Claims (15)
1. Molekula polipeptida dvostruke specifičnosti koja obuhvaća prvi polipeptidni lanac i drugi polipeptidni lanac, pri čemu:
a) prvi polipeptidni lanac sadrži
ai) prvu varijabilnu domenu (VD1) antitijela,
aii) prvu varijabilnu domenu (VR1) T-staničnog receptora (TCR) i
aiii) prvi povezivač (LINK1) koji povezuje spomenute domene;
b) drugi polipeptidni lanac sadrži
bi) drugu varijabilnu domenu (VR2) TCR-a,
bi) drugu varijabilnu domenu (VD2) antitijela, i
biii) drugi povezivač (LINK2) koji povezuje spomenute domene;
pri čemu se spomenuta prva varijabilna domena (VD1) i spomenuta druga varijabilna domena (VD2) povezuju kako bi se stvorilo prvo vezno mjesto (VD1)(VD2) koje specifično veže površinski antigen ljudske imunološke efektorske stanice;
pri čemu je prva varijabilna domena (VR1) jedna od TCR Vα domene i TCR Vβ domene, a druga varijabilna domena (VR2) je druga od TCR Vα domene i TCR Vβ domene, spomenuta prva varijabilna domena (VR1 ) i spomenuta druga varijabilna domena (VR2) povezuju se da bi stvorile drugo vezno mjesto (VR1) (VR2) koje se specifično veže za MHC-povezani peptidni epitop;
pri čemu su spomenuta dva polipeptidna lanca spojena s humanim IgG zglobnim domenama i/ili humanim IgG Fc domenama ili njihovim dimerizirajućim dijelovima;
pri čemu su navedena dva polipeptidna lanca povezana kovalentnim i/ili nekovalentnim vezama između spomenutih zglobnih domena i/ili Fc-domena;
pri čemu je spomenuta polipeptidna molekula dvostruke specifičnosti sposobna istodobno vezati molekulu stanične površine i MHC-povezani peptidni epitop; i
pri čemu je redoslijed varijabilnih domena u dva polipeptidna lanca odabran između VD1-VR1 i VR2-VD2 ili VD2-VR2 i VR1-VD1.1. A polypeptide molecule of dual specificity comprising a first polypeptide chain and a second polypeptide chain, wherein:
a) the first polypeptide chain contains
ai) the first variable domain (VD1) of the antibody,
aii) the first variable domain (VR1) of the T-cell receptor (TCR) i
aiii) the first connector (LINK1) that connects the mentioned domains;
b) the second polypeptide chain contains
bi) the second variable domain (VR2) of the TCR,
bi) the second variable domain (VD2) of the antibody, i
biii) the second linker (LINK2) that connects the mentioned domains;
wherein said first variable domain (VD1) and said second variable domain (VD2) are linked to form a first binding site (VD1)(VD2) that specifically binds a human immune effector cell surface antigen;
wherein the first variable domain (VR1) is one of the TCR Vα domain and the TCR Vβ domain, and the second variable domain (VR2) is the other of the TCR Vα domain and the TCR Vβ domain, said first variable domain (VR1 ) and said second variable domain ( VR2) associate to form a second binding site (VR1) (VR2) that specifically binds to an MHC-associated peptide epitope;
wherein said two polypeptide chains are joined with human IgG hinge domains and/or human IgG Fc domains or their dimerizing parts;
wherein the two polypeptide chains are linked by covalent and/or non-covalent bonds between the mentioned joint domains and/or Fc-domains;
wherein said polypeptide molecule of dual specificity is capable of simultaneously binding a cell surface molecule and an MHC-associated peptide epitope; and
wherein the order of the variable domains in the two polypeptide chains is selected between VD1-VR1 and VR2-VD2 or VD2-VR2 and VR1-VD1.
2. Molekula polipeptida dvostruke specifičnosti u skladu s patentnim zahtjevom 1, pri čemu sekvence linkera LINK1 i/ili LINK2 sadrže najmanje jedan motiv sekvence odabran između GGGS, GGGGS, TVLRT, TVSSAS i TVLSSAS.2. A dual specificity polypeptide molecule according to claim 1, wherein the linker sequences LINK1 and/or LINK2 contain at least one sequence motif selected from GGGS, GGGGS, TVLRT, TVSSAS and TVLSSAS.
3. Polipeptidna molekula dvostruke specifičnosti prema bilo kojem od zahtjeva 1 ili 2, pri čemu spomenuti prvi i drugi polipeptidni lanac nadalje sadrže najmanje zglobnu domenu i Fc domenu ili njihove dijelove izvedene iz humanog IgG1, IgG2 ili IgG4, pri čemu navedena Fc domena poželjno sadrži barem jednu mutaciju efektorske funkcije prigušivanja u slijedu SEQ ID br. 50 (ELLGGP) humanog IgG1, po mogućnosti gdje je navedena mutacija utišavanja efektorske funkcije generirana jednom ili više mutacija E1P, L2V, L3A i bez ostatka na položaju 4 SEQ ID br. 50.3. A double-specificity polypeptide molecule according to any one of claims 1 or 2, wherein said first and second polypeptide chains further contain at least a hinge domain and an Fc domain or parts thereof derived from human IgG1, IgG2 or IgG4, wherein said Fc domain preferably contains at least one mutation of the effector function of silencing in the sequence of SEQ ID no. 50 (ELLGGP) human IgG1, preferably where said effector function silencing mutation is generated by one or more mutations E1P, L2V, L3A and no residue at position 4 of SEQ ID no. 50.
4. Molekula polipeptida dvostruke specifičnosti u skladu s patentnim zahtjevom 3, pri čemu navedena Fc domena sadrži CH3 domenu koja sadrži barem jednu mutaciju koja olakšava stvaranje heterodimera, po mogućnosti, gdje spomenute mutacije sadrže odgovarajuće mutacije.4. A dual-specificity polypeptide molecule according to claim 3, wherein said Fc domain contains a CH3 domain containing at least one mutation that facilitates heterodimer formation, preferably, where said mutations contain corresponding mutations.
5. Polipeptidna molekula dvostruke specifičnosti prema bilo kojem od zahtjeva 3 ili 4, pri čemu navedena Fc domena sadrži CH2 i CH3 domenu(e) koje sadrže najmanje dva dodatna ostatka cisteina.5. A polypeptide molecule of dual specificity according to any one of claims 3 or 4, wherein said Fc domain comprises CH2 and CH3 domain(s) containing at least two additional cysteine residues.
6. Molekula polipeptida dvostruke specifičnosti u skladu s bilo kojim od patentnih zahtjeva 1 do 5, pri čemu spomenute domene izvedene od antitijela VD1 i VD2 prikazuju konstruirani disulfidni most koji uvodi kovalentnu vezu između VD1 i VD2 i gdje se navedeni cisteini uvode u okvirno područje (FR) 4 u slučaj VL i okvirno područje 2 u slučaju VH.6. A dual-specificity polypeptide molecule according to any one of claims 1 to 5, wherein said antibody-derived domains VD1 and VD2 display an engineered disulfide bridge that introduces a covalent bond between VD1 and VD2 and wherein said cysteines are introduced into the framework region ( FR) 4 in the case of VL and frame area 2 in the case of VH.
7. Polipeptidna molekula dvostruke specifičnosti prema bilo kojem od zahtjeva 1 do 6, pri čemu je poznato da spomenuta molekula površinske stanice inducira aktivaciju imunoloških stanica ili je barem jedna odabrana iz skupine koju čine CD3, CD4, CD7, CD8, CD10 , CD11b, CD11c, CD14, CD16, CD18, CD22, CD25, CD28, CD32a, CD32b, CD33, CD41, CD41b, CD42a, CD42b, CD44, CD45RA, CD49, CD55, CD56, CD61, CD64, CD68, CD94, CD90 , CD117, CD123, CD125, CD134, CD137, CD152, CD163, CD193, CD203c, CD235a, CD278, CD279, CD287, Nkp46, NKG2D, GITR, FcεRI, TCRα/β i TCRγ/δ, HLA-DR, po mogućnosti molekula stanične površine je CD3γ, CD3δ ili CD3ε.7. Polypeptide molecule of dual specificity according to any one of claims 1 to 6, wherein said surface cell molecule is known to induce the activation of immune cells or at least one is selected from the group consisting of CD3, CD4, CD7, CD8, CD10, CD11b, CD11c , CD14, CD16, CD18, CD22, CD25, CD28, CD32a, CD32b, CD33, CD41, CD41b, CD42a, CD42b, CD44, CD45RA, CD49, CD55, CD56, CD61, CD64, CD68, CD94, CD90 , CD117, CD123 , CD125, CD134, CD137, CD152, CD163, CD193, CD203c, CD235a, CD278, CD279, CD287, Nkp46, NKG2D, GITR, FcεRI, TCRα/β and TCRγ/δ, HLA-DR, preferably the cell surface molecule CD3γ. , CD3δ or CD3ε.
8. Polipeptidna molekula dvostruke specifičnosti prema bilo kojem od zahtjeva 1 do 7, pri čemu prvi polipeptidni lanac sadrži SEQ ID br. 28, SEQ ID br. 29 i SEQ ID br. 30; a drugi polipeptidni lanac sadrži SEQ ID br. 31, SEQ ID br. 32 i SEQ ID br. 30.8. A polypeptide molecule of dual specificity according to any one of claims 1 to 7, wherein the first polypeptide chain contains SEQ ID no. 28, SEQ ID NO. 29 and SEQ ID NO. 30; and the second polypeptide chain contains SEQ ID no. 31, SEQ ID NO. 32 and SEQ ID NO. 30.
9. Polipeptidna molekula dvostruke specifičnosti prema bilo kojem zahtjevu od 3 do 8, pri čemu Fc regija u prvom polipeptidnom lancu sadrži SEQ ID br. 26 ili SEQ ID br. 47 (Fc1), a Fc regija u drugom polipeptidnom lancu SEQ ID br. 27 ili SEQ ID br. 48 (Fc2).9. A polypeptide molecule of dual specificity according to any one of claims 3 to 8, wherein the Fc region in the first polypeptide chain contains SEQ ID no. 26 or SEQ ID NO. 47 (Fc1), and the Fc region in the second polypeptide chain of SEQ ID no. 27 or SEQ ID NO. 48 (Fc2).
10. Molekula polipeptida dvostruke specifičnosti prema zahtjevu 1, pri čemu prvi lanac polipeptida sadrži SEQ ID br. 16, SEQ ID br. 43, SEQ ID br. 45, SEQ ID br. 51, SEQ ID br. 53, SEQ ID br. 55, ili SEQ ID br. 57, a pri čemu drugi polipeptidni lanac sadrži SEQ ID br. 17, SEQ ID 44, SEQ ID br. 46, SEQ ID br. 52, SEQ ID br. 54, SEQ ID br. 56, ili SEQ ID br. 58.10. A double-specificity polypeptide molecule according to claim 1, wherein the first polypeptide chain contains SEQ ID no. 16, SEQ ID NO. 43, SEQ ID NO. 45, SEQ ID NO. 51, SEQ ID NO. 53, SEQ ID NO. 55, or SEQ ID NO. 57, and the second polypeptide chain contains SEQ ID no. 17, SEQ ID 44, SEQ ID no. 46, SEQ ID NO. 52, SEQ ID NO. 54, SEQ ID NO. 56, or SEQ ID NO. 58.
11. Polipeptidna molekula dvostruke specifičnosti prema bilo kojem od zahtjeva 1 do 10, pri čemu je spomenuto prvo mjesto vezanja (VD1)(VD2) koje veže stanični površinski antigen spomenutih imunih stanica humanizirano; i/ili navedeno drugo vezno mjesto (VR1)(VR2) koje veže spomenuti MHC-povezani peptidni epitop je zrelo u afinitetu i/ili sazrelo za stabilnost.11. A polypeptide molecule of dual specificity according to any one of claims 1 to 10, wherein said first binding site (VD1)(VD2) that binds the cell surface antigen of said immune cells is humanized; and/or said second binding site (VR1)(VR2) that binds said MHC-associated peptide epitope is affinity matured and/or stability matured.
12. Nukleinska kiselina koja kodira prvi polipeptidni lanac i/ili drugi polipeptidni lanac prema bilo kojem od zahtjeva 1 do 11, ili ekspresijski vektor koji sadrži najmanje jednu od spomenutih nukleinskih kiselina.12. Nucleic acid encoding the first polypeptide chain and/or the second polypeptide chain according to any one of claims 1 to 11, or an expression vector containing at least one of the mentioned nucleic acids.
13. Stanica domaćin koja sadrži i po potrebi eksprimira vektor kako je definirano u zahtjevu 12.13. A host cell containing and optionally expressing the vector as defined in claim 12.
14. Farmaceutski pripravak, koji sadrži molekulu polipeptida dvostruke specifičnosti prema bilo kojem zahtjevu od 1 do 11, nukleinsku kiselinu ili ekspresijski vektor prema zahtjevu 12 ili stanicu prema zahtjevu 13, zajedno s jednim ili više farmaceutski prihvatljivih nosača ili pomoćnih tvari.14. A pharmaceutical preparation, containing a polypeptide molecule of dual specificity according to any of claims 1 to 11, a nucleic acid or an expression vector according to claim 12 or a cell according to claim 13, together with one or more pharmaceutically acceptable carriers or excipients.
15. Polipeptidna molekula dvostruke specifičnosti prema bilo kojem od zahtjeva 1 do 11, nukleinska kiselina ili ekspresijski vektor prema zahtjevu 12, stanica prema zahtjevu 13 ili farmaceutski pripravak prema zahtjevu 14, za uporabu u medicini, poželjno za upotrebu u prevenciji ili liječenju zaraznih bolesti.15. Polypeptide molecule of dual specificity according to any of claims 1 to 11, nucleic acid or expression vector according to claim 12, cell according to claim 13 or pharmaceutical preparation according to claim 14, for use in medicine, preferably for use in the prevention or treatment of infectious diseases.