Diastereomer-Specific Effects of Double-Stranded Peptides Conjugated with -L-Tyr-L-Phe- or -L-Tyr-D-Phe- Residues on Tyrosine Phosphorylation and Inhibition of srctsNRK, A431, MCF-7, and DU145 Cell Growth

Abstract

Signal transduction factors that mediate cell division and growth such as protein tyrosine and serine kinases play important roles in many proliferative diseases. Binding of ligands to specific cell-surface receptors leads to aberrant signals, such as gene deletion and abnormal protein function, which lead to abnormal proliferation. 1,2) Better understanding these pathways can potentially aid in designing drugs that inhibit malignant neoplastic proliferation. Tyrosine (Tyr), with its phenol side chain, provides a phosphorylation site for tyrosine kinases, which play an important role in signal transduction (such as SH2 and SH3 domains).3) From a pharmacological viewpoint, drugs which contain Tyr residues in their structure compete for tyrosine kinase phosphorylation and may affect the signal transduction of living cells.Recently, we reported the relationship between optical isomers and growth inhibition activity by using a set of parallel, double-stranded peptides, bis(y-Phe-x-Phe) 2 -spacer(S) (7) (symbols x and yϭL-or D-configurations, AAϭamino acid, and Sϭ(CH 2 ) 12 , with two Phe residues conjugated to a polyamine spacer as a model of neoplastic targeting. [4][5][6] In order to elucidate the relationship between chirality and effect of Tyr on cell growth inhibition by the double-stranded peptides, we synthesized the bis(y-Tyr-x-Phe) 2 -(CH 2 ) 12 (6) replacing y-Phe-x-Phe with y-Tyr-x-Phe sequence. To elucidate the mechanism of action of the compound, we assayed (i) the change in the level of tyrosine phosphorylation of total proteins of src ts NRK cells, and (ii) the inhibition of epidermal growth factor receptor protein tyrosine kinase domain (EGFRKD) autophosphorylation 7,8) following treatment with 6 or 7. The proliferation of these cells is regulated by tyrosine phosphorylation and dephosphorylation, by tyrosine kinases and tyrosine phosphatases, respectively.The results showed that bis(L-Tyr-L-Phe)-N,N-dodecane-1,12-diamine (6a) and bis(L-Tyr-L-Ile)-N,N-dodecane-1,12-diamine (6e), containing the L-Tyr-L-Phe and L-Tyr-L-Ile sequences, markedly increased the tyrosine phosphorylation of 32-34 kDa proteins in src ts NRK cells. However, the profiles of the total phosphotyrosine of proteins from cells treated with 7a and 7b, containing L-Phe-L-Phe and D-Phe-LPhe sequences, are strikingly different from the profiles of those treated with 6a and 6e. The action of 6a on the tyrosine phosphorylation of proteins in src ts NRK cells is very similar to that of the tyrosine kinase inhibitor, erbstatin.9) The EGF-R protein tyrosine kinase, a transmembrane glycoprotein, is one target for the treatment of proliferative diseases. 8,10) Our results show that compounds 6a and 6b are inhibitors of EGF-R tyrosine phosphorylation.Thus, we believe that the double-stranded peptides, 6a, b and 6c, d, can be used to control the inhibitory effect on proliferation shown by both diastereomers, (D-, L-) and (L-, D-), and we describe new insights which could explain the different inhibitory effects on proliferation of Phe and Tyr, (...

Potent 2,2-Diphenyl-1-picrylhydrazyl Radical-Scavenging Activity of Novel Antioxidants, Double-Stranded Tyrosine Residues Conjugating Pyrocatechol

“…19,20) Boc (Boc, tert-butoxycarbonyl)-protected tyrosine residue, Boc-LTyr(OBzl) was conjugated to a triethylenedioxide spacer to yield protected 2a by the C-activating method using N,N-carbonyldiimidazole (CDI) in dry CHCl 3 (Chart 1). The crude compound 2a was purified using chromatography over neu-tral silica gel and the Boc-protecting group was removed by treatment with 90% trifluoroacetic acid (TFA) to produce the free form 3a.…”

“…19,20) The peptide and related ligands were detected on thin-layer chromatography (TLC) plates using iodine vapor or UV absorption. Silica gel column chromatography was performed on silica gel 60N (100 mesh, neutral; Kanto Chemical Co., Tokyo, Japan).…”

Development of New Double-Stranded Phenylalanyl Chelators Using .ETA.-.CHI. Diagrams and Binding Constants for Chelators and Lanthanide Ions

“…Removal of the Boc groups by trifluoroacetic acid (TFA) afforded the free compound (12a) in good yield. 16,19) Conjugated 2,3-bis(benzyloxy)benzoic acid was prepared from 2,3-dihydroxybenzoic acid, 16) with the N-terminal of 12a afforded by treatment with DEPC (phosphorocyanidate) on dry N,N-dimethylformamide (DMF). Deprotection of the Bzl (benzyl) group in 13a using 5% Pd/C-H 2 gave a new peptide chelator (14a) in 65% yield.…”

“…18,19) The chelator could expect flexible conformation due to a backbone conjugated with two amino acid residues to the N-terminus of a spacer, -O-CH 2 -CH 2 -O-, and its binding site is Cat, not an amido group. Chelator 1 provides a coordinate bond complex with Ln 3ϩ of the largest ionic radius in 10 mM N- (2-hydroxyethyl) Complexes of chelator 1 with Ln 3ϩ may be applied in the pharmacological field as chemotherapeutic agents and luminescent sensors, 8) which are a subject of many recent studies.…”

Design, Synthesis and Antitumor Activities of Novel Quinazolinone Derivatives as Potential EGFR Inhibitors

“…[6][7][8] Based on the key role of EGFR in antitumor effects, many small molecule inhibitors have been developed. [9][10][11][12] However, with the use of drugs, side effects and drug resistance problems have emerged. [13][14][15] Currently, EGFR small molecule inhibitors have evolved to the fourth generation.…”