We have continued our collaboration with Dr. Robert Hoffman at Anticancer, Inc. in San Diego. The methionine addiction of cancer cells is known as the Hoffman effect. While non-cancer cells in culture can utilize homocysteine in place of methionine for cellular growth, most cancer cells require exogenous methionine for proliferation. It has been suggested that a biochemical basis of this effect is the increased utilization of methionine for S-adenosylmethionine, the major methyl donor for a variety of cellular methyltransferases. Recent studies have pointed to the role of S-adenosylmethionine-dependent protein arginine methyltransferases (PRMTs) in cell proliferation and cancer. To further understand the biochemical basis of the methionine addiction of cancer cells, we compared protein arginine methylation in two previously described isogenic cell lines, a methionine-addicted 143B human osteosarcoma cell line and its less methionine-dependent revertant. We utilized antibodies to detect the asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) products of PRMTs in polypeptides from cellular extracts and purified histone preparations of these cell lines fractionated by SDS-PAGE. Importantly, we observed little to no differences in the banding patterns of ADMA- and SDMA-containing species between the osteosarcoma parental and revertant cell lines. Furthermore, enzymatic activity assays using S-adenosyl-ʟ-[methyl-3H] methionine, recombinantly purified PRMT enzymes, cell lysates, and specific PRMT inhibitors revealed no major differences in radiolabeled polypeptides on SDS-PAGE gels. Taken together, these results suggest that changes in protein arginine methylation may not be major contributors to the Hoffman effect and that other consequences of methionine addiction may be more important in the metastasis and malignancy of osteosarcoma and potentially other cancers.

Publication: Holtz, A. G., Lowe, T. L., Aoki, Y., Kubota, Y., Hoffman, R. M., and Clarke, S. G. (2023) “Asymmetric and Symmetric Protein Arginine Methylation in Methionine-Addicted Human Cancer Cells” PLoS ONE 18(12): e0296291. (DOI: 10.1371/journal.pone.0296291). PMCID: PMC10745221.

We are following up on this study to determine the mechanism of the Hoffman effect. We have begun to analyze the differences in the exomic DNA sequences, mRNA expression, and metabolomics of the methionine-dependent and methionine-independent cell lines. The goal is to understand how cancer cells use methylation pathways to enhance their survival. These studies may well suggest therapeutic approaches to controlling human tumor growth.