Cgap-pna multivalent peptide nucleic acid ligand display
A peptide nucleic acid and amino acid technology, applied in the field of cGAP‑PNA multivalent peptide nucleic acid ligand display, can solve the problem that the number of ligands cannot be strictly limited
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[0082] In the preparation of the constructs of the invention, gamma-substituents may be introduced based on their function in the interaction with specific receptors or other biological interaction sites. In a particular PNA, more than one substituent may be utilized. In a particular L-PNA:PNA(GAP) scaffold, different L-PNAs may contain the same or different substituents depending on the target moiety.
[0083] In addition to the PNAs described above, any of a variety of PNA variations known in the art may be used. Known PNA macromolecules include macromolecules represented by the following structures.
[0084]
[0085] Natural and unnatural bases can be used in these structures and are well known to those skilled in the art.
[0086] Scheme 1 depicts a method for the synthesis of γ-substituted monomers that can be used to prepare L-PNA. The gamma substituents can serve as points for further functionalization. These monomers can be converted to L-PNA by methods known in...
Embodiment 1
[0122] This example demonstrates the generation of an initial library of ligand-modified PNA conjugates and the multivalent profile of the conjugates according to one embodiment of the invention.
[0123] To generate a multivalent library of ligand-modified PNA conjugates (L-PNA), the high-affinity AR antagonist, xanthine amine congener (XAC), was synthesized via a γ-side chain derived from lysine (γ- Lys) conjugated to PNA oligomer ( Figure 1A). Ligands attached to this side chain within the L-PNA oligomer do not interfere with the ability of L-PNA to bind complementary DNA sequences via conventional Watson Crick base pairing. A series of PNA oligomers (each consisting of 12 nucleobases) were synthesized in which one, two or three γ-Lys side chains were incorporated into the sequence ( Figures 1B-1D ). The primary amine at the end of the side chain of γ-Lys serves as the attachment point for the XAC ligand. Two mini-PEG (8-amino-3,6-dioxahanoic acid) linkers inserted bet...
Embodiment 2
[0128] This example demonstrates ligand spacing on the binding of L-PNA:DNA complexes according to one embodiment of the invention.
[0129] The initial results obtained showed that the L-PNA:DNA complex carrying two XAC ligands bound significantly better than the corresponding monovalent complex. Next, the effect of ligand spacing was assessed. Examine a series of bivalent constructs in which the two γ-Lys side chains carrying the XAC ligand are systematically displaced along the PNA backbone ( Figure 3A-3B ). To minimize the electrostatic impact of negative charges on the DNA phosphodiester backbone, the DNA was replaced with a sequence-complementary PNA. It was established that PNA:PNA duplexes maintain regular nucleobase pairing in the double helix. Experimental results suggest that DNA may have a negative effect on binding because A1 P than A1 D Effective 8-fold (p=0.0015). The resulting L-PNA:PNA duplex is charge-neutral and does not undergo charge-charge with the...
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