Abstract:
OBJECTIVE:To develop recombinant factor IX (FIX) variants with augmented clotting activity. RESULTS:We generated three new variants, FIX-YKALW, FIX-ALL and FIX-LLW, expressed in SK-Hep-1 cells and characterized in vitro and in vivo. FIX-YKALW showed the highest antigen expression level among the variants (2.17 µg-mL), followed by FIX-LLW (1.5 µg-mL) and FIX-ALL (0.9 µg-mL). The expression level of FIX variants was two-five fold lower than FIX-wild-type (FIX-WT) (4.37 µg-mL). However, the biological activities of FIX variants were 15-31 times greater than FIX-WT in the chromogenic assay. Moreover, the new variants FIX-YKALW, FIX-LLW and FIX-ALL also presented higher specific activity than FIX-WT (17, 20 and 29-fold higher, respectively). FIX variants demonstrated a better clotting time than FIX-WT. In hemophilia B mice, we observed that FIX-YKALW promoted hemostatic protection. CONCLUSION:We have developed three improved FIX proteins with potential for use in protein replacement therapy for hemophilia B.
journal_name
Biotechnol Lettjournal_title
Biotechnology lettersauthors
Sousa Bomfim A,Corrêa de Freitas MC,Picanço Castro V,Abreu Soares Neto M,Pádua R,Covas DT,Sousa Russo EMdoi
10.1007/s10529-020-03040-7subject
Has Abstractpub_date
2021-01-01 00:00:00pages
143-152issue
1eissn
0141-5492issn
1573-6776pii
10.1007/s10529-020-03040-7journal_volume
43pub_type
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