JP2015120696A - アルファ2,3−およびアルファ2,6−シアリル化を含む組換えfsh - Google Patents
アルファ2,3−およびアルファ2,6−シアリル化を含む組換えfsh Download PDFInfo
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- JP2015120696A JP2015120696A JP2014262056A JP2014262056A JP2015120696A JP 2015120696 A JP2015120696 A JP 2015120696A JP 2014262056 A JP2014262056 A JP 2014262056A JP 2014262056 A JP2014262056 A JP 2014262056A JP 2015120696 A JP2015120696 A JP 2015120696A
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- sialylation
- fsh
- rfsh
- sialic acid
- recombinant fsh
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Classifications
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- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
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- C07K14/575—Hormones
- C07K14/59—Follicle-stimulating hormone [FSH]; Chorionic gonadotropins, e.g.hCG [human chorionic gonadotropin]; Luteinising hormone [LH]; Thyroid-stimulating hormone [TSH]
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- A61K38/22—Hormones
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Abstract
Description
vivo効力は減少しており、より短期間の血漿半減期を示した(D’Antonio
et al.1999)。排卵周期の後期中に循環するより塩基性のFSHが、エストラジオールレベルの増加により引き起こされる下垂体前葉でのα2,3シアリル−トランスフェラーゼの下方制御に起因することも報告されている(Damian−Matsumara et al.1999.Ulloa−Aguirre et al.2001)。α2,6シアリル−トランスフェラーゼに関する結果は報告されていない。
れているため、直接的に比較することができない。下垂体/血清/尿のFSHは、α2,3およびα2,6結合のシアル酸を両方とも含有しており、前者が圧倒的に多い。しかしながら、CHO細胞に由来する組換え体は、α2,3のみを含有する(Kagawa et al、1988、Takeuchi et al、1988、Svensson et al.、1990)。これは、後者のシアル酸含有量が全体的に低いことに加えて、天然産物と現行の組換え産物との間にある別の相異である。
出されるアシアロ糖タンパク質(ASGP)受容体により媒介されることが見出された(実施例9)。これは、この受容体の別の基質を過剰量で使用して、ASGP受容体を一時的にブロックすることにより実証された。受容体をアシアロフェツインでブロックすると、この高度にシアリル化された物質の予測通りのクリアランスが回復した(図9)。これは、ブロックが克服されて、α2,6結合の高度にシアリル化されたrFSHの速やかなクリアランスが再開されるまで、数時間維持された。
〜14mol/mol、例えば12mol/mol〜14mol/mol、例えば12mol/mol〜13mol/molであってもよい。本発明のrFSHは、ヒト細胞系で産生または発現させてもよい。
せてもよい。rFSHは、内因性シアリルトランスフェラーゼ活性によりもたらされるα2,6結合シアル酸(α2,6シアリル化)を含んでいてもよい。
公知の任意の方法により任意の適切な供給源(例えば、尿)から精製することができる。組換えLHを発現および精製する方法は、当技術分野で周知である。
することができる。
ヒトFSH
FSHアルファポリペプチドの遺伝子のコード領域を、FiddesおよびGoodman(1981年)に従って使用した。この配列は、AH007338として寄託されており、構築時には、このタンパク質配列の他の変異体は存在しなかった。この配列は、本明細書中では配列番号1として参照される。
α2,3−シアリルトランスフェラーゼ:ベータガラクトシドアルファ−2,3−シアリルトランスフェラーゼ4(α2,3−シアリルトランスフェラーゼ、ST3GAL4)の遺伝子のコード領域を、KitagawaおよびPaulson(1994年)に従って使用した。この配列は、L23767として寄託されており、本明細書中では配列番号3として参照される。
FSH発現ベクターの構築
FSHアルファポリペプチド(AH007338、配列番号1)のコード配列、およびFSHベータポリペプチド(NM_003032、配列番号2)を、プライマーの組合せFSHa−fwおよびFSHa−revならびにFSHb−fwおよびFSHb−recをそれぞれ使用して、PCRにより増幅した。
FSHa-fw 5'-CCAGGATCCGCCACCATGGATTACTACAGAAAAATATGC-3'
FSHa-rev 5'-GGATGGCTAGCTTAAGATTTGTGATAATAAC-3'
FSHb-fw 5'-CCAGGCGCGCCACCATGAAGACACTCCAGTTTTTC-3'
FSHb-rev 5'-CCGGGTTAACTTATTATTCTTTCATTTCACCAAAGG-3'
号1および配列番号2の通りの正確な配列を含有していた。プラスミドpFSH A+B#17をトランスフェクション用に選択した(図1)。
ST3発現ベクターの構築
ベータガラクトシドアルファ−2,3−シアリルトランスフェラーゼ4(ST3、L23767、配列番号3)のコード配列を、プライマーの組合せ2,3STfwおよび2,3STrevを使用してPCRにより増幅した。
2,3STfw 5'-CCAGGATCCGCCACCATGTGTCCTGCAGGCTGGAAGC-3'
2,3STrev 5'-TTTTTTTCTTAAGTCAGAAGGACGTGAGGTTCTTG-3'
ST6発現ベクターの構築
ベータガラクトサミドアルファ−2,6−シアリルトランスフェラーゼ1(ST6、NM_003032、配列番号4)のコード配列を、プライマーの組合せ2,6STfwおよび2,6STrevを使用してPCRにより増幅した。
2,6STfw 5'-CCAGGATCCGCCACCATGATTCACACCAACCTGAAG-3'
2,6STrev 5'-TTTTTTTCTTAAGTTAGCAGTGAATGGTCCGG-3'
PER.C6細胞におけるpFSH A+Bの安定的発現。クローンのトランスフェクション単離およびスクリーニング。
FSHを産生するPer.C6クローンを、単一プラスミドからFSHの両ポリペプチド鎖を発現させることにより産生した(実施例1を参照)。
料の等電点特性を測定した(実施例6)。代表的な試料を図4に示す。IEFからの情報を使用して、代謝クリアランス率を分析するためのクローンを選択した(実施例9)。十分な生産性および品質を有するクローン(005、104、179、223、144)を選択して、シアリルトランスフェラーゼで操作した。
シアリル化のレベルは、α2,3−またはα2,6−シアリルトランスフェラーゼを過剰発現する細胞で増加する。PER.C6細胞を発現するFSHにおけるpST3またはpST6の安定的な発現;クローンのトランスフェクション単離およびスクリーニング。
FSHの両ポリペプチド鎖を既に発現しているPer.C6細胞においてα2,3シアリルトランスフェラーゼまたはα2,6シアリルトランスフェラーゼを別々のプラスミド(実施例2および3を参照)から発現させることにより、高度にシアリル化されたFSHを産生するPer.C6クローンを生成した(実施例4を参照)。実施例4で示したようなPER.C6(登録商標)細胞から生成した4つのクローンを、生産性、良好な増殖特性、機能的タンパク質の産生、および産生されたFSH(幾らかのシアリル化を含む)などの特徴によって選択した。
Per.C6が産生したFSHアイソフォームのpIの等電点電気泳動による分析。
電気泳動は、電場による荷電された分子の溶媒中の移動と定義される。電場中の生体分子の移動度は、電場の強さ、分子の正味電荷、分子のサイズおよび形状、分子が移動する媒体のイオン強度および特性に依存するだろう。
るPer.C6 FSHアイソフォームを視覚化することができる。
Per.C6 FSHのシアル酸結合の分析
レクチンに基づくグリカン識別法を使用して、複合糖質を分析した。この方法では、ニトロセルロースに結合した糖タンパク質および複合糖質を特徴づけることができる。レクチンは、特定の部分、例えばα2,3結合シアル酸を選択的に認識する。添加されたレクチンは、ステロイドハプテンジゴキシゲニンと結合し、これによって結合したレクチンの免疫学的検出が可能になる。
(α2−3)シアル酸を示した。
致している(Kagawa et al、1988、Takeuchi et al、1988、Svensson et al.、1990)。
総シアル酸の定量
シアル酸は、単糖類と考えられているタンパク質結合炭水化物であり、ガラクトース、マンノース、グルコサミン、ガラクトサミン、およびフコースのような他のモノサッカライドとの組合せで生じる。
α2,3、α2,6、およびα2,8シアル酸の相対量の定量
精製rFSH(実施例11)のα2,3、α2,6、およびα2,8シアル酸の相対パーセント量を、公知の技術を使用して測定した。
5%)であり、α2,6シアリル化は15%〜35%(例えば、21.46%)、およびα2,8シアリル化は0.1%〜3%であることが見出された。
モノ、ジ、トリ、およびテトラアンテナシアリル化構造の相対量の定量
精製rFSH(実施例11)から抽出したグリカンにあるモノ、ジ、トリ、およびテトラシアリル化構造の相対パーセント量を、公知の技術を使用して測定した。
rFSHの代謝クリアランス率の測定
Per.C6 FSH試料の代謝クリアランス率(MCR)を測定するために、意識のある雌ラット(1クローン当たり3匹の動物)の尾部静脈に、rFSHのボーラスを時点ゼロで注射した(1〜10μg/ラット、試料のELISA定量に基づいて、DRG EIA 1288)。血液試料(400μl)を、試験試料注射の1、2、4、8、12、24、および32時間後に尾部先端から採取した。血清を遠心分離により収集し、ELISA(DRG EIA 1288)によりFSH含有量についてアッセイした。
cer and Ashwell、1971.Van Lenten and Ashwell、1972)。ASGP受容体および結合した脱シアリ化糖タンパク質は、細胞内に内部移行し、そこで受容体が再利用され、リガンドが分解される(Regoeczi et al、1978、Steer and Ashwell、1980)。
されることが示された(図8)。α2,3−シアリルトランスフェラーゼによる操作は、その結果として、基準物質と同様のMCRを有するクローンをもたらし(図9)、シアル酸含有量が様々であったことは、FSHのアイソフォームで知られていることと一致していた(図10)。
Steelman−Pohley in vivoアッセイ
FSHのシアル酸含有量の増加が、その結果として生物学的効果の増加をもたらすことを示すために、高度にシアリル化されたFSH、例えば実施例5で産生されたものなど、によるラット卵巣の重量増加を検査した。
産生および精製の概要
無血清培地に懸濁培養したPER.C6細胞中でFSHを産生するための手順を開発した。その手順は下記に記載されており、数個のFSH産生PER.C6細胞系に適用した。
降し、続いて1×106細胞/mlの密度でVPRO培地+6mM L−グルタミンに懸濁した。その後、細胞を37℃、5%CO2、および100rpmで7〜10日間振とうフラスコで培養した。この期間中に、細胞は107細胞/mlを超える密度に増殖した。細胞生存率が低下し始めた後、培地を回収した。細胞を、1000rpmで5分間遠心沈降し、上清をFSHの定量および精製に使用した。FSHの濃度を、ELISA(DRG
EIA 1288)を使用して測定した。
Andersen CY, Westergaard LG, and van Wely M. (2004). FSH isoform composition of commercial gonadotrophin preparations: a neglected aspect? Reprod Biomed Online.
9(2), 231-236.
Arey BJ, Stevis PE, Deecher DC, Shen ES, Frail DE, Negro-Vilar A, and Lopez FJ. (1997) Induction of promiscuous G protein coupling of the follicle-stimulating hormone (FSH) receptor: a novel mechanism for transducing pleiotropic actions of FSH isoforms. Mol Endocrinol. 11(5), 517-526.
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卵胞刺激ホルモンアルファポリペプチド
受託番号AH007338
FSHアルファのヌクレオチド配列
1 ATGGATTACT ACAGAAAATA TGCAGCTATC TTTCTGGTCA CATTGTCGGT GTTTCTGCAT
61 GTTCTCCATT CCGCTCCTGA TGTGCAGGAT TGCCCAGAAT GCACGCTACA GGAAAACCCA
121 TTCTTCTCCC AGCCGGGTGC CCCAATACTT CAGTGCATGG GCTGCTGCTT CTCTAGAGCA
181 TATCCCACTC CACTAAGGTC CAAGAAGACG ATGTTGGTCC AAAAGAACGT CACCTCAGAG
241 TCCACTTGCT GTGTAGCTAA ATCATATAAC AGGGTCACAG TAATGGGGGG TTTCAAAGTG
301 GAGAACCACA CGGCGTGCCA CTGCAGTACT TGTTATTATC ACAAATCTTA A
FSHアルファのタンパク質配列
1 MKTLQFFFLF CCWKAICCNS CELTNITIAI EKEECRFCIS INTTWCAGYC YTRDLVYKDP
61 ARPKIQKTCT FKELVYETVR VPGCAHHADS LYTYPVATQC HCGKCDSDST DCTVRGLGPS
121 YCSFGEMKE
配列番号2
卵胞刺激ホルモンベータポリペプチド
受託番号NM_000510
FSHベータのヌクレオチド配列
1 ATGAAGACAC TCCAGTTTTT CTTCCTTTTC TGTTGCTGGA AAGCAATCTG CTGCAATAGC
61 TGTGAGCTGA CCAACATCAC CATTGCAATA GAGAAAGAAG AATGTCGTTT CTGCATAAGC
121 ATCAACACCA CTTGGTGTGC TGGCTACTGC TACACCAGGG ATCTGGTGTA TAAGGACCCA
181 GCCAGGCCCA AAATCCAGAA AACATGTACC TTCAAGGAAC TGGTATATGA AACAGTGAGA
241 GTGCCCGGCT GTGCTCACCA TGCAGATTCC TTGTATACAT ACCCAGTGGC CACCCAGTGT
301 CACTGTGGCA AGTGTGACAG CGACAGCACT GATTGTACTG TGCGAGGCCT GGGGCCCAGC
361 TACTGCTCCT TTGGTGAAAT GAAAGAATAA
FSHベータのタンパク質配列
1 MKTLQFFFLF CCWKAICCNS CELTNITIAI EKEECRFCIS INTTWCAGYC YTRDLVYKDP
61 ARPKIQKTCT FKELVYETVR VPGCAHHADS LYTYPVATQC HCGKCDSDST DCTVRGLGPS
121 YCSFGEMKE
配列番号3
ベータガラクトシドアルファ−2,3−シアリルトランスフェラーゼ4
受託番号L23767
ST3GAL4のヌクレオチド配列
1 ATGTGTCCTG CAGGCTGGAA GCTCCTGGCC ATGTTGGCTC TGGTCCTGGT CGTCATGGTG
61 TGGTATTCCA TCTCCCGGGA AGACAGGTAC ATCGAGCTTT TTTATTTTCC CATCCCAGAG
121 AAGAAGGAGC CGTGCCTCCA GGGTGAGGCA GAGAGCAAGG CCTCTAAGCT CTTTGGCAAC
181 TACTCCCGGG ATCAGCCCAT CTTCCTGCGG CTTGAGGATT ATTTCTGGGT CAAGACGCCA
241 TCTGCTTACG AGCTGCCCTA TGGGACCAAG GGGAGTGAGG ATCTGCTCCT CCGGGTGCTA
301 GCCATCACCA GCTCCTCCAT CCCCAAGAAC ATCCAGAGCC TCAGGTGCCG CCGCTGTGTG
361 GTCGTGGGGA ACGGGCACCG GCTGCGGAAC AGCTCACTGG GAGATGCCAT CAACAAGTAC
421 GATGTGGTCA TCAGATTGAA CAATGCCCCA GTGGCTGGCT ATGAGGGTGA CGTGGGCTCC
481 AAGACCACCA TGCGTCTCTT CTACCCTGAA TCTGCCCACT TCGACCCCAA AGTAGAAAAC
541 AACCCAGACA CACTCCTCGT CCTGGTAGCT TTCAAGGCAA TGGACTTCCA CTGGATTGAG
601 ACCATCCTGA GTGATAAGAA GCGGGTGCGA AAGGGTTTCT GGAAACAGCC TCCCCTCATC
661 TGGGATGTCA ATCCTAAACA GATTCGGATT CTCAACCCCT TCTTCATGGA GATTGCAGCT
721 GACAAACTGC TGAGCCTGCC AATGCAACAG CCACGGAAGA TTAAGCAGAA GCCCACCACG
781 GGCCTGTTGG CCATCACGCT GGCCCTCCAC CTCTGTGACT TGGTGCACAT TGCCGGCTTT
841 GGCTACCCAG ACGCCTACAA CAAGAAGCAG ACCATTCACT ACTATGAGCA GATCACGCTC
901 AAGTCCATGG CGGGGTCAGG CCATAATGTC TCCCAAGAGG CCCTGGCCAT TAAGCGGATG
961 CTGGAGATGG GAGCTATCAA GAACCTCACG TCCTTCTGA
ST3GAL4のタンパク質配列
1 MCPAGWKLLA MLALVLVVMV WYSISREDRY IELFYFPIPE KKEPCLQGEA ESKASKLFGN
61 YSRDQPIFLR LEDYFWVKTP SAYELPYGTK GSEDLLLRVL AITSSSIPKN IQSLRCRRCV
121 VVGNGHRLRN SSLGDAINKY DVVIRLNNAP VAGYEGDVGS KTTMRLFYPE SAHFDPKVEN
181 NPDTLLVLVA FKAMDFHWIE TILSDKKRVR KGFWKQPPLI WDVNPKQIRI LNPFFMEIAA
241 DKLLSLPMQQ PRKIKQKPTT GLLAITLALH LCDLVHIAGF GYPDAYNKKQ TIHYYEQITL
301 KSMAGSGHNV SQEALAIKRM LEMGAIKNLT SF
配列番号4
ベータガラクトサミドアルファ−2,6シアリルトランスフェラーゼ1
受託番号NM_003032
ST6GAL1のヌクレオチド配列
1 ATGATTCACA CCAACCTGAA GAAAAAGTTC AGCTGCTGCG TCCTGGTCTT TCTTCTGTTT
61 GCAGTCATCT GTGTGTGGAA GGAAAAGAAG AAAGGGAGTT ACTATGATTC CTTTAAATTG
121 CAAACCAAGG AATTCCAGGT GTTAAAGAGT CTGGGGAAAT TGGCCATGGG GTCTGATTCC
181 CAGTCTGTAT CCTCAAGCAG CACCCAGGAC CCCCACAGGG GCCGCCAGAC CCTCGGCAGT
241 CTCAGAGGCC TAGCCAAGGC CAAACCAGAG GCCTCCTTCC AGGTGTGGAA CAAGGACAGC
301 TCTTCCAAAA ACCTTATCCC TAGGCTGCAA AAGATCTGGA AGAATTACCT AAGCATGAAC
361 AAGTACAAAG TGTCCTACAA GGGGCCAGGA CCAGGCATCA AGTTCAGTGC AGAGGCCCTG
421 CGCTGCCACC TCCGGGACCA TGTGAATGTA TCCATGGTAG AGGTCACAGA TTTTCCCTTC
481 AATACCTCTG AATGGGAGGG TTATCTGCCC AAGGAGAGCA TTAGGACCAA GGCTGGGCCT
541 TGGGGCAGGT GTGCTGTTGT GTCGTCAGCG GGATCTCTGA AGTCCTCCCA ACTAGGCAGA
601 GAAATCGATG ATCATGACGC AGTCCTGAGG TTTAATGGGG CACCCACAGC CAACTTCCAA
661 CAAGATGTGG GCACAAAAAC TACCATTCGC CTGATGAACT CTCAGTTGGT TACCACAGAG
721 AAGCGCTTCC TCAAAGACAG TTTGTACAAT GAAGGAATCC TAATTGTATG GGACCCATCT
781 GTATACCACT CAGATATCCC AAAGTGGTAC CAGAATCCGG ATTATAATTT CTTTAACAAC
841 TACAAGACTT ATCGTAAGCT GCACCCCAAT CAGCCCTTTT ACATCCTCAA GCCCCAGATG
901 CCTTGGGAGC TATGGGACAT TCTTCAAGAA ATCTCCCCAG AAGAGATTCA GCCAAACCCC
961 CCATCCTCTG GGATGCTTGG TATCATCATC ATGATGACGC TGTGTGACCA GGTGGATATT
1021 TATGAGTTCC TCCCATCCAA GCGCAAGACT GACGTGTGCT ACTACTACCA GAAGTTCTTC
1081 GATAGTGCCT GCACGATGGG TGCCTACCAC CCGCTGCTCT ATGAGAAGAA TTTGGTGAAG
1141 CATCTCAACC AGGGCACAGA TGAGGACATC TACCTGCTTG GAAAAGCCAC ACTGCCTGGC
1201 TTCCGGACCA TTCACTGCTA A
0p−
ST6GAL1のタンパク質配列
1 MIHTNLKKKF SCCVLVFLLF AVICVWKEKK KGSYYDSFKL QTKEFQVLKS LGKLAMGSDS
61 QSVSSSSTQD PHRGRQTLGS LRGLAKAKPE ASFQVWNKDS SSKNLIPRLQ KIWKNYLSMN
121 KYKVSYKGPG PGIKFSAEAL RCHLRDHVNV SMVEVTDFPF NTSEWEGYLP KESIRTKAGP
181 WGRCAVVSSA GSLKSSQLGR EIDDHDAVLR FNGAPTANFQ QDVGTKTTIR LMNSQLVTTE
241 KRFLKDSLYN EGILIVWDPS VYHSDIPKWY QNPDYNFFNN YKTYRKLHPN QPFYILKPQM
301 PWELWDILQE ISPEEIQPNP PSSGMLGIII MMTLCDQVDI YEFLPSKRKT DVCYYYQKFF
361 DSACTMGAYH PLLYEKNLVK HLNQGTDEDI YLLGKATLPG FRTIHC
Claims (41)
- α2,3−およびα2,6−シアリル化を含む組換えFSH(rFSH)。
- 6mol/mol以上のシアル酸含有量[シアル酸モル数対タンパク質モル数の比率として表される]を有する、請求項1に記載の組換えFSH。
- 6mol/mol〜15mol/molのシアル酸含有量を有する、請求項1または請求項2に記載の組換えFSH。
- 総シアリル化の10%以上がα2,3−シアリル化である、請求項1から3のいずれかに記載の組換えFSH。
- 総シアリル化の65〜85%の量のα2,3−シアリル化を含む、請求項1から4のいずれかに記載の組換えFSH。
- 総シアリル化の70〜80%の量のα2,3−シアリル化を含む、請求項1から5のいずれかに記載の組換えFSH。
- 総シアリル化の50%以下がα2,6−シアリル化である、請求項1から6のいずれかに記載の組換えFSH。
- 総シアリル化の15〜35%の量のα2,6−シアリル化を含む、請求項1から7のいずれかに記載の組換えFSH。
- 総シアリル化の20〜30%の量のα2,6−シアリル化を含む、請求項1から8のいずれかに記載の組換えFSH。
- α2,8−シアリル化をさらに含む、請求項1から9のいずれかに記載の組換えFSH。
- シアル酸含有量が6質量%以上である、請求項1から10のいずれかに記載の組換えFSH。
- ヒト細胞系で産生または発現される、請求項1から11のいずれかに記載の組換えFSH。
- Per.C6細胞系、Per.C6由来細胞系、または修飾Per.C6細胞系で産生または発現される、請求項1から12のいずれかに記載の組換えFSH。
- 前記細胞系が、α2,3−シアリルトランスフェラーゼを使用して修飾されている、請求項12または13に記載の組換えFSH。
- 内因性シアリルトランスフェラーゼ活性によりもたらされるα2,6結合シアル酸(α2,6−シアリル化)を含む、請求項12から14のいずれかに記載の組換えFSH。
- ヒト細胞系で発現される組換えFSH。
- 総シアリル化の10%以上がα2,3−シアリル化である、請求項16に記載の組換えFSH。
- 総シアリル化の65〜85%の量のα2,3−シアリル化を含む、請求項16または17に記載の組換えFSH。
- 総シアリル化の50%以下がα2,6−シアリル化である、請求項16から18のいずれかに記載の組換えFSH。
- 総シアリル化の15〜35%の量のα2,6−シアリル化を含む、請求項16から19のいずれかに記載の組換えFSH。
- 6mol/mol以上のシアル酸含有量[シアル酸モル数対タンパク質モル数の比率として表される]を含む、請求項16から20のいずれかに記載の組換えFSH。
- α2,3−シアリル化およびα2,6−シアリル化を含む、請求項16または21のいずれかに記載の組換えFSH。
- α2,3−およびα2,6−シアリル化を含む組換えFSH調製物。
- 医薬品用調製物である、請求項23に記載の調製物。
- 6mol/mol以上のシアル酸含有量[シアル酸モル数対タンパク質モル数の比率として表される]を有する、請求項23または24に記載の調製物。
- 総シアリル化の10%以上がα2,3−シアリル化である、請求項23から25のいずれかに記載の調製物。
- 総シアリル化の65〜85%の量のα2,3−シアリル化を含む、請求項23から26のいずれかに記載の調製物。
- 総シアリル化の50%以下がα2,6−シアリル化である、請求項23から27のいずれかに記載の調製物。
- 総シアリル化の15〜35%の量のα2,6−シアリル化を含む、請求項23から28のいずれかに記載の調製物。
- ヒト細胞系で産生または発現される、請求項23から29のいずれかに記載の調製物。
- α2,3−シアリル化およびα2,6−シアリル化を含むrFSHを含む医薬組成物。
- 総シアリル化の10%以上がα2,3−シアリル化である、請求項31に記載の医薬組成物。
- 総シアリル化の65〜85%の量のα2,3−シアリル化を含む、請求項31または32のいずれかに記載の医薬組成物。
- 総シアリル化の50%以下がα2,6−シアリル化である、請求項31から33のいずれかに記載の医薬組成物。
- 総シアリル化の15〜35%の量のα2,6−シアリル化を含む、請求項31から33のいずれかに記載の医薬組成物。
- 請求項1から22のいずれかに記載のrFSHおよび/または請求項23から30のいずれかに記載の調製物を含む医薬組成物。
- hCGおよび/またはLHをさらに含む、請求項31から36のいずれかに記載の医薬組成物。
- 不妊症治療に使用するための、請求項31から37のいずれかに記載の医薬組成物。
- 不妊症を治療する方法であって、請求項1から22のいずれかに記載のrFSH、および/または請求項23から30のいずれかに記載の調製物、および/または請求項31から37のいずれかに記載の医薬組成物を含む組成物を、対象に投与するステップを含む方法。
- 不妊症治療用の薬剤の製造における、請求項1から22のいずれかに記載のrFSHおよび/または請求項23から30のいずれかに記載のrFSH調製物の使用。
- ヒト細胞系でrFSHを産生または発現させるステップを含む、請求項1から22のいずれかに記載のrFSHおよび/または請求項23から30のいずれかに記載のrFSH調製物の生産方法。
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HRP20150968T4 (hr) | 2010-09-29 | 2023-11-10 | Ferring B.V. | Pripravak za uporabu za liječenje neplodnosti |
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CN103619358B (zh) * | 2011-03-31 | 2017-02-15 | 辉凌公司 | 药物制剂 |
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WO2013093760A2 (en) * | 2011-12-19 | 2013-06-27 | Grifols, S.A. | Compositions, methods, and kits for preparing sialylated recombinant proteins |
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JP6652334B2 (ja) | 2014-05-31 | 2020-02-19 | Jcrファーマ株式会社 | ウリジンとn−アセチル−d−マンノサミンとを含有する培地 |
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RU2704252C2 (ru) | 2015-04-17 | 2019-10-25 | Ферринг Б.В. | Композиция для лечения бесплодия |
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GB201603280D0 (en) | 2016-02-24 | 2016-04-13 | Ferring Bv | Stable liquid gonadotropin formulation |
EP3382014A1 (en) | 2017-03-29 | 2018-10-03 | CEVEC Pharmaceuticals GmbH | Recombinant glycoproteins with reduced antennary fucosylation |
EP3441471A1 (en) | 2017-08-08 | 2019-02-13 | CEVEC Pharmaceuticals GmbH | Use of constitutively active variants of growth factor receptors as selection makers for the generation of stable producer cell lines |
EP3973982B1 (en) | 2017-09-01 | 2024-07-31 | Ferring B.V. | Composition for controlled ovarian stimulation |
US20210038694A1 (en) | 2018-04-30 | 2021-02-11 | Ferring B.V. | Composition for controlled ovarian stimulation |
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TW202237173A (zh) | 2020-12-09 | 2022-10-01 | 荷蘭商菲林公司 | 用於受控的卵巢刺激之組成物及方法 |
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