Tailored collagen binding of albumin-fused hyperactive coagulation factor IX dictates in vivo distribution and functional properties

Kristin Hovden Aaen; Maria Francesca Testa; Jeannette Nilsen; Rebecca Tarantino; Cesare Canepari; Mascia Benedusi; Sopisa Benjakul; Mari Nyquist-Andersen; Marie Leangen Herigstad; Alessio Cantore; Giuseppe Valacchi; Inger Sandlie; Francesco Bernardi; Mirko Pinotti; Alessio Branchini; Jan Terje Andersen

doi:10.1038/s41467-025-62955-9

Tailored collagen binding of albumin-fused hyperactive coagulation factor IX dictates in vivo distribution and functional properties

Nat Commun. 2025 Sep 29;16(1):8433. doi: 10.1038/s41467-025-62955-9.

Authors

Kristin Hovden Aaen^#^{1

2

3}, Maria Francesca Testa^#⁴, Jeannette Nilsen^{1

2

3}, Rebecca Tarantino⁴, Cesare Canepari^{5

6}, Mascia Benedusi⁷, Sopisa Benjakul^{1

2

3}, Mari Nyquist-Andersen^{1

2

3}, Marie Leangen Herigstad^{1

2

3}, Alessio Cantore^{5

6}, Giuseppe Valacchi^{8

9

10}, Inger Sandlie¹¹, Francesco Bernardi⁴, Mirko Pinotti⁴, Alessio Branchini¹², Jan Terje Andersen^{13

14

15}

Affiliations

¹ Institute of Clinical Medicine, Department of Pharmacology, University of Oslo, Oslo, Norway.
² Department of Immunology, Oslo University Hospital Rikshospitalet, Oslo, Norway.
³ Precision Immunotherapy Alliance (PRIMA), University of Oslo, Oslo, Norway.
⁴ Department of Life Sciences and Biotechnology, University of Ferrara, Ferrara, Italy.
⁵ San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, Milan, Italy.
⁶ Vita-Salute San Raffaele University, Milan, Italy.
⁷ Department of Neuroscience and Rehabilitation, University of Ferrara, Ferrara, Italy.
⁸ Department of Environmental Sciences and Prevention, University of Ferrara, Ferrara, Italy.
⁹ Department of Neurosciences and Rehabilitation; Animal Science Department, NC Research Campus, Plants for Human Health Institute, NC State University, Kannapolis, NC, USA.
¹⁰ Department of Food and Nutrition, Kyung Hee University, Seoul, Republic of Korea.
¹¹ Department of Biosciences, University of Oslo, Oslo, Norway.
¹² Department of Life Sciences and Biotechnology, University of Ferrara, Ferrara, Italy. brnlss@unife.it.
¹³ Institute of Clinical Medicine, Department of Pharmacology, University of Oslo, Oslo, Norway. j.t.andersen@medisin.uio.no.
¹⁴ Department of Immunology, Oslo University Hospital Rikshospitalet, Oslo, Norway. j.t.andersen@medisin.uio.no.
¹⁵ Precision Immunotherapy Alliance (PRIMA), University of Oslo, Oslo, Norway. j.t.andersen@medisin.uio.no.

^# Contributed equally.

Abstract

The efficacy of hemophilia B (HB) replacement therapy is evaluated by coagulation factor IX (FIX) activity in plasma, although FIX bound to extravascular type IV collagen (Col4) also contributes to efficient hemostasis. Here, we investigated the impact of engineering FIX for improved (K5R) or reduced (K5A) Col4 binding on the pharmacokinetic properties of FIX Padua, fused to human serum albumin (HSA_QMP) engineered for favorable neonatal Fc receptor (FcRn) engagement. Hyperactive features and extended plasma half-life in human FcRn expressing mice, attributed to FIX Padua and HSA_QMP engineering, respectively, was confirmed. In HB mice, Padua_KA-HSA_QMP exhibited negligible extravascular distribution and the highest plasma levels at early time points followed by the steepest decay. Conversely, Padua_KR-HSA_QMP showed increased extravascular distribution and a 3-fold longer functional half-life (80 hours). These findings support the use of Padua_KA-HSA_QMP and Padua_KR-HSA_QMP as hyperactive short- or long-term therapeutics, respectively, with opportunities for tailored HB replacement therapy.

MeSH terms

Animals
Collagen Type IV* / metabolism
Collagen* / metabolism
Factor IX* / genetics
Factor IX* / metabolism
Factor IX* / pharmacokinetics
Half-Life
Hemophilia B* / blood
Hemophilia B* / genetics
Hemophilia B* / metabolism
Hemophilia B* / therapy
Histocompatibility Antigens Class I / genetics
Histocompatibility Antigens Class I / metabolism
Humans
Male
Mice
Mice, Inbred C57BL
Protein Binding
Protein Engineering
Receptors, Fc / genetics
Receptors, Fc / metabolism
Recombinant Fusion Proteins* / genetics
Recombinant Fusion Proteins* / metabolism
Recombinant Fusion Proteins* / pharmacokinetics
Serum Albumin, Human* / genetics
Serum Albumin, Human* / metabolism

Substances

Factor IX
Fc receptor, neonatal
Receptors, Fc
Serum Albumin, Human
Histocompatibility Antigens Class I
Recombinant Fusion Proteins
Collagen Type IV
Collagen

Abstract

MeSH terms

Substances

Grants and funding