Treating stroke and other diseases without inhibiting N-type calcium channels
Inventors
Belmares, Michael P. • Garman, Jonathan David • Lu, Peter S. • Salter, Michael W. • Tymianski, Michael
Assignees
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Publication Number
US-9061070-B2
Publication Date
2015-06-23
Expiration Date
Abstract
The invention provides methods for treating stroke and compositions for use in the same. The methods employ a chimeric peptide of an active peptide and an internalization peptide. The internalization peptide is a tat variant that promotes uptake of itself and a linked active peptide into a cell without substantial binding to N-type calcium channels. Use of the tat variant allows treating of stroke free of certain side effects associated with binding to N-type calcium channels. Tat variant peptides can also be linked to other active agent for use in treating other diseases.
Core Innovation
The invention relates to chimeric agents comprising an active agent and an internalization peptide that promotes uptake of the chimeric agent into cells. The internalization peptide is described as a variant of the tat peptide YGRKKRRQRRR (SEQ ID NO:1) that has reduced capacity to bind to an N-type calcium channel relative to the tat peptide. A specific internalization peptide amino acid sequence comprising FGRKKRRQRRR (SEQ ID NO:135) is provided.
The disclosed chimeric agents are designed to address NMDAR/PSD-95 excitotoxicity in stroke pathology while aiming to avoid N-type calcium channel-mediated side effects. The document describes linking inhibitory active peptides associated with PSD-95/NMDA receptor modulation to internalization peptide variants. The central improvement is stated as reducing binding/inhibition of N-type calcium channels through the tat variant while preserving cellular uptake.
The document further describes an optional quantitative binding threshold in terms of Kd for N-type calcium channel binding, including kD greater than 10 nM, for the internalization peptide variant. The disclosure includes discussion of screening and assessment, including receptor binding assessment and functional assessment using calcium channel inhibition measurements.
Claims Coverage
The partial content identifies three independent claims. Across these claims, the main inventive features focus on a chimeric agent using a tat-peptide internalization variant with reduced N-type calcium channel binding, an isolated internalization peptide defined by a specific amino acid sequence, and an improvement in a chimeric agent that specifies that same internalization peptide sequence.
Chimeric agent with reduced N-type calcium channel binding internalization peptide
An isolated chimeric agent comprising an active agent and an internalization peptide that promotes uptake into cells, wherein the internalization peptide is a variant of the tat peptide YGRKKRRQRRR (SEQ ID NO:1) that has reduced capacity to bind to an N-type calcium channel relative to the tat peptide and the variant has an amino acid sequence comprising FGRKKRRQRRR (SEQ ID NO:135).
Isolated internalization peptide sequence
An isolated internalization peptide having an amino acid sequence comprising FGRKKRRQRRR (SEQ ID NO:135).
Improvement in chimeric agent specifying the internalization peptide sequence
In a chimeric agent comprising an internalization peptide linked to an active agent, the improvement wherein the internalization peptide has an amino acid sequence comprising FGRKKRRQRRR (SEQ ID NO:135).
Together, the independent claims define chimeric agents and internalization peptides using the specific tat variant sequence FGRKKRRQRRR (SEQ ID NO:135), with the key inventive premise that this variant provides reduced capacity to bind to an N-type calcium channel while supporting cellular uptake in chimeric constructs.
Stated Advantages
Reduced binding/inhibition of N-type calcium channels to avoid N-type channel-mediated side effects while preserving cellular uptake.
Neuroprotection in rat stroke models as indicated by reduction of infarct volume and associated functional/disability indices.
Documented Applications
Treating stroke and other CNS diseases.
Animal stroke models including MCAO and pial vessel occlusion/permanent ischemia, with infarct volume reduction and functional/disability index assessment.
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