Assay method, peptides and reagent kits for testing mrp-specific inhibitors
OwnerEva Bakos, Balazs Sarkadi +3inventors
InventorsEva Bakos, Balazs Sarkadi, Andras Varadi, Gabor Tusnadi, Raymond Evers
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Abstract
The invention relates to a method for testing L0-specific inhibitors of transporter proteins of the MRP-family and for testing inhibitors specific to the predicted amphipatic helical region of the L0 region, the tested inhibitors L0 peptide, mutants thereof, polynucleotides coding the peptides. Vectors and hosts cells comprising the polynucleotides and reagent kits for testing and developing the inhibitors. The invention is especially useful for combatting multidrug resistance in cancer patients and in the field of related research.
Claims18
An assay method for testing L0-specific inhibitors of transporter proteins of the MRP family, comprising assaying an L0 peptide exposed to the effect of a potential inhibitor, to determine whether at least one L0-function was retained or at least partially lost.
A method of any of clams 3 or 4 comprising assaying L0 peptide to determine whether at least one of the following functions was retained:
- membrane binding, rescuing the core region of the protein to the physiological membrane compartment and/or restoring transporter protein function.
An assay method for screening potential inhibitors a transporter protein of the MRP family said inhibitor being specific for a segment of the L0 region predicted to be an amphipatic helix comprising determining whether the potential inhibitor exhibits affinity for the segment predicted to be an amphipatic helix.
An assay method for testing inhibitors of MRP family transporter proteins specific for an L0 region segment predicted to be an amphipatic helix, comprising assaying a transporter protein of the MRP family having exposed to the effect of a potential inhibitor;
- a functional transporter protein fragment comprising L0 region;
- or an L0 peptide and a protein consisting of a transporter protein core region (or a functional fragment thereof) said peptide and said protein being not linked covalently;
- a) in respect of specific membrane routing and/or glycosilation of the protein or its fragment, and
- b) in respect of any other function, preferably transporting capability of the protein, membrane binding of the L0 peptide and/or MRP1 function restoring capability of the, L0 peptide, and
- if the assayed functions of a) are retained and if the assayed functions of b) are at least partially lost considering the result as the selective inhibiting effect of the inhibitor specific for the segment predicted to be an amphipatic helix.
L0 peptide or a mutant or homologue thereof retaining at least partly at least one L0-function, for use in testing, designing or developing L0-specific inhibitors of transporter proteins of the MRP family, in multidrug resistance research or as a validated drug target.
Mutant L0 peptide carrying a mutation in the peptide segment predicted to be an amphipatic helix, preferably in the segment consisting of amino acids 221 to 227, said mutation being preferably a deletion or a substitution.
A polynucleotide comprising a sequence coding L0 region of a transporter protein of the MRP family for use in expressing L0 peptide.
Dependencies are inferred from the claim text.