Pyridoxal-5’-phosphate (PLP, the active form of vitamin B6) is an essential cofactor in numerous biochemical pathways, including amino acid metabolism, glycogenolysis, and one-carbon transfers. In cancer cells, both PLP and PLP-dependent enzymes promote survival and proliferation, making PLP biosynthesis a promising target for therapeutic intervention. In humans, PLP is synthesized from dietary precursors via a salvage pathway involving two key enzymes: pyridoxal kinase (PDXK) and pyridox(am)ine-5’-phosphate oxidase (PNPO). These enzymes are frequently upregulated in aggressive tumor types, including breast, ovarian, and hematologic cancers. Silencing PDXK or PNPO in cellular and mouse models has been shown to reduce cancer cell viability and migration. Our strategy focuses on chemically inhibiting PNPO as a novel antineoplastic approach. PNPO is particularly suitable for drug targeting due to its susceptibility to feedback inhibition by PLP. We screened various PLP analogues on recombinant human PNPO and identified PLP-rhodanine as a potent mixed-type inhibitor. We further characterized its binding profile and proposed a plausible metabolic precursor. In parallel, we investigated the off-target effects of hydrazide-containing drugs—such as isoniazid, carbidopa, phenelzine, and hydralazine—which are known to deplete PLP independently of their primary therapeutic functions. We hypothesize that these compounds form adducts with endogenous PLP, not only depleting PLP levels but also directly inhibiting PNPO. To test this, we synthesized and studied two such adducts: PLP-isoniazid, which acts as a potent mixed-type inhibitor, and PLP-carbidopa, which displays weaker, noncompetitive inhibition. These findings suggest a dual strategy: direct PNPO inhibition via rationally designed analogues and the potential repurposing of existing hydrazide drugs as anticancer agents. Ongoing studies in cellular models will further evaluate their therapeutic potential.
Inhibiting pyridoxal 5'-phosphate biosynthesis as an antitumor approach / Graziani, Claudio; Antonelli, Lorenzo; Fiorillo, Annarita; Ilari, Andrea; Vetica, Fabrizio; Di Salvo, Martino Luigi; Paiardini, Alessandro; Tramonti, Angela; Contestabile, Roberto. - (2025). (Intervento presentato al convegno 63th national congress of the Italian Society for Biochemistry and Molecular Biology tenutosi a Palermo, Italy).
Inhibiting pyridoxal 5'-phosphate biosynthesis as an antitumor approach
Claudio Graziani;Lorenzo Antonelli;Annarita Fiorillo;Fabrizio Vetica;Martino Luigi Di Salvo;Alessandro Paiardini;Angela Tramonti;Roberto Contestabile
2025
Abstract
Pyridoxal-5’-phosphate (PLP, the active form of vitamin B6) is an essential cofactor in numerous biochemical pathways, including amino acid metabolism, glycogenolysis, and one-carbon transfers. In cancer cells, both PLP and PLP-dependent enzymes promote survival and proliferation, making PLP biosynthesis a promising target for therapeutic intervention. In humans, PLP is synthesized from dietary precursors via a salvage pathway involving two key enzymes: pyridoxal kinase (PDXK) and pyridox(am)ine-5’-phosphate oxidase (PNPO). These enzymes are frequently upregulated in aggressive tumor types, including breast, ovarian, and hematologic cancers. Silencing PDXK or PNPO in cellular and mouse models has been shown to reduce cancer cell viability and migration. Our strategy focuses on chemically inhibiting PNPO as a novel antineoplastic approach. PNPO is particularly suitable for drug targeting due to its susceptibility to feedback inhibition by PLP. We screened various PLP analogues on recombinant human PNPO and identified PLP-rhodanine as a potent mixed-type inhibitor. We further characterized its binding profile and proposed a plausible metabolic precursor. In parallel, we investigated the off-target effects of hydrazide-containing drugs—such as isoniazid, carbidopa, phenelzine, and hydralazine—which are known to deplete PLP independently of their primary therapeutic functions. We hypothesize that these compounds form adducts with endogenous PLP, not only depleting PLP levels but also directly inhibiting PNPO. To test this, we synthesized and studied two such adducts: PLP-isoniazid, which acts as a potent mixed-type inhibitor, and PLP-carbidopa, which displays weaker, noncompetitive inhibition. These findings suggest a dual strategy: direct PNPO inhibition via rationally designed analogues and the potential repurposing of existing hydrazide drugs as anticancer agents. Ongoing studies in cellular models will further evaluate their therapeutic potential.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
