3可為胺基酸序列KDELKDELKDEL(序列編號第16號)。「NESK」一詞係指一包含核輸出信號及ER滯留序列之融合胜肽。在本發明之一實施例中，NESK為胺基酸序列LQKKLEELELAKDEL(序列編號第12號)。各組之腫瘤大小及無腫瘤動物數均加以記錄(第7及8圖)。疫苗PE407 -E7-K3、RAP1-PE268-313 -E7-K3及RAP1-PE268-313 -E7-NESK以AS04C為佐劑時，均對腫瘤生長具有顯著之抑制效果。就抑制腫瘤生長及提高無腫瘤動物百分比之效果而言，疫苗RAP1-PE268-313 -E7-NESK係優於RAP1-PE268-313 -E7-K3，且遠優於PE407 -E7-K3。
範例8
本範例製造出下列融合蛋白：PE313 -NES-抗原-K、PE1-252 -PE268-313 -NES-抗原-K、PE1-252 -PE280-313 -NES-抗原-K。此外，融合蛋白PE313 -抗原-NESK之PE部位Ia片段(PE1-252 )以RAP1部位3(序列編號第8號)、A2M最小(序列編號第9號)、HIV-Tat最小(序列編號第10號)或HSPs最小(序列編號第11號)取代後，分別製造出融合蛋白RAP1部位3-PE253-313 - 抗原-NESK、A2M-PE253-313 -抗原-NESK、Tat-PE253-313 -抗原-NESK及HSP-PE253-313 -抗原-NESK、RAP1部位3-PE268-313 -抗原-NESK、A2M-PE268-313 -抗原-NESK、Tat-PE268-313 -抗原-NESK及HSP-PE268-313 -抗原-NESK疫苗、RAP1部位3-PE280-313 -抗原-NESK、A2M-PE280-313 -抗原-NESK、Tat-PE280-313 -抗原-NESK及HSP-PE280-313 -抗原-NESK、RAP1部位3-PE253-313 -NES-抗原-K、A2M-PE253-313 -NES-抗原-K、Tat-PE253-313 -NES-抗原-K及HSP-PE253-313 -NES-抗原-K、RAP1部位3-PE268-313 -NES-抗原-K、A2M-PE268-313 -NES-抗原-K、Tat-PE268-313 -NES-抗原-K及HSP-PE268-313 -NES-抗原-K疫苗、RAP1部位3-PE280-313 -NES-抗原-K、A2M-PE280-313 -NES-抗原-K、Tat-PE280-313 -NES-抗原-K及HSP-PE280-313 -NES-抗原-K。上列疫苗能否增強細胞媒介免疫反應亦以類似前述之方法檢測。
表1顯示製造各種融合蛋白所用胜肽之序列編號。
綜合上述，分析結果已證明一融合蛋白若含有一位於N端之APC結合部位、一移位部位，並接續一病原體的一抗原，以及一位於羧基端之NESK融合胜肽，則其增強細胞媒介免疫反應之效果、抑制腫瘤生長之 效果及/或提高無腫瘤動物百分比之效果均優於羧基端無NESK融合胜肽之PE-融合蛋白。
本說明書雖已繪示及描述本發明之多種實施例，但熟習此項技術之人士仍可加以修改及改良。本發明並不限於本說明書所繪示之特定形態，所有未脫離本發明精神與範圍之修改方式均涵蓋在後附之申請專利範圍中。
以上實施例與範例之選擇與描述，係為解釋本發明之原理及其實際應用，使熟習此項技術之人士得以利用本發明及各種實施例，或依實際需要而以不同方式修改之。熟習此技術之人士可在不偏離本案精神與範圍之情況下，輕易想見多種替代實施例。因此，本發明之範圍取決於後附申請專利範圍，而非由以上敘述及其中所描述之實施範例加以定義。
本說明書所引用及討論之參考文獻可能包括專利、專利申請案及各種出版物。之所以引用及/或討論該些參考文獻乃為闡明本發明之內容，而非承認該些參考文獻為本發明之「先前技術」。本專利說明書所引用與討論之所有參考文獻，在此以引用之方式將其全文併入本文，且其併入之程度一如各參考文獻係分別以引用之方式併入本文。
<110> 生控基因疫苗股份有限公司
<120> 可用作誘發抗原專一性T細胞反應之免疫原性增強劑之融合蛋白
<140> TW 102144581
<141> 2013-12-05
<150> US/61733879
<151> 2012-12-05
<160> 29
<170> PatentIn version 3.5
<210> 1
<211> 252
<212> PRT
<213> 綠膿桿菌
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<212> PRT
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<212> PRT
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<212> PRT
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<211> 407
<212> PRT
<213> 綠膿桿菌
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<212> PRT
<213> 人類
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<212> PRT
<213> 人工序列
<220>
<223> A2M最小
<400> 9
<210> 10
<211> 24
<212> PRT
<213> 人工序列
<220>
<223> HIV-Tat最小
<400> 10
<210> 11
<211> 641
<212> PRT
<213> 人工序列
<220>
<223> HSPs最小
<400> 11
<210> 12
<211> 15
<212> PRT
<213> 人工序列
<220>
<223> NESK
<400> 12
<210> 13
<211> 11
<212> PRT
<213> 人工序列
<220>
<223> NES一致序列
<220>
<221> misc_feature
<222> (2)..(3)
<223> Xaa可為任一種天然胺基酸
<220>
<221> misc_feature
<222> (6)..(7)
<223> Xaa可為任一種天然胺基酸
<220>
<221> misc_feature
<222> (9)..(9)
<223> Xaa可為任一種天然胺基酸
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<221> misc_feature
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<223> Xaa可為任一種天然胺基酸
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<212> PRT
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<223> NES
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<223> 內質網(ER)滯留序列
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<212> PRT
<213> 人工序列
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<223> K3
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<223> 截短之CSFV E2
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<223> FMDV VP1胜肽
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<223> FMDV 3A胜肽
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<223> FMDV VP1-3A胜肽
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<223> NDV F胜肽
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<223> NDV HN胜肽
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<223> NDV FHN胜肽
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Claims (20)

  1. 一種融合蛋白，包含：(a)一抗原呈現細胞(APC)結合部位或一CD91受體結合部位，其位於融合蛋白之N端；(b)一移位胜肽，其就長度而言具有34-112個胺基酸殘基，且包含一與序列編號第4、2、3或6號至少90%相同之胺基酸序列，移位胜肽位於APC結合部位或CD91受體結合部位之C端；(c)一病原體的一抗原，其位於移位胜肽之C端；(d)一核輸出信號，其包含序列編號第13號之胺基酸序列；及(e)一內質網(ER)滯留序列，其位於融合蛋白之C端。
  2. 如申請專利範圍第1項之融合蛋白，其中APC結合部位或CD91受體結合部位係一多胜肽，多胜肽包含一與下列各項所組成之群組中選出之序列至少90%相同之胺基酸序列：序列編號第1及8至11號。
  3. 如申請專利範圍第1項之融合蛋白，其中核輸出信號包含序列編號第14號之胺基酸序列。
  4. 如申請專利範圍第1項之融合蛋白，其中內質網滯留序列包含序列編號第15號之胺基酸序列。
  5. 如申請專利範圍第1項之融合蛋白，其中核輸出信號係位於移位胜肽與抗原之間。
  6. 如申請專利範圍第1項之融合蛋白，其中核輸出信號係位於抗原與內質網滯留序列之間。
  7. 如申請專利範圍第1項之融合蛋白，其中核輸出信號及ER滯留序列形成一融合胜肽，融合胜肽包含一與序列編號第12號至少90%相同之胺基酸序列。
  8. 如申請專利範圍第1項之融合蛋白，其中移位胜肽就長度而言具有34-61個胺基酸殘基。
  9. 如申請專利範圍第1項之融合蛋白，其中APC結合部位或CD91受體結合部位完全不含綠膿桿菌外毒素A(PE)結合部位I之胺基酸序列。
  10. 如申請專利範圍第9項之融合蛋白，其中APC結合部位或CD91受體結合部位包含序列編號第8號之胺基酸序列。
  11. 如申請專利範圍第1項之融合蛋白，其中APC結合部位或CD91受體結合部位之胺基酸序列為序列編號第1號。
  12. 如申請專利範圍第11項之融合蛋白，其中移位胜肽就長度而言具有34-61個胺基酸殘基。
  13. 如申請專利範圍第12項之融合蛋白，其中核輸出信號及ER滯留序列形成一融合胜肽，融合胜肽之胺基酸序列與序列編號第12號至少90%相同。
  14. 如申請專利範圍第11項之融合蛋白，其中移位胜肽就長度而言具有34-46個胺基酸殘基。
  15. 如申請專利範圍第10項之融合蛋白，其中移位胜肽就長度而言具有34-61個胺基酸殘基。
  16. 如申請專利範圍第15項之融合蛋白，其中核輸出信號及ER滯留序列形成一融合胜肽，融合胜肽之胺基酸序列與序列編號第12號至少90%相同。
  17. 如申請專利範圍第1項之融合蛋白，其中抗原係一病原體之兩個以上抗原胜肽之融合抗原。
  18. 如申請專利範圍第1項之融合蛋白，其實質上由下列各項組成：(a)一抗原呈現細胞(APC)結合部位或一CD91受體結合部位，其位於融合蛋白之N端；(b)一移位胜肽，其就長度而言具有34-112個胺基酸殘基，且包含一與序列編號第4、2、3或6號至少90%相同之胺基酸序列，移位胜肽位於APC結合部位或CD91受體結合部位之C端；(c)一病原體的一抗原，其位於移位胜肽之C端；(d)一核輸出信號，其包含序列編號第13號之胺基酸序列；及(e)一內質網(ER)滯留序列，其位於融合蛋白之C端。
  19. 一種疫苗組合物，包含如申請專利範圍第1至18項任一項之融合蛋白及一佐劑。
  20. 一種如申請專利範圍第1至18項任一項之融合蛋白的應用，係用於製備一種誘發並增強對病原體抗原具有專一性之T細胞反應的藥物。
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Families Citing this family (24)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11246915B2 (en) 2010-09-15 2022-02-15 Applied Molecular Transport Inc. Cholix toxin-derived fusion molecules for oral delivery of biologically active cargo
CN109553688A (zh) * 2012-12-05 2019-04-02 生控基因疫苗股份有限公司 用作诱发抗原特异性t细胞反应的免疫原性增强剂的融合蛋白
CN107249629A (zh) * 2015-02-26 2017-10-13 生控基因疫苗股份有限公司 包含免疫原性蛋白质及组合佐剂并用以诱发抗原特异性t细胞反应的疫苗组合物
WO2016196383A1 (en) * 2015-06-01 2016-12-08 Reber Genetics Co., Ltd. Vaccine compositions against porcine reproductive and respiratory syndrome and porcine circovirus associated diseases
JP2018521983A (ja) 2015-07-16 2018-08-09 バイオカイン セラピューティックス リミテッド がんを治療するための組成物および方法
KR102153303B1 (ko) * 2015-11-23 2020-09-09 뵈링거 잉겔하임 애니멀 헬스 유에스에이 인코포레이티드 Fmdv 및 e2 융합 단백질 및 이의 용도
WO2018052854A1 (en) * 2016-09-19 2018-03-22 Thevax Genetics Vaccine Co., Ltd. Hepatitis b therapeutic vaccines
US11186618B2 (en) * 2016-09-21 2021-11-30 The Research Foundation For Microbial Diseases Of Osaka University Dendritic-cell-targeted peptide, fusion peptide utilizing said peptide, and vaccine utilizing said fusion peptide
CN107991481A (zh) * 2016-10-27 2018-05-04 武汉科前生物股份有限公司 一种检测猪伪狂犬病毒和口蹄疫病毒的二联阻断elisa抗体检测试剂盒及其应用
CN107937354A (zh) * 2017-11-10 2018-04-20 南京天邦生物科技有限公司 2型猪圆环病毒及其应用
PL3762009T3 (pl) 2018-03-08 2022-09-12 Applied Molecular Transport Inc. Pochodzące z toksyny konstrukty dostarczające do dostarczania doustnego
CN109134667A (zh) * 2018-09-19 2019-01-04 天康生物股份有限公司 融合蛋白及其制备方法、应用、表达系统和疫苗
JP2022512976A (ja) 2018-11-07 2022-02-07 アプライド モレキュラー トランスポート インコーポレイテッド 異種ペイロードの経口送達のためのコリックス由来担体
CN109593136B (zh) * 2018-12-26 2021-03-19 天康生物股份有限公司 禽副粘病毒融合蛋白及其制备方法、应用和用于鸽子的apmv疫苗
KR20220012256A (ko) * 2019-05-24 2022-02-03 프로비바 테라퓨틱스 (홍콩) 리미티드 Il-2 조성물 및 이의 사용 방법
CN110051832B (zh) * 2019-05-31 2020-11-10 四川农业大学 一种犬恶丝虫病疫苗
CN110452928A (zh) * 2019-08-13 2019-11-15 成都天邦生物制品有限公司 一株pk-15稳定细胞株的构建及应用
AU2020329290A1 (en) * 2019-08-13 2022-03-24 Elpis Biopharmaceuticals Engineered interleukin-2 receptor beta agonists
CN110669142B (zh) * 2019-09-30 2021-10-12 湖南农业大学 融合rgd的猪圆环病毒2型病毒样颗粒、突变型感染性克隆及其制备方法和应用
US20230173049A1 (en) * 2019-12-31 2023-06-08 The Johns Hopkins University Fusion proteins and methods of use thereof
CN111548395A (zh) * 2020-05-25 2020-08-18 中国农业科学院兰州兽医研究所 一种口蹄疫病毒二价多表位重组病毒样颗粒及其应用
KR102507298B1 (ko) 2020-12-21 2023-03-08 충남대학교산학협력단 면역증강을 위한 조성물
CN112812171B (zh) * 2021-01-22 2022-04-08 浙江辉肽生命健康科技有限公司 具有氨基酸结构vvrkplnkegkkp的生物活性肽及其制备方法和应用
CN114539429B (zh) * 2022-03-24 2022-07-29 上海普铭生物科技有限公司 融合蛋白组合物及其应用

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090214570A1 (en) * 2005-05-18 2009-08-27 Trinity Biosystems, Inc. Methods and compositions for immunizing against chlamydia infection
US20120213811A1 (en) * 2008-01-31 2012-08-23 Healthbanks Usa Inc. Chimeric hiv fusion proteins as vaccines

Family Cites Families (37)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US843505A (en) * 1905-10-09 1907-02-05 Alexandre Leonard Tombelaine Safety closing device for miner's lamps.
PT651805E (pt) * 1992-07-17 2007-02-28 Dana Farber Cancer Inst Inc Método de ligação intracelular de moléculas-alvo
US5712149A (en) * 1995-02-03 1998-01-27 Cell Genesys, Inc. Chimeric receptor molecules for delivery of co-stimulatory signals
US6451592B1 (en) * 1996-04-05 2002-09-17 Chiron Corporation Recombinant alphavirus-based vectors with reduced inhibition of cellular macromolecular synthesis
US20050119470A1 (en) * 1996-06-06 2005-06-02 Muthiah Manoharan Conjugated oligomeric compounds and their use in gene modulation
DE19651443A1 (de) * 1996-12-11 1998-06-18 Hoechst Ag Selbstverstärkende, pharmakologisch kontrollierbare Expressionssysteme
US7314632B1 (en) 1997-07-11 2008-01-01 The United States Of America As Represented By The Secretary Of The Department Of Health And Human Services Pseudomonas exotoxin A-like chimeric immunogens
DK1000163T3 (da) * 1997-07-11 2006-02-06 Us Gov Health & Human Serv Pseudomonas-exotoksin A-lignende kimære immunogener
ATE359084T1 (de) * 1998-02-20 2007-05-15 Univ Miami Modifizierter hitzeschockprotein/peptidantigen komplex
WO1999045127A2 (en) * 1998-03-06 1999-09-10 Oxford Biomedica (Uk) Limited Enhanced prodrug activation
US20030215421A1 (en) * 1999-07-21 2003-11-20 Mcdonald John R. Methods and compositions for treating secondary tissue damage and other inflammatory conditions and disorders
US20020061848A1 (en) * 2000-07-20 2002-05-23 Ajay Bhatia Compounds and methods for treatment and diagnosis of chlamydial infection
WO2000049041A1 (fr) * 1999-02-19 2000-08-24 Sumitomo Electric Industries, Ltd. Preparations proteiques
JP4637368B2 (ja) * 1999-04-14 2011-02-23 ノバルティス バクシンズ アンド ダイアグノスティックス，インコーポレーテッド アルファウイルスに基づくベクター系を利用する免疫応答を生成するための組成物および方法
EP1222289B1 (en) * 1999-10-20 2008-04-16 The Johns Hopkins University School Of Medicine Chimeric immunogenic compositions and nucleic acids encoding them
US8128922B2 (en) * 1999-10-20 2012-03-06 Johns Hopkins University Superior molecular vaccine linking the translocation domain of a bacterial toxin to an antigen
US7030219B2 (en) * 2000-04-28 2006-04-18 Johns Hopkins University B7-DC, Dendritic cell co-stimulatory molecules
CA2453528C (en) * 2001-08-01 2011-07-26 Cellomics, Inc. Novel fusion proteins and assays for molecular binding
US7235631B2 (en) * 2002-02-07 2007-06-26 Mayo Foundation For Medical Education And Research ICOS mutants
WO2004087754A1 (en) * 2003-04-03 2004-10-14 Yissum Research Development Company Of The Hebrew University Of Jerusalem Broad-spectrum in-vivo effective superantigen toxin antagonists based on the interaction between cd28 and the superantigen and uses thereof
IL164799A0 (en) * 2002-04-25 2005-12-18 Univ Connecticut Using heat shock proteins to improve the therapeutic benefit of a non-vaccine treatment modality
CA2515123A1 (en) * 2003-02-04 2004-08-19 University Of Connecticut Health Center Immunogenic cd91 ligand-antigenic molecule complexes and fusion proteins
EP1594437A2 (en) * 2003-02-04 2005-11-16 University of Connecticut Health Center Immunogenic cd91 ligand-antigenic molecule complexes and fusion proteins
US7595054B2 (en) * 2003-06-09 2009-09-29 Healthbanks Biotech Co., Ltd. Fusion antigen used as vaccine
US7335361B2 (en) 2003-06-09 2008-02-26 Animal Technology Institute Taiwan Fusion antigen used as vaccine
EP1756147A2 (en) * 2004-06-01 2007-02-28 Innogenetics N.V. Peptides for inducing a ctl and/or htl response to hepatitis c virus
JP4474264B2 (ja) * 2004-08-20 2010-06-02 生寶生物科技股▲ふん▼有限公司 子宮頸癌抑制の融合蛋白
WO2006135428A2 (en) * 2004-10-04 2006-12-21 Trinity Biosystems, Inc. Methods and compositions for inducing an immune response against multiple antigens
US7465455B2 (en) 2006-07-05 2008-12-16 Healthbanks Biotech Co., Ltd. Fusion protein of porcine reproductive and respiratory syndrome virus as PRRS vaccine
WO2008047243A2 (en) * 2006-08-29 2008-04-24 Forhumantech. Co., Ltd. Pharmaceutical composition for suppression of apoptosis and method for delivering the same
MX2009002893A (es) * 2006-09-18 2009-07-10 Raptor Pharmaceutical Inc Tratamiento de trastornos hepaticos mediante la administracion de conjugados de la proteina asociada al receptor (rap).
US7887801B2 (en) * 2007-07-13 2011-02-15 Topotarget Germany Ag Optimized DNA and protein sequence of an antibody to improve quality and yield of bacterially expressed antibody fusion proteins
WO2009036349A1 (en) * 2007-09-12 2009-03-19 Anaphore, Inc. Hsp70-based treatment for autoimmune diseases
EP2078726A1 (en) * 2008-01-09 2009-07-15 Vision 7 GmbH Secretable HIV entry inhibitory peptides for therapy of HIV infection
WO2010040023A2 (en) * 2008-10-03 2010-04-08 Government Of The United States Of America, As Represented By The Secretary, Department Of Health & Human Services Methods and compositions for protein delivery
PL391627A1 (pl) * 2010-06-25 2012-01-02 Adamed Spółka Z Ograniczoną Odpowiedzialnością Przeciwnowotworowe białko fuzyjne
CN109553688A (zh) * 2012-12-05 2019-04-02 生控基因疫苗股份有限公司 用作诱发抗原特异性t细胞反应的免疫原性增强剂的融合蛋白

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090214570A1 (en) * 2005-05-18 2009-08-27 Trinity Biosystems, Inc. Methods and compositions for immunizing against chlamydia infection
US20120213811A1 (en) * 2008-01-31 2012-08-23 Healthbanks Usa Inc. Chimeric hiv fusion proteins as vaccines

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
M Kakar et al. "Optimizing the protein switch: Altering nuclear import and export signals, and ligand binding domain", Journal of Controlled Release, 2007, Vol. 120, p. 220~232. *

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