Synthetic conjugates and uses thereof
Inventors
Zion, Todd C. • Lancaster, Thomas M.
Assignees
SmartCells Inc • National Institutes of Health NIH • US Department of Health and Human Services
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Publication Number
US-8940690-B2
Publication Date
2015-01-27
Expiration Date
2030-01-27
Abstract
The present disclosure provides a cross-linked material comprising conjugates which include two or more separate affinity ligands bound to a non-polymeric framework, wherein the molecular weight of the non-polymeric framework is less than 10,000 Da; and multivalent cross-linking agents that non-covalently bind the affinity ligands of the conjugates and thereby cross-link the conjugates to form a cross-linked material, wherein the non-covalent bonds between the multivalent cross-linking agents and the affinity ligands are competitively dissociated in the presence of excess amounts of a target molecule. The present disclosure also provides methods of making and methods of using these materials. In other aspects, the present disclosure provides exemplary conjugates including conjugates for use in glucose responsive cross-linked materials.
Core Innovation
The invention provides a cross-linked material comprising conjugates that include two or more separate affinity ligands bound to a non-polymeric framework having a molecular weight less than 10,000 Da, and multivalent cross-linking agents that non-covalently bind the affinity ligands and thereby cross-link the conjugates to form a cross-linked material. These non-covalent bonds are competitively dissociated in the presence of excess amounts of a target molecule, enabling controlled release of conjugates.
The problem being solved is that prior art controlled-release drug delivery systems are incapable of releasing drugs at intervals and concentrations proportional to the amount of a molecular indicator in the body, such as glucose. Existing conjugates using glycosylated polymers present difficulties including high molecular weight leading to slow absorption and reduced bioactivity. Enzymatically degradable conjugates face drawbacks of manufacturing challenges and unwanted degradation, with species differences in enzymatic activity complicating pharmacokinetics and regulatory approval. There is therefore a need for conjugates that resist enzymatic degradation, have low molecular weight to preserve pharmacokinetics and bioactivity, and yet can form insoluble cross-linked materials responsive to target molecules.
The invention overcomes these problems by developing families of high affinity ligands conjugated to low molecular weight, non-polymeric frameworks to create well-characterized conjugates that preserve bioactivity, resist enzymatic degradation, and form insoluble cross-linked materials when combined with multivalent cross-linking agents such as lectins. These materials respond to varying concentrations of target molecules by competitively dissociating the non-covalent bonds, enabling controlled release of conjugates such as insulin conjugates for glucose-responsive applications.
Claims Coverage
The patent includes one independent claim describing a cross-linked material with specific molecular and functional characteristics related to conjugates and cross-linking agents.
Cross-linked material with conjugates comprising an anti-diabetic drug and two or more separate affinity ligands bound to a low molecular weight non-polymeric framework
The material includes conjugates with an anti-diabetic drug and two or more separate affinity ligands bound to a non-polymeric framework having molecular weight less than 10,000 Da.
Cross-linking by multivalent agents with competitively dissociable non-covalent bonds
Multivalent cross-linking agents non-covalently bind the affinity ligands of the conjugates to form a cross-linked material, wherein these non-covalent bonds are competitively dissociated in the presence of excess amounts of a target molecule.
Affinity ligands comprising saccharides for glucose as the target molecule
When the target molecule is glucose, the two or more separate affinity ligands comprise a saccharide such as glucose, mannose, glucosamine, mannosamine, methylglucose, methylmannose, ethylglucose, or ethylmannose, including aminoethylglucose (AEG), aminoethylmannose (AEM), aminoethylbimannose (AEBM) or aminoethyltrimannose (AETM).
Conjugate general structures
The conjugates have defined general formulas including formula (I) and subformulas (VIa), (VIb), and (VIc), specifying molecular branching and component arrangements.
Insulin molecules as the anti-diabetic drug and conjugation sites
The anti-diabetic drug can be an insulin molecule conjugated to the non-polymeric framework via specific amino acid residues such as A1, B1, or B29.
Use of lectins as multivalent cross-linking agents, including modified lectins
The multivalent cross-linking agent includes a lectin, which may be covalently bonded to a recognition element that competes with the target molecule and conjugates for lectin binding, with the lectin having higher affinity for the conjugate affinity ligands than for the recognition element.
The claims cover a cross-linked material comprising conjugates with an anti-diabetic drug and multiple affinity ligands bound to a low molecular weight non-polymeric framework, cross-linked by multivalent agents through competitively dissociable non-covalent bonds. The affinity ligands and drug, particularly insulin, conformation, and the use of lectins—including chemically modified lectins—as cross-linking agents, are claimed. The structure and composition of conjugates and their specific molecular features are also claimed.
Stated Advantages
Conjugates of the invention exhibit significantly enhanced physical and chemical stability compared to native insulin under accelerated conditions.
The conjugates maintain in vivo bioactivity indistinguishable from fresh conjugates after accelerated stability testing.
Conjugates are rapidly absorbed and eliminated in vivo with pharmacokinetics similar to native insulin despite high stability and non-enzymatic degradability.
Materials formed from these conjugates and modified lectins exhibit glucose-responsive controlled release of conjugates in vitro and in vivo.
Binding-site modification of lectins reduces T-cell mitogenicity and improves safety profiles while retaining functional cross-linking ability with conjugates.
The low molecular weight of conjugates allows formulation into injectable dispersions with high drug loading, suitable for large-scale manufacturing and administration using standard syringes and needles.
Documented Applications
Use of cross-linked materials for controlled release of conjugates in response to target molecules, particularly glucose-responsive drug delivery materials for diabetes treatment.
Application of conjugates and materials in therapeutic drug delivery, including insulin and other anti-diabetic drugs for physiological regulation.
Use in in vivo treatments of diabetes and hyperglycemia by subcutaneous administration to achieve glucose-responsive insulin delivery.
Use in in vitro or in vivo chemical sensors including fluorescence resonance energy transfer (FRET)-based glucose sensors and viscosity-based glucose sensors.
Use of modified lectins as multivalent cross-linking agents with reduced mitogenicity and improved safety for forming glucose-responsive materials.
Use of the materials in artificial feedback systems that release bioactive conjugates in response to dynamic levels of endogenous or exogenous target molecules.
Use includes combination therapies with other anti-diabetic agents such as insulin sensitizers and insulin secretagogues.
Provided are kits comprising injectable dispersions of cross-linked materials for controlled drug delivery, facilitating repeated administration and storage.
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