The invention is in the field of inhibitors for the enzyme polyisoprenylated methylated protein methyl esterase (PMPMEase). The invention is also in the field of methods for medical treatment using inhibitors of the enzyme PMPMEase. The invention is further in the field of using PMPMEase inhibitors for diagnostic or clinical applications.
Protein polyisoprenylation and subsequent methylation are essential modifications on a significant proportion of eucaryotic proteins. The modifications are a series of post-translational modifications involving motifs such as -CAAX wherein C is cysteine, A is any aliphatic amino acid, and X is any amino acid. The modifications include polyisoprenylation of the cysteine of the -CAAX motif (on the sulfur), proteolysis of the carboxyl-terminal three amino acids (AAX), and methylation of the carboxyl group of the cysteine. In the polyisoprenylation step, a 15 carbon (trans,trans-farnesyl) or 20 carbon (all trans-geranylgeranyl)hydrocarbon group is covalently added to the protein. The prenylation pathway is shown in FIG. 1.
Proteins such as the G-gamma subunits of heterotrimeric G-proteins of the G-protein coupled receptors, nuclear lamins, and guanine nucleotide-binding proteins such as Ras are polyisoprenylated and undergo methylation. Polyisoprenylated proteins serve numerous functions in cells, including receptor signaling, vesicular trafficking, cell proliferation, differentiation and apoptosis.
The only reversible step in the process is the last step, methylation. Two enzymes mediate this final state of polyisoprenylated proteins. Polyisoprenylated protein methyl transferase (PPMTase), also known as isoprenyl carboxylmethyl transferase (ICMT), transfers a methyl group from S-adenosyl-L-methionine (SAM) to the C-terminal —COO− to form the methylated polyisoprenylated protein. PPMTase is essential to the developing embryo; knockout mice lacking PPMTase activity do not survive through mid-gestation. The second of the two enzymes, polyisoprenylated methylated protein methyl esterase (PMPMEase), hydrolyzes the methyl esters of polyisoprenylated proteins to form the original proteins with free —COO− groups.
PPMTase and PMPMEase counterbalance the effects of each other. It is conceivable that the methylated and demethylated forms of prenylated proteins may be variously preferred for functional interactions by different protein targets, thus rendering PPMTase and PMPMEase very important moderators of polyisoprenylated protein function. Accordingly, manipulation of these enzymes should render significant effects on many cellular functions.
U.S. Pat. No. 5,202,456 to Rando teaches compounds that inhibit the methylation step of the prenylation process. The compounds have the structure W-Y-Q-Z or W-Y-Z, where W is a farnesyl or geranylgeranyl group or substituted farnesyl or geranylgeranyl group, Y is certain sulfur and selenium moieties, Q is substituted hydrocarbon moieties, and Z is —COOH or salts or esters (preferably salts or esters (preferably methyl, ethyl, or propyl) thereof, —CN, or —SO3 or salts or esters (preferably methyl, ethyl, or propyl) thereof.
U.S. Pat. No. 5,574,025 to Anthony teaches compounds which inhibit the prenylation of several proteins. The compounds are inhibitors of farnesyl-protein transferase and geranylgeranyl-protein transferase and are disclosed as being useful as chemotherapeutic agents.
U.S. Pat. No. 5,705,528 to Kloog teaches farnesyl derivatives which are inhibitors for prenylated protein methyltransferase enzyme (PPMTase). These compounds also are described as useful anti-cancer agents. Kloog hypothesized that focus on the reversible methylation step in the prenylation process would have less harmful effects than the use of inhibitors for the irreversible prenylation step itself.
U.S. Pat. No. 6,372,793 to Lamango et al. teaches compounds and methods for the treatment of neurological dysfunction. The compounds reverse an imbalance in methylation/demethylation of certain proteins. The compounds are prenyl cysteine compounds and analogs.
None of the above discussed prior art have focused on inhibitors for the enzyme that demethylates the prenylated protein, PMPMEase.
It is known that PMPMEase is inhibited by various organophosphorus compounds (OPs). This is interesting for a number of reasons. Parkinson's disease (PD)-like dyskinesias of a delayed nature have been reported following intoxication with OPs. It is known that OPs inhibition of an esterase known as neuropathy target esterase (NTE) is associated with organophosphate-induced delayed neuropathy (OPIDN). OPs inhibition of NTE in experimental animals causes the degeneration of long nerves towards the cell bodies.
It is known that excessive macromolecular carboxymethylation caused by intracerebroventricular injections of SAM results in PD-like effects of an analogous time profile. The SAM-induced PD-like effects are completely blocked by polyisoprenyl-L-cysteine (PC) analogs which are modeled around the C-terminal end of prenylated proteins.
Since PPMTase and PMPMEase act in reverse to each other, inhibition of PMPMEase and thus inhibition of the demethylation of prenylated proteins could have similar effects as this excessive methylation of prenylated proteins. PMPMEase inhibitors could be useful as research chemicals and could have clinical usefulness as antineoplastic agents.
PMPMEase, through its possible regulation of the functions of various types of polyisoprenylated proteins, may exert profound effects on various intracellular events and consequently on animal physiology. Putative substrates include both heterotrimeric guanine nucleotide-binding proteins (G-proteins), monomeric G-proteins, nuclear lamins, etc. These proteins mediate processes ranging from neurotransmitter signaling, cytoskeletal and intracellular transportation functions, cell proliferation, differentiation, and apoptosis. It could be inferred from this that aberrant levels of PMPMEase activity would be expressed through disease states such as cancers, neurodegenerative, and neuropsychiatric disorders.