MilliporeSigma
  • Home
  • Search Results
  • Potentiating the cellular targeting and anti-tumor activity of Dp44mT via binding to human serum albumin: two saturable mechanisms of Dp44mT uptake by cells.

Potentiating the cellular targeting and anti-tumor activity of Dp44mT via binding to human serum albumin: two saturable mechanisms of Dp44mT uptake by cells.

Oncotarget (2015-04-08)
Angelica M Merlot, Sumit Sahni, Darius J R Lane, Ashleigh M Fordham, Namfon Pantarat, David E Hibbs, Vera Richardson, Munikumar R Doddareddy, Jennifer A Ong, Michael L H Huang, Des R Richardson, Danuta S Kalinowski
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.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Bovine Serum Albumin, heat shock fraction, pH 7, ≥98%
Sigma-Aldrich
Trichloroacetic acid, ≥99.0% (titration)
Sigma-Aldrich
Trichloroacetic acid, BioXtra, ≥99.0%
Sigma-Aldrich
Ibuprofen, ≥98% (GC)
Sigma-Aldrich
Sodium azide, ReagentPlus®, ≥99.5%
Sigma-Aldrich
Sodium azide, BioXtra
Sigma-Aldrich
Trichloroacetic acid, suitable for electrophoresis, suitable for fixing solution (for IEF and PAGE gels), ≥99%
Sigma-Aldrich
Trichloroacetic acid, ACS reagent, ≥99.0%
Sigma-Aldrich
Albumin from human serum, lyophilized powder, essentially globulin free, ≥99% (agarose gel electrophoresis)
Sigma-Aldrich
apo-Transferrin human, powder, ≥98% (agarose gel electrophoresis)
Sigma-Aldrich
Ibuprofen, meets USP testing specifications
Sigma-Aldrich
Monoclonal Anti-β-Actin antibody produced in mouse, clone AC-74, ascites fluid
Sigma-Aldrich
Trypan Blue, powder, BioReagent, suitable for cell culture
Sigma-Aldrich
Methylamine solution, 2.0 M in methanol
Sigma-Aldrich
Methylamine solution, 2.0 M in THF
Sigma-Aldrich
Trypan Blue, Dye content 60 %
Sigma-Aldrich
Trichloroacetic acid, BioUltra, ≥99.5% (T)
Sigma-Aldrich
Trichloroacetic acid, ACS reagent, for the determination of Fe in blood according to Heilmeyer, ≥99.5%
Sigma-Aldrich
Methylamine solution, 33 wt. % in absolute ethanol ((denatured with 1% toluene))
Sigma-Aldrich
Sodium pyruvate, Hybri-Max, powder, suitable for hybridoma
Sigma-Aldrich
Sodium pyruvate, powder, BioReagent, suitable for cell culture, suitable for insect cell culture, ≥99%
Sigma-Aldrich
Sodium pyruvate, powder, BioXtra, suitable for mouse embryo cell culture
Sigma-Aldrich
Sodium pyruvate, BioXtra, ≥99%
Sigma-Aldrich
Sodium pyruvate, ReagentPlus®, ≥99%
Sigma-Aldrich
Sodium azide, BioUltra, ≥99.5% (T)
Sigma-Aldrich
Sodium azide, purum p.a., ≥99.0% (T)
Sigma-Aldrich
Protease from Streptomyces griseus, powder, BioReagent, suitable for mouse embryo cell culture, ≥3.5 units/mg solid
Supelco
Ibuprofen
Sigma-Aldrich
Sodium pyruvate, anhydrous, free-flowing, Redi-Dri, ReagentPlus®, ≥99%
Sigma-Aldrich
Ethylenediaminetetraacetic acid, anhydrous, crystalline, BioReagent, suitable for cell culture