Recombinant bifunctional protein targeting cd47 and her2

Abstract

The present disclosure provides a recombinant fusion protein containing an extracellular Ig-like domain of a signal-regulator protein (SIRP), linked via a linker, to a paratope of an Ig-like anti-HER2 antibody at the N-terminus of a heavy chain or a light chain constituting the paratope. The present disclosure also provides a polynucleotide encoding the recombinant fusion protein, an expression vector containing the polynucleotide, a method for producing the recombinant protein and a method for treating a disease caused by over-expression of CD47 and / or HER2.

Description

CROSS REFERENCE TO RELATED APPLICATIONS[0001]This application is a continuation application of U.S. patent application Ser. No. 16 / 535,075 filed Aug. 8, 2018, which claims priority of U.S. provisional patent application Ser. No. 62 / 716,356 filed Aug. 8, 2018.SEQUENCE LISTING[0002]The instant application contains a Sequence Listing which has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety. Said ASCII copy, is named, 5552500019SL.txt and is 39.0 kbytes in size.FIELD OF THE INVENTION[0003]The present disclosure relates to a recombinant fusion protein targeting CD47 and HER2, and the preparation and use thereof, especially its use in tumor therapies.BACKGROUND OF THE INVENTION[0004]Antibody therapies are approved in various jurisdictions to treat a wide range of cancers, and have significantly improved patient outcomes (Komeev K V et al., (2017) TLR-signaling and proinflammatory cytokines as drivers of tumorigenesis. Cytokine 89: 127...

AI Technical Summary

Benefits of technology

[0014]However, bispecific or multi-specific antibodies present significant design challenges as a number of issues have to be considered, including compatibility of the molecules, the resulting antibody's affinity, stability and pharmaceutical properties. It is well recognized that simply linking antibodies or proteins together does not necessarily result in synergetic / advantageous effects. A recombinant antibody disclosed in the present disclosure, comprising SIRPαD1, linked by a linker, to Erbitux (Cetuximab), has been proved to have inferior anti-tumor activity compared to Erbitux or SIRPαD1-Fc alone in the HT-29 or NCl-H1975 tumor model (see Example 8).

[0015]Through diligent efforts, the present inventors, however, have successfully designed a recombinant bispecific protein that accurately targets both CD47 and HER2 and shows better anti-tumor activity than ordinary single antigen targeting antibodies.SUMMARY OF THE INVENTION

[0016]The present disclosure discloses a recombinant fusion protein, comprising an extracellular Ig-like domain of a signal-regulator protein (SIRP), linked via a linker, to a paratope of an Ig-like anti-HER2 antibody at the N-terminus of a heavy chain or a light chain constituting the paratope, wherein the recombinant fusion protein bind to CD47, HER2 and FcR simultaneously. Binding to CD47s on cancer cells blocks the interaction of CD47s with SIRPs on macrophages and thus releases the check on macrophages by SIRP-mediated inhibitory signals; while binding to HER2s on cancer cells inhibits the uncontrolled tumor cell growth; and at the same time, binding to FcRs on NK cells or macrophages stimulates targeted cancer cell killings by NK cells or macrophages.

[0017]In an embodiment, either paratope of the Ig-like anti-HER2 antibody is linked to an extracellular Ig-like domain of signal-regulator protein (SIRP) at the N-terminus of the heavy chain or the light chain constituting the paratope. In an embodiment, each paratope of the Ig-like anti-HER2 antibody is linked to an extracellular Ig-like domain of signal-regulator protein (SIRP) at the N-terminus of the heavy chain or the light chain constituting that paratope. In one embodiment, each paratope of the Ig-like anti-HER2 antibody is linked to an extracellular Ig-like domain of signal-regulator protein (SIRP) at the N-terminus of the heavy chain constituting that paratope. In one embodiment, each paratope of the Ig-like anti-HER2 antibody is linked to an extracellular Ig-like domain of signal-regulator protein (SIRP) at the N-terminus of the light chain constituting that paratope. In one embodiment, one paratope of the Ig-like anti-HER2 antibody is linked to an extracellular Ig-like domain of signal-regulator protein (SIRP) at the N-terminus of the heavy chain constituting that paratope, and the other paratope is linked to an extracellular Ig-like domain of signal-regulator protein (SIRP) at the N-terminus of the light chain constituting that paratope. In some embodiment, one paratope of the anti-HER2 antibodies may be linked to two copies of the extracellular Ig-like domain of signal-regulator protein (SIRP) at the N-terminus of the heavy chain and light chain constituting that paratope.

[0018]In one embodiment, the signal-regulatory protein in the recombinant fusion protein may be SIRPα, and the extracellular Ig-like domain of the signal-regulatory protein may be the first extracellular Ig-like domain of SIRPα (SIRPαD1). The extracellular Ig-like domain of the signal-regulatory protein, such as SIRPαD1, can bind to CD47 on the cell surfaces of cancer / tumor cells and thus block the interaction of CD47 with SIRPs on the cell surfaces of macrophages.

[0019]In one embodiment, the SIRPαD1 has the nucleic acid sequence and amino acid sequence set forth in SEQ ID NOs: 1 and 2, respectively. In some embodiments, the SIRPαD1 may comprise an amino acid sequence having at least 80%, 85%, 90%, 95%, 98% or 99% identity to SEQ ID NO: 2, wherein the SIRPαD1 can bind to CD47 on the cell surfaces of cancer / tumor cells and block the interaction of CD47 with SIRPs on the cell surfaces of macrophages.

Problems solved by technology

Once bound to a cancer antigen, antibodies may induce antibody-dependent cell-mediated cytotoxicity, activate the complement system, or prevent a receptor from interacting with its ligand, all of which may lead to cell death.

Antibodies have significantly advanced our ability to treat cancers, yet clinical studies have shown many patients do not adequately respond to monospecific therapy.

However, bispecific or multi-specific antibodies present significant design challenges as a number of issues have to be considered, including compatibility of the molecules, the resulting antibody's affinity, stability and pharmaceutical properties.

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