Chimeric activators: quantitatively designed protein therapeutics and uses thereof

Abstract

Aspects of the invention provide methods for harnessing the potential of proteins that occur naturally (e.g., in humans) and that have serious but finite toxicity. Aspects of the invention relate to a quantitative systems-biological and structural approach to design a class of chimeric proteins that avoid the toxicity of protein drugs while retaining their desired activities. In particular, chimeric proteins containing a variant form of a natural protein fused to a targeting moiety may be administered to a subject to target a signal (e.g., induction of apoptosis) to particular cells without having a generalized toxic effect in the subject.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]This application is a continuation of co-pending U.S. application Ser. No. 12 / 594,747 filed Apr. 16, 2010, which is a 35 U.S.C. §371 National Phase Entry Application of International Application No. PCT / US08 / 04435 filed Apr. 5, 2008, which designates the U.S. and which claims the benefit under 35 U.S.C. §119(e) from U.S. provisional applications Ser. No. 60 / 910,390 filed Apr. 5, 2007, Ser. No. 61 / 005,775 filed Dec. 7, 2007 and Ser. No. 61 / 065,937 filed Feb. 15, 2008, the entire contents of which are herein incorporated by reference.SEQUENCE LISTING[0002]The instant application contains a Sequence Listing which has been submitted in ASCII format via EFS-Web and is hereby incorporated by reference in its entirety. Said ASCII copy, created on Jun. 19, 2017, is named Sequence-Listing-12594747.txt and is 56,625 bytes in size.FIELD OF THE INVENTION[0003]The invention relates to chimeric proteins, fusion proteins, and therapeutic proteins.BACKGR...

AI Technical Summary

Benefits of technology

[0009]Aspects of the invention relate to methods and compositions for specifically targeting the activity of naturally-occurring proteins to certain cells or tissues where their activity can be therapeutically useful. In some aspects, the invention provides therapeutic applications for naturally-occurring proteins that otherwise have serious but finite toxicity. Methods and compositions of the invention can be used to target the cell-activating properties of a protein to a subset of target cells without activating non-target cells. Aspects of the invention relate to chimeric proteins that include a variant of a cell-activating element (also referred to herein as an Activity element) that has reduced cell-activating properties relative to its wild-type counterpart. The chimeric proteins also include a cell-targeting element (also referred to herein as a targeting element) that preferentially binds to target cells of interest. The cell-activating variant and the cell-targeting element are connected via a linker. According to aspects of the invention, the activating and targeting elements are selected so that the resulting chimeric protein only activates cells that are bound by the targeting element. The activating element variant is selected such that it can only activate a cell when present in a sufficiently high amount (e.g., concentration) on or near a cell-surface. The targeting element is selected such that it can target a sufficient amount of the activating element to the surface of a target cell in order to activate it. In some embodiments, the linker is selected so that it allows simultaneous binding of the activating and targeting elements to different receptors on the target cell. Accordingly, chimeric proteins of the invention can be used to avoid or reduce unwanted side effects caused by the activating element activating non-target cells.

[0010]Aspects of the invention are particularly useful for targeting the activity of toxic activating elements to target cells (e.g., to target cytotoxic elements to cancer cells) without killing surrounding non-target cells. Aspects of the invention can be useful to harness certain functions of natural proteins that otherwise may be toxic (or have serious toxic side effects) when administered therapeutically. For example, IFNα is normally secreted by virus-infected cells and promotes the death of cells in the immediate vicinity. TNF is secreted in response to the presence of bacterial molecules such as lipopolysaccharide and is thought to play an important role in wound-healing. Other similar molecules include, but are not limited to, IFNβ, IL-2, IFNγ, and IL-1. These therapeutic proteins are usually administered systemically and all have serious side effects, but nonetheless can be very useful. Erythropoietin can also have serious side effects when treating anemia associated with cancer, since erythropoietin can stimulate the growth and survival of certain cancer cells. However, by using less active variants of these proteins in combination with targeting elements of the invention, the general toxicity of protein drugs can be avoided while retaining desired activities (e.g., cell killing activities, enzymatic activities, signal transduction activities, etc., or any combination thereof).

[0014]Accordingly, aspects of the invention relate to chimeric activators including an activity element and a targeting element, optionally joined by a linker, wherein the targeting element binds to a first binding site on a target cell, and wherein the activity element binds to a second binding site on a target cell, and wherein the linker length has been optimized for maximizing binding efficiency of the activity element to the second binding site when the targeting element is bound to the first binding site. In some embodiments the first binding site is a ligand binding site, a receptor, or an epitope on a cell surface. In some embodiments the second binding site is a signal-transduction receptor on a cell surface. In some embodiments the targeting element is an antibody, an antibody fragment, an Fc region, a natural or synthetic ligand, or a protein interaction motif. In some embodiments the activity element is a cytokine, a cytotoxic protein, a regulatory protein, or an active fragment of any one thereof.

[0016]Aspects of the invention relate to methods for manufacturing a chimeric activator including an activity element and a targeting element joined by a linker, wherein the targeting element binds to a first binding site on a target cell, and wherein the activity element binds to a second binding site on a target cell, and wherein the linker length has been optimized for maximizing binding efficiency of the activity element to the second binding site when the targeting element is bound to the first binding site. In some embodiments methods include: i) optimizing the linker length as a function of the concentration of the first and second binding sites, the size of the activity element and targeting element, the minimum distance between the binding sites of the targeting element and the activity element, and the affinities of the activity element and the targeting element for the first and second binding sites; and ii) preparing a chimeric activator having an optimal linker length.

[0018]Aspects of the invention relate to methods for targeted cellular regulation, the method including: administering to a subject a therapeutically effective amount of a chimeric activator including an activity element and a targeting element joined by a linker, wherein the targeting element binds to a first binding site on a target cell, and wherein the activity element binds to a second binding site on a target cell, and wherein the linker length has been optimized for maximizing binding efficiency of the activity element to the second binding site when the targeting element is bound to the first binding site.

[0019]Further described herein are methods for treating cancer, the method including: administering to a subject having a cancer a therapeutically effective amount of a composition including a chimeric activator including an activity element and a targeting element joined by a linker, wherein the targeting element binds to a first binding site on a target cell, and wherein the activity element binds to a second binding site on a target cell, and wherein the linker length has been optimized for maximizing binding efficiency of the activity element to the second binding site when the targeting element is bound to the first binding site.

Problems solved by technology

Erythropoietin can also have serious side effects when treating anemia associated with cancer, since erythropoietin can stimulate the growth and survival of certain cancer cells.

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