Methods for DNA-dependent targeting of a cell permeant antibody
Inventors
Weisbart, Richard H. • Nishimura, Robert N.
Assignees
US Department of Veterans Affairs
Publication Number
US-11453725-B2
Publication Date
2022-09-27
Expiration Date
2036-02-18
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Abstract
The invention provides methods for selective targeting of live cells, which have undergone or are undergoing radiation or chemotherapy, at a site of interest with a cell-penetrating polypeptide. In one embodiment of the invention, the method comprises contacting the live cells with a cell-penetrating polypeptide comprising cell-penetrating determinants so that the cell-penetrating polypeptide binds extracellular DNA near or around the live cells so as to form a complex or association therewith such that the complex or associated polypeptide-DNA so bound bind the live cells and penetrates the live cells thereby selectively targeting live cells at a site of interest with a cell-penetrating polypeptide.
Core Innovation
The invention provides methods for selectively targeting live cells at a site of interest, particularly cells that have undergone or are undergoing radiation or chemotherapy, using a cell-penetrating polypeptide that comprises cell-penetrating determinants. The method involves contacting live cells with the cell-penetrating polypeptide so that it binds extracellular DNA near or around the cells, forming a complex that penetrates the live cells. This allows selective targeting of live cells at the site with the polypeptide.
Additionally, the invention addresses methods for targeting live cells near cellular injury using cell-penetrating polypeptides optionally joined to therapeutic agents. The method includes administering the polypeptide at or near the injury site where it binds extracellular DNA from the injury, forming complexes that bind and penetrate the live cells. This approach selectively targets live cells at injury sites with the polypeptide, which may comprise a therapeutic agent to inhibit injury or induce cell death.
The problem being solved arises from the need to achieve targeted delivery of therapeutic agents to live cells, especially in contexts such as cancer therapy or tissue injury, minimizing systemic toxicity and enhancing efficacy. The background describes lupus anti-DNA autoantibodies, specifically 3E10, which uniquely penetrate live cells and nuclei without harming normal cells, and can be used as molecular delivery vehicles. However, the mechanism of how these antibodies penetrate nuclei and selectively target cells was not well understood. The invention solves the problem by discovering that penetration requires extracellular DNA, enabling targeted therapies that exploit extracellular DNA binding to selectively deliver therapeutic agents to live cells in situ.
Claims Coverage
The patent contains multiple independent claims centered on methods involving the selective targeting of live cells using cell-penetrating polypeptides, specifically 3E10 antibodies or their fragments, and purified DNA binding.
Method of selective targeting of live cells using 3E10 antibody and purified DNA
A method comprising introducing purified DNA near live cells at a site of interest; administering a cell-penetrating polypeptide comprising cell-penetrating determinants before, after, or concurrently with the purified DNA; allowing the polypeptide to bind the DNA forming complexes; and contacting live cells with the complexes to bind and penetrate them, thereby selectively targeting live cells. The polypeptide is a 3E10 antibody or single chain Fv fragment thereof.
Adjunct therapy to chemotherapy or radiation therapy
Using the selective targeting method as an adjunct to chemotherapy or radiation therapy, where the chemotherapy or radiation may be administered concurrently or before the targeting method.
Conjugation of cell-penetrating polypeptide to therapeutic agent
The 3E10 cell-penetrating polypeptide used in the targeting method may be conjugated to a therapeutic agent to enhance treatment effects.
Use of nucleoside salvage pathway dependence for cell penetration
The cell penetration by the polypeptide-DNA complex depends on a salvage pathway, particularly a nucleoside salvage pathway mediated by equilibrative nucleoside transporters such as ENT2.
Targeting of cells in various injury or disease sites with purified DNA
Methods apply to targeting live cells at sites of injury including ischemic tissues, physical trauma, chemical burn, radiation injury, and various cancers, using purified DNA and the 3E10 antibody-based polypeptide.
Introducing DNA by various administration routes or implants
Introducing or producing purified DNA at or near live cells by injection, microinjection, microprojectile, or implantation, including implants releasing DNA in a single burst or over time, enabling selective cell targeting with the cell-penetrating polypeptide.
Methods for inhibiting cellular injury or inducing cell death
Administering the 3E10 cell-penetrating polypeptide conjugated to therapeutic agents at or near sites of cellular injury, contacting with purified DNA to form complexes that penetrate live cells, to inhibit injury or induce cell death selectively.
Methods for enhancing chemotherapy or radiation therapy effects
Enhancing effects of chemotherapy or radiation by selectively targeting live cells using the 3E10 antibody polypeptide and purified DNA methods.
Methods for increasing uptake and efficacy in targeting tumors
Introducing purified or artificial DNA near tumor or cancer cells followed by administering the 3E10 cell-penetrating polypeptide, allowing formation of complexes to increase uptake and enhance efficacy in tumor targeting.
The claims cover innovative methods of selectively targeting live cells at sites of interest, particularly live cells in tumors or injury sites, using 3E10 antibodies or fragments that bind extracellular purified DNA to form complexes which penetrate live cells. Additional inventive features include the dependence on nucleoside salvage pathways for cell penetration, use of therapeutic conjugates, adjunct use with chemotherapy or radiation, and enhancement of targeting efficacy by introducing purified or artificial DNA near target cells.
Stated Advantages
Selective targeting of live cells at sites of injury or tumors reduces potential systemic toxicity and increases therapeutic effectiveness.
Enhanced nuclear penetration of 3E10 scFv in the presence of extracellular DNA allows selective delivery to cells in areas of high cell turnover or ischemia.
Preferential accumulation of 3E10 antibody in tumor cells and ischemic tissues improves stability and efficacy of delivery with minimal off-target effects.
The method enables targeted therapy that can be used adjunctively with chemotherapy or radiation to improve treatment outcomes.
Documented Applications
Selective targeting and treatment of tumors, including gliomas and various cancers with high cell turnover.
Selective targeting of live cells at sites of injury such as intracranial injury, brain injury, myocardial infarction, skin injury, liver injury, gastrointestinal injury, lung injury, eye injury, kidney injury, pancreas injury, peritoneal injury, bone injury, nasopharyngeal injury, uterine injury, cervical injury, breast injury, organ injury, tissue injury, burn or radiation injury.
Inhibition of cellular injury or induction of cell death at sites of injury or disease using therapeutic agents conjugated to cell-penetrating polypeptides.
Enhancement of chemotherapy or radiation therapy efficacy by selective targeting of live cells via cell-penetrating polypeptides and extracellular DNA.
Increasing uptake and enhancing efficacy of 3E10 antibody-based polypeptides in targeting tumors or cancer cells by introducing purified or artificial DNA near the target cells.
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