Methyl (E)-(3-(3,4-dihydroxyphenyl)acryloyl)tryptophanate (MHAT) is an O-methyl ester of javamide-II showing strong anti-inflammatory activity. Therefore, in this study, MHAT was chemically synthesized, and its effects on p38 MAP kinase, NF-κB, and monocyte chemotactic factor-1 (MCP-1) expression were investigated in LPS-stimulated differentiated THP-1 cells. MHAT inhibited p38 MAP kinase with an IC50 of 12 μM, and the inhibition was supported by an in silico model showing that its binding to p38 MAP kinase was stronger than that of SB203580. At the concentration of 20 μM, the p38 inhibition reduced ATF-2 phosphorylation by 55% (P < 0.05). Additionally, MHAT inhibited NF-κB (p65) phosphorylation by 30% (P < 0.05) at the same concentration, suggesting that MHAT was able to reduce NF-κB transcriptional activity. This supposition was confirmed by the NF-κB reporter assay, demonstrating that MHAT (20 μM) could suppress NF-κB transcriptional activity by 29% (P < 0.05) in the NF-κB reporter (Luc)-HEK293 cell line. As expected, the treatment with MHAT (5–40 μM) significantly inhibited MCP-1 mRNA expression by 9–73% (P < 0.05) and the production of MCP-1 protein by 10–70% (P < 0.05) in the THP-1 cells. Furthermore, MHAT was found to inhibit RANTES expression as well in the same THP-1 cells, supporting its purported inhibition of p38 MAP kinase and NF-κB. All these data suggest that MHAT is a potent compound that can inhibit MCP-1 production by suppressing p38 kinase/ATF-2 phosphorylation and NF-κB in the differentiated THP-1 cells.