Modified soluble FGF receptor Fc fusions method
The invention relates to modified soluble FGF receptor Fc fusions comprising a fusion of a soluble fragment or domain of the FGF receptor part (targeting or binding moiety) with an Fc region of an immunoglobulin part (effector function moiety), having improved biological activity including ADCC/CDC activities, compositions containing them, and method of producing such modified soluble FGF receptor Fc fusion molecules.
1. A method for treating cancer overexpressing fibroblast growth factor 2 (FGF2) comprising administering to a subject with said cancer a therapeutically effective amount of a modified soluble FGF receptor Fc fusion comprising a fusion of a soluble FGF receptor 2 (sFGFR2) moiety with an Fc region of an immunoglobulin, wherein at least the 5 th N-glycosylation site of the FGFreeceptor moeity is occupied, and at most 45% of the N-glycans of the FGF receptor moiety have no sialyl group, wherein the FGF receptor Fc fusion binds FGF, and wherein the FGF receptor Fc fusion exhibits reduced aggregation or improved productivity relative to a soluble FGF receptor Fc fusion lacking the N-glycan at the 5th N-glycosylation site from the N-terminus.
2. The method of claim 1 , further comprising administering an additional therapeutic agent.
3. The method of claim 2 , wherein the additional therapeutic agent is an anti-angiogenic agent or a chemotherapeutic agent.
4. The method of claim 3 , wherein the anti-angiogenic agent is tumor necrosis factor, an antagonist of an acidic or basic fibroblast growth factor (FGF), hepatocyte growth factor (HGF), tissue factor (TF), protein C, protein S, platelet-derived growth factor (PDGF), or HER2 receptor.
5. The method of claim 3 , wherein the chemotherapeutic agent is selected from the group consisting of: anti-microtubule agents; platinum coordination complexes; alkylating agents; antibiotic agents; topoisomerase II inhibitors; antimetabolites; topoisomerase I inhibitors; hormones and hormone analogues; signal transduction pathway inhibitors; non-receptor tyrosine kinase angiogenesis inhibitors; immunotherapeutic agents; pro-apoptotic agents; and cell cycle signaling inhibitors.
6. The method of claim 3 , wherein the chemotherapeutic agent is selected from the group consisting of: taxol and taxotere.
7. The method of claim 1 , wherein the cancer is selected from the group consisting of: carcinoma, bladder cancer, breast cancer, colon cancer, cancer of the head and neck, kidney cancer, renal cell carcinoma, liver cancer, lung cancer, cancer of the ovary, cancer of the pancreas, stomach cancer, cancer of the cervix, thyroid cancer, skin cancer, squamous cell carcinoma, hematopoietic tumors of lymphoid lineage, leukemia, acute lymphocytic leukemia, acute lymphoblastic leukemia, B-cell lymphoma, T-cell lymphoma, Burkitt's lymphoma, hematopoietic tumors of myeloid lineage, acute and chronic myelogenous leukemias, promyelocytic leukemia, tumors of mesenchymal origin, fibrosarcoma, rhabdomyoscarcoma, melanoma, seminoma, tetratocarcinoma, neuroblastoma, glioma, tumors of the central and peripheral nervous system, astrocytoma, neuroblastoma, glioma, schwannomas, tumors of mesenchymal origin, fibrosarcoma, rhabdomyoscarama, osteosarcoma, melanoma, xeroderma pigmentosum, keratoactanthoma, seminoma, thyroid follicular cancer, and teratocarcinoma.
8. The method of claim 1 , wherein the cancer is selected from the group consisting of: melanoma, leukemia, renal cancer, colon cancer, ovarian cancer, prostate cancer, lung cancer, bladder cancer, breast cancer, and head and neck cancer.
9. The method of claim 1 , wherein, in addition, the 3 rd , 4 th , 6 th , and 7 th N-glycosylation sites of the FGF receptor moiety are occupied with a N-glycan.
10. The method of claim 9 , wherein at least 7 N-glycosylation sites of the FGF receptor moiety are occupied with a N-glycan.
11. The method of claim 10 , wherein all N-glycosylation sites of the FGF receptor moiety are occupied with a N-glycan.
12. The method of claim 1 , wherein the average number of sialic acid per N-glycan of the FGF receptor moiety is 0.9 or above.
13. The method of claim 12 , wherein the average number of sialic acid per N-glycan of the FGF receptor moiety is 1.2 or above.
14. The method of claim 1 , wherein the K D value of the fusion for FGF2 measured by Biacore™ is comprised between 1 and 5 nM.
15. The method of claim 14 , wherein the K D value of the fusion for FGF2 measured by Biacore™ is around 1.5 nM.
16. The method of claim 1 , wherein the fusion possesses ADCC and/or CDC activities.
17. The method of claim 1 , wherein the N-glycans of the fusion are 60-100% fucosylated.
18. The method of claim 1 , wherein the modified soluble FGF receptor Fc fusion comprises 3 mannose residues, a mean of 1.5 to 3.0 galactose residues, a mean of 3.5 to 5 of N-acetylglucosamine residues, and a mean of 0.6 to 1 fucose residues per molecule of glycan.
19. The method of claim 1 , wherein the N-glycans of the fusion are 0-60% fucosylated.
20. The method of claim 1 , wherein the FGF receptor is FGF receptor 2 isotype IIIc.
21. The method of claim 1 , wherein the FGF receptor soluble domain has a sequence as set forth in SEQ ID NO: 4, or a sequence having an identity of at least 95% with SEQ ID NO: 4.
22. The method of claim 1 , wherein the Fc portion has a sequence as set forth in SEQ ID NO: 6, or a sequence having an identity of at least 95% with the SEQ ID NO: 6.
23. The method of claim 1 , wherein the modified soluble FGF receptor Fc fusion further comprises a linker sequence of at least 3 amino acid residues.
24. The method of claim 23 , wherein the linker sequence is SAL (Ser-Ala-Leu).
25. The method of claim 1 , wherein the modified soluble FGF receptor Fc fusion has a polypeptide sequence as set forth in SEQ ID NO: 2, or a sequence having an identity of at least 95% with the SEQ ID NO: 2.
26. The method of claim 1 , wherein the modified soluble FGF receptor Fc fusion further comprises the signal peptide of SEQ ID NO: 8.