title: Potentiating the cellular targeting and anti-tumor activity of Dp44mT via binding to human serum albumin: two saturable mechanisms of Dp44mT uptake by cells.
abstract: Di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT) demonstrates potent anti-cancer activity. We previously demonstrated that 14C-Dp44mT enters and targets cells through a carrier/receptor-mediated uptake process. Despite structural similarity, 2-benzoylpyridine 4-ethyl-3-thiosemicarbazone (Bp4eT) and pyridoxal isonicotinoyl hydrazone (PIH) enter cells via passive diffusion. Considering albumin alters the uptake of many drugs, we examined the effect of human serum albumin (HSA) on the cellular uptake of Dp44mT, Bp4eT and PIH. Chelator-HSA binding studies demonstrated the following order of relative affinity: Bp4eT≈PIH>Dp44mT. Interestingly, HSA decreased Bp4eT and PIH uptake, potentially due to its high affinity for the ligands. In contrast, HSA markedly stimulated Dp44mT uptake by cells, with two saturable uptake mechanisms identified. The first mechanism saturated at 5-10 µM (B(max):1.20±0.04 × 10⁷ molecules/cell; K(d):33±3 µM) and was consistent with a previously identified Dp44mT receptor/carrier. The second mechanism was of lower affinity, but higher capacity (B(max):2.90±0.12 × 10⁷ molecules/cell; K(d):65±6 µM), becoming saturated at 100 µM and was only evident in the presence of HSA. This second saturable Dp44mT uptake process was inhibited by excess HSA and had characteristics suggesting it was mediated by a specific binding site. Significantly, the HSA-mediated increase in the targeting of Dp44mT to cancer cells potentiated apoptosis and could be important for enhancing efficacy.
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Angelica M
authors.initials: AM
authors.lastname: Merlot
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Sumit
authors.initials: S
authors.lastname: Sahni
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Darius J R
authors.initials: DJ
authors.lastname: Lane
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Ashleigh M
authors.initials: AM
authors.lastname: Fordham
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Namfon
authors.initials: N
authors.lastname: Pantarat
authors.affiliation: Faculty of Pharmacy, The University of Sydney, Sydney, NSW, Australia.
authors.forename: David E
authors.initials: DE
authors.lastname: Hibbs
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Vera
authors.initials: V
authors.lastname: Richardson
authors.affiliation: Faculty of Pharmacy, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Munikumar R
authors.initials: MR
authors.lastname: Doddareddy
authors.affiliation: Faculty of Pharmacy, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Ong
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Michael L H
authors.initials: ML
authors.lastname: Huang
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Des R
authors.initials: DR
authors.lastname: Richardson
authors.affiliation: Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Danuta S
authors.initials: DS
authors.lastname: Kalinowski
mesh_terms: D000970:Antineoplastic Agents / Q000493:pharmacokinetics / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D004354:Drug Screening Assays, Antitumor; D056945:Hep G2 Cells; D006801:Humans; D061986:MCF-7 Cells; D008958:Models, Molecular; D012709:Serum Albumin / Q000378:metabolism*; D013882:Thiosemicarbazones / Q000493:pharmacokinetics / Q000494:pharmacology*
chemical_list: D000970:Antineoplastic Agents; D012709:Serum Albumin; D013882:Thiosemicarbazones; C539263:di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone
keywords: Dp44mT; albumin; anti-tumor targeting; human serum albumin
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