JP6997619B2 - 液性免疫の抑制または増進のための、vistaアゴニスト及びvistaアンタゴニストの使用 - Google Patents
液性免疫の抑制または増進のための、vistaアゴニスト及びvistaアンタゴニストの使用 Download PDFInfo
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Description
本出願は、2014年6月11日出願の米国仮出願第62/010,736号に対する優先権を主張する。この出願は、参照によってその全体が本明細書に組み込まれる。
本発明は、必要とする対象における、VISTAが媒介する液性免疫阻害の阻害方法または無効化方法を提供し、当該方法は、例えば、B細胞免疫が抑制される癌または感染性疾患状態を有する対象における、VISTAアンタゴニストの単独投与、または別の免疫アゴニストと関連させた投与を含む。
本発明の詳細を記載する前に、下記の定義を提供する。
実施例において使用した材料及び方法
マウス.7週齢の雄性C57BL/6(B6)マウスをNational Institutes of Health(メリーランド州、ベゼスダ)から購入し、Dartmouth Medical School Animal facilityにおいて飼育し、約8~10週齢となったところで使用した。B6へと完全に戻し交配させたiNOSノックアウトマウスの繁殖ペアは、Jackson Labs(メイン州、バーハーバー)から入手し、当該繁殖ペアは、元は、以前に説明された(Laubachら 1995)ようにして得られたものである。また、B6へと戻し交配させたものであり、報告に沿って得たVISTAノックアウト繁殖ペアは、から入手した。
この実施例は、図1の実験に関するものである。こうした実験では、脾臓細胞によるVISTAの発現は、インビボでLP-BM5が感染することで増加することが示された。図1(A)は、フローサイトメトリー分析を含むものであり、非感染野生型B6マウスまたは感染後5週間もしくは8.5週間の野生型B6マウスに由来する脾臓細胞のVISTA発現パターンを示している。割合は、VISTAの発現に陽性である細胞を示し、イタリック体である値は、VISTAに陽性である細胞のMFIを示す。図1(B)は、脾臓細胞表面のVISTAの発現であり、下記の群のCD11b+細胞のVISTAを示す。すなわち、記載の非感染マウス系または感染マウス系から、その後に精製することなく得た細胞と、5wpi-LP-BM5マウスから精製したLy6C+CD11b+脾臓細胞と、である。示されている結果パターンは、2回の追加実験を代表するものである。
この実施例は、図2の実験に関するものである。こうした実験は、Ly6C+CD11b+で精製した脾臓細胞が、B細胞増殖と、T細胞増殖とをインビトロで抑制するために、異なる機構的必要要件を有していることを示している。図2(A及びB)は、レスポンダー対抑制因子(R:S)の比を3:1とし、ナイーブB6レスポンダー脾臓細胞を、a-VISTA及びLNMMAで事前に処理したLy6C+CD11b+MDSCと混合し、(A)a-CD40及びIL-4、または(B)ConAで3日間刺激した実験の結果を示す。
この実施例は、図3の実験に関するものである。こうした実験では、レスポンダー対抑制因子(R:S)の比を3:1として、ナイーブVISTA-/-レスポンダー脾臓細胞を、α-VISTAで事前処理したLy6C+CD11b+MDSCと混合し、a-CD40及びIL-4で3日間刺激した。示されている結果パターンは、3回の追加実験を代表するものである。*はp<0.05、**はP<0.01、***はP<0.001、有意差なし(ns.)は、P>0.05であることを示す。
この実施例は、図4の実験に関するものである。こうした実験では、LP-BM5に感染後、5週間が経過した、記載の種のマウスから精製した単球性MDSC(Ly6C+CD11b+)が、ナイーブVISTA-/-脾臓細胞の増殖を異なるパターンで抑制していることを示しており、これはフローサイトメトリーによって評価したものである。CFSEで標識したVISTA-/-レスポンダー細胞を、ct-CD40+IL4の存在下(+)または非存在下(-)かつ、単球性MDSCの存在下または非存在下で3日間培養した。すなわち、(a)(-)と、(b)(+)(灰色の影付き曲線)と、(c)(+)、野生型のMDSC(黒色の開曲線)と、(d)(+)、DNMMAで処理した野生型のMDSC(黒色の開曲線)と、(e)(+)、LNMMAで処理した野生型のMDSCと、(f)(+)、INOS-/-MDSC(点線の曲線)と、(g)(+)と、(h)(+)、VISTA-/-MDSC及び野生型のMDSCの対比と、である。4日間のインキュベーションの終了時点で、すべての培養物に由来する細胞を、ct-CD19と蛍光色素が結合したmAbで染色し、FACSによってCFSE希釈も評価した。CFSE標識したVISTA-/-レスポンダーは、%=最初の増殖ピーク中の細胞の割合eであり、イタリック体の数=CFSE+細胞のMFIである。示されている結果パターンは、1回の追加実験を代表するものである。
この実施例は、図5の実験に関するものである。こうした実験では、B細胞増殖の単球性の抑制が、2つの主な機構であるiNOS/NO及びVISTAに依存することが示された。5Aの実験では、LP-BM5に感染した5w.p.の野生型マウスまたはVISTA-/-マウスから精製したLy6C+CD11b+MDSCをct-VISTA遮断抗体または対照HIg、LNMMAまたは対照DNMMAで事前に処理してから、VISTA-/-レスポンダー細胞、ct-CD40、及びIL-4と共に3日間培養した。5(B)の実験では、パネル(A)に示すものと同一の処理レジメンに、ct-VISTA及びLNMMAの両方を含む組み合わせレジメンを含めて、野生型のLy6C+CD11b+MDSCを事前処理した。レスポンダーが取り込んだ3Hチミジンのcpmによって、(A)及び(B)の両方の抑制%を計算した。示されている結果パターンは、2回の追加実験(A)及び1回の追加実験(B)を代表するものである。*は、P<0.05、**は、P<0.01、有意差なし(ns.)はP>0.05であることを示す。
LP-BM5レトロウイルスによる感染後にMDSCが増殖するため、VISTAの細胞表面発現は、感染後(p.i.)に初めて評価した。5w.p.i.の時点で、VISTA+脾臓細胞の割合は増加していなかったが、VISTAのMFIは増加し、陽性ピークの形状が変化した(図1A)。この変化は、非感染B6マウスにおいて、CD4 T細胞がVISTAを優位に発現するという報告(Wangら 2011)と、インビボでの刺激に際して増殖するCD11b+VISTA+細胞などの細胞型のさらなる存在と、の両方に一致するものであり、本明細書では、我々が報告したLP-BM5感染が、MDSCを増殖させている(我々の参考文献)。8.5w.p.i.の時点で、VISTAの発現増加は、より明白であった(図1A)。非感染B6マウスと、5w.p.i.の野生型B6マウス、VISTA-/-B6マウス、及びINOS-/-B6マウスと、に由来する細胞による、VISTA及びCD11bの同時発現をさらに染色して対比させることで、抗VISTAmAbの特異性を確認し、高度に濃縮された単球性MDSC集団における、VISTAの発現増加を実証した(図1B)(以前にLy6Cも発現していることを示した(我々の参考文献を参照のこと。本明細書では、データ未掲載)。さらに、Wangら(2011)による研究によって予測されていたように、他の主なVISTA+集団は、CD4 T細胞コンパートメントであった(データ未掲載)。
B細胞応答性に対するVISTAの作用、具体的には、MSDCによって媒介される、B細胞応答性に対するVISTAの抑制作用に基づき、VISTAアンタゴニストを単独または他の免疫活性化因子もしくは免疫アゴニストと関連させて使用してよく、具体的には、B細胞応答性の増加が治療的に望ましい状態を治療または防止するために、アゴニストである抗CD40抗体またはCD40LポリペプチドなどのCD40アゴニストなどの、液性免疫を増進するものと関連させて使用してよい。そのような状態には、具体的には、B細胞が応答する癌が含まれ、例えば、抗体応答が疾患病理に関与するか、または予防免疫もしくは治療免疫に関与するものである。そのような癌の状態の例には、限定はされないが、カルシノーマ、リンパ腫、芽細胞腫、肉腫(脂肪肉腫を含む)、神経内分泌腫瘍、中皮腫、シュワン腫(schwanoma)、髄膜腫、腺癌、黒色腫、卵巣癌、乳癌、肺癌、脳癌、消化器癌、及び白血病またはリンパ系腫瘍が含まれる。
マウスVISTA配列識別番号:1
1 mgvpavpeas sprwgtllla iflaasrglv aafkvttpys lyvcpegqna tltcrilgpv
61 skghdvtiyk twylssrgev qmckehrpir nftlqhlqhh gshlkanash dqpqkhglel
121 asdhhgnfsi tlrnvtprds glycclviel knhhpeqrfy gsmelqvqag kgsgstcmas
181 neqdsdsita aalatgaciv gilclplill lvykqrqvas hrraqelvrm dsntqgienp
241 gfettppfqg mpeaktrppl syvaqrqpse sgryllsdps tplsppgpgd vffpsldpvp
301 dspnseai
マウスVISTA配列識別番号:2
1 mgvpavpeas sprwgtllla iflaasrglv aafkvttpys lyvcpegqna tltcrilgpv
61 skghdvtiyk twylssrgev qmckehrpir nftlqhlqhh gshlkanash dqpqkhglel
121 asdhhgnfsi tlrnvtprds glycclviel knhhpeqrfy gsmelqvqag kgsgstcmas
181 neqdsdsita aalatgaciv gilclplill lvykqrqvas hrraqelvrm dssntqgien
241 pgfettppfq gmpeaktrpp lsyvaqrqps esgryllsdp stplsppgpg dvffpsldpv
301 pdspnseai
マウスVISTA配列識別番号:3
1 mgvpavpeas sprwgtllla iflaasrglv aafkvttpys lyvcpegqna tltcrilgpv
61 skghdvtiyk twylssrgev qmckehrpir nftlqhlqhh gshlkanash dqpqkhglel
121 asdhhgnfsi tlrnvtprds glycclviel knhhpeqrfy gsmelqvqag kgsgstcmas
181 neqdsdsita aalatgaciv gilclplill lvykqrqvas hrraqelvrm dssntqgien
241 pgfettppfq gmpeaktrpp lsyvaqrqps esgryllsdp stplsppgpg dvffpsldpv
301 pdspnseai
ヒトVISTA配列識別番号:4
1 mgvptaleag swrwgsllfa lflaaslgpv aafkvatpys lyvcpegqnv tltcrllgpv
61 dkghdvtfyk twyrssrgev qtcserrpir nltfqdlhlh hgghqaants hdlaqrhgle
121 sasdhhgnfs itmrnltlld sglycclvve irhhhsehrv hgamelqvqt gkdapsncvv
181 ypsssqdsen itaaalatga civgilclpl illlvykqrq aasnrraqel vrmdsniqgi
241 enpgfeaspp aqgipeakvr hplsyvaqrq psesgrhlls epstplsppg pgdvffpsld
301 pvpdspnfev i
参考文献
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2.Bowen,J.L.,and J.K.Olson.2009.Innate immune CD11b+Gr-1+ cells,suppressor cells,affect the immune response during Theiler’s virus-induced demyelinating disease.J.Immunol.183:6971-6980.
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4.Chen,S.,S.M.F.Akbar,M.Abe,Y.Hiasa,and M.Onji.2011.Immunosuppressive functions of hepatic myeloid-derived suppressor cells of normal mice and in a murine model of chronic hepatitis B virus.Clin.Exp.Immunol.166:134-142.
5.Day C.L.,D.E.aufmann,P.Kiepiela,J.A.Brown,E.S.Moodley,S.Reddy,E.W.Mackey,J.D.Miller,A.J.Leslie,C.DePierres,Z.Mncube,J.Duraiswamy,B.Zhu,Q.Eichbaum,M.Altfeld,E.J.Wherry,E.M.Coovadia,P.J.R.Goulder,P.Klenerman,R.Ahmed,G.J.Freeman,and B.D.Walker.2006.PD-1 expression on HIV-specific T cells is associated with T-cell exhaustion and disease progression.Nature.443:350-354.
6.Gabrilovich,D.I.,and S.Nagaraj.2009.Myeloid-derived suppressor cells as regulators of the immune system.Nat.Rev.Immunol.9:162-174.
7.Garg,A.and S.A.Spector.2014.HIV type 1 gp120-induced expansion of myeloid derived suppressor cells is dependent on Interleukin 6 and suppresses immunity.J.Inf.Dis.209(3):441-451.
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Claims (6)
- VISTAアンタゴニストである抗VISTA抗体の有効量の、以下のいずれかに使用するための医薬の製造における使用:
(i) 必要とする対象における、VISTAが媒介する液性免疫阻害の阻害または無効化;
(ii) 必要とする対象における、抗原特異的な抗体応答を含む、B細胞増殖またはB細胞応答の増加;
(iii) 必要とする対象における、抗原または治療抗体に対して誘発される液性免疫応答の促進;あるいは
(iv) 治療ワクチンまたは予防ワクチンによって誘発される液性免疫応答の促進、
ここで、前記(i)~(iv)における使用は、iNOS/NO阻害物質である別の医薬の使用をさらに含む、前記使用。 - 前記対象が、癌または感染性疾患状態を有するか、あるいはB細胞増殖またはB細胞応答の増加を必要とする、請求項1に記載の使用。
- 腫瘍抗原、自己抗原、アレルゲン、または感染病原体の抗原から選択される抗原の使用をさらに含む、請求項1または2に記載の使用。
- 前記iNOS/NO阻害物質が、一酸化窒素もしくは一酸化窒素の合成を阻害する化合物、アルギニン誘導体、NG-ニトロ-L-アルギニンメチルエステル、NG-エチル-L-アルギニン、N-イミノエチル-L-アルギニン、L-NG-メチルアルギニン、NG-ニトロ-L-アルギニン、NG-ニトロ-L-アルギニンメチルエステル、ロバスタチン、フェニル酢酸のナトリウム塩(NaPA)、FPT阻害物質II、N-アセチルシステイン(NAC)、cAMP、L-NMA、またはLNMMAから選択される、請求項1、2、または3に記載の使用。
- 前記対象が:(i)ウイルス、細菌、または寄生虫によって引き起こされる感染性疾患を有するか;(ii)進行癌、転移性癌、及び/または固形腫瘍を有するか;あるいは(iii)肉腫、黒色腫、卵巣癌、肺癌、リンパ腫、白血病、または骨髄性の癌を有する、請求項1、2、または3に記載の使用。
- PD-1アンタゴニストまたはPD-L1アンタゴニストの使用をさらに含む、請求項1に記載の使用。
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JP7026169B2 (ja) | 2022-02-25 |
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