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Discovery and Basic Research

Category: Poster Abstract

(W1030-04-24) Diethyldithiocarbamate Copper Nanoparticle Overcomes Resistance in Cancer Therapy without Inhibiting P-glycoprotein

Wednesday, October 25, 2023
10:30 AM - 11:30 AM ET


Purpose: Drug resistance towards taxane-based chemotherapy is a major issue in the treatment of prostate cancer(1). Efforts to inhibit these transporters have been limited by toxic side effects. diethyldithiocarbamate (DDC),an active metabolite of Disulfiram (DSF) has potent anticancer activities(2). However, the poor water solubility of Cu(DDC)2 limited the applications(3). This study aims to develop an formulation of Cu(DDC)2 and explore the potential and mechanism of Cu(DDC)2 NP against drug-resistant prostate and non-small-cell lung cancer (NSCLC) cells.

Methods: Cu(DDC)2 NP were prepared with the stabilized metal ion ligand complex method (SMILE) method (4) and the stabilizer PEG2K-PLA1.8K was synthesized with the ring-opening polymerization method. The anticancer effects of Cu(DDC)2 NP were measured using
In vitro MTT assay, colony-forming, tumor spheroids, annexin V apoptosis assay and in-vivo xenograft mice models. The mechanisms of overcoming resistance by Cu(DDC)2 NP were studied using additional experiments and molecular docking(PDB ID: 3G60).

Results: The NP had a Cu(DDC)2 core surrounded by PEG-PLA stabilizers, resulting in high drug concentration(50-fold increase compared to conventional method). The NP showed a particle size of 46.6 ± 0.3 nm, and the formation of Cu(DDC)2 complex was confirmed by absorption peak and mass spectrum. The Cu(DDC)2 NP shows potential anticancer effects on drug-resistant prostate and lung cancer cell lines and their parental drug-sensitive cells. The Cu(DDC)2 NP also showed anti-cancer potential in xenograft tumor model. (Fig1-2).Mechanism results indicate that Cu(DDC)2 NP is not a substrate of P-gp and thus can avoid P-gp mediated drug efflux. Further, the Cu(DDC)2 NP does not inhibit the activity or the expression of P-gp.(Fig3)

Conclusion: Novel copper diethyldithiocarbamate nanoparticles developed in our studies can effectively treat drug-resistant cancers. This nanoparticle can overcome drug resistance without inhibiting P-gp and thus avoid implications associated with P-gp inhibitors.

References: 1.G. Sonpavde, A. Huang, L. Wang, O. Baser, R. Miao Taxane chemotherapy vs antiandrogen agents as first-line therapy for metastatic castration-resistant prostate cancer.
2.A. McMahon, W. Chen, F. Li Old wine in new bottles: advanced drug delivery systems for disulfiram-based cancer therapy J. Control. Release, 319 (2020), pp. 352-359
3.X. Kang, Y. Cai, Q. Wang, C. Wang, W. Chen, W. Yang, et al.Near-infrared light triggered activation of pro-drug combination cancer therapy and induction of immunogenic cell death. Int. J. Pharm., 607 (2021), Article 120972
4.W. Chen, W. Yang, P. Chen, Y. Huang Li F. Disulfiram copper nanoparticles prepared with a stabilized metal ion ligand complex method for treating drug-resistant prostate cancers. ACS Appl. Mater. Interfaces 2018, 10, 48, 41118–41128

Acknowledgements:This work was financially supported by the following resources: Auburn University start-up fund and Auburn University LAUNCH Innovation Grant Program (F. Li).The authors declare that there is no conflict of interest.

Fig1. Preparation, characterization and anti-cancer effects of Cu(DDC)2 NP. (A1) Schematic presentation showing the NP has a Cu(DDC)2 complex core with PEG-PLA on the surface. (A2) Drug concentration in the nanoparticles using conventional thin-film dispersion method and SMILE method. (A3) TEM photo. (B1) UV–Vis spectrum shows the formation of Cu(DDC)2 complex in the NP formulation as indicated by the absorption peak at 435 nm. (B2) The formation of Cu(DDC)2 was also confirmed with the MS analysis.(C) Summary of IC50 values and resistance index, n = 3. (D) Colony-forming assay. (E) Cytotoxicity on tumor spheroids. Scale bars = 300 μm. (D) DU145-TXR cells were treated with control (cell culture medium), Cu(DDC)2 NP (500 nM), and positive control (staurosporine 10 μM) for 5 h. (E) Cell apoptosis analyzed with flow cytometry after staining with FITC-Annexin V and propidium iodide (PI), n = 3.

Fig2. (A)Cu(DDC)2 NP overcomes drug resistance by avoiding P-gp mediated drug efflux.(B)The p-gp levels in sensitive and PTX resistant cell lines. was established with drug-resistant r cells. (C) Tumor growth of DU145-TXR prostate xenograft tumor model. (D) tumor weight; (E) photo of tumor; and (F) body weight. Data are presented as the mean ± SD, n = 4, **, P < 0.01.*, P < 0.05

Fig. 3.(A1-A2)The docking of paclitaxel and Cu(DDC)2 with P-gp (PDB code: 3G60). (A1)Paclitaxel has strong interactions with P-gp: Molecular receptor surface area of P-gp and 2D presentation of the interaction of paclitaxel with amino acid residues of P-gp. (A2)Cu(DDC)2 has negligible interaction with P-gp: Molecular receptor surface area of P-gp and 2D presentation indicates no interaction between Cu(DDC)2 with amino acid residues of P-gp.
(B1). Schematic presentation showing the effects of different treatments on the P-gp activities. (B2) P-gp activities of DU145-TXR cells treated with control, a P-gp inhibitor (lapatinib, 5 μM), and Cu(DDC)2 NP (500 nM) were determined with calcein-AM as a P-gp substrate. (B3) P-gp activities/calcein AM accumulation in DU145-TXR tumor spheroid. Scale bars = 1 mm. (B4) Western blot analysis of P-gp protein levels in cultured DU145-TXR cells and in vivo tumor tissues treated with Cu(DDC)2 NP. Data are presented as the mean ± SD, n = 3, ***, P < 0.001.