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Multi-small molecule conjugations as new targeted delivery carriers for tumor therapy.

Shan L, Liu M, Wu C, Zhao L, Li S, Xu L, Cao W, Gao G, Gu Y - Int J Nanomedicine (2015)

Bottom Line: In vitro and acute toxicity studies demonstrated the low toxicity of the prodrug formulations compared with the free drug.Notably, compared with free PTX and with PTX-loaded macromolecular carriers from our previous study, this multi-small molecule-conjugated strategy improved the water solubility, loading rate, targeting ability, antitumor activity, and toxicity profile of PTX.These results support the use of multi-small molecules as tumor-targeting drug delivery systems.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pharmaceutical Biotechnology, School of Biology and Food Engineering, Suzhou University, Suzhou, People's Republic of China.

ABSTRACT
In response to the challenges of cancer chemotherapeutics, including poor physicochemical properties, low tumor targeting ability, and harmful side effects, we developed a new tumor-targeted multi-small molecule drug delivery platform. Using paclitaxel (PTX) as a model therapeutic, we prepared two prodrugs, ie, folic acid-fluorescein-5(6)-isothiocyanate-arginine-paclitaxel (FA-FITC-Arg-PTX) and folic acid-5-aminofluorescein-glutamic-paclitaxel (FA-5AF-Glu-PTX), composed of folic acid (FA, target), amino acids (Arg or Glu, linker), and fluorescent dye (fluorescein in vitro or near-infrared fluorescent dye in vivo) in order to better understand the mechanism of PTX prodrug targeting. In vitro and acute toxicity studies demonstrated the low toxicity of the prodrug formulations compared with the free drug. In vitro and in vivo studies indicated that folate receptor-mediated uptake of PTX-conjugated multi-small molecule carriers induced high antitumor activity. Notably, compared with free PTX and with PTX-loaded macromolecular carriers from our previous study, this multi-small molecule-conjugated strategy improved the water solubility, loading rate, targeting ability, antitumor activity, and toxicity profile of PTX. These results support the use of multi-small molecules as tumor-targeting drug delivery systems.

No MeSH data available.


Related in: MedlinePlus

Synthetic scheme and structures of FA-FITC-Arg-PTX (A) and FA-5AF-Glu-PTX (B).Notes: (A) a, Fmoc-Arg(Pbf)-PTX; b, FA-NHS; c, NH2- Arg(Pbf)-PTX; d, FA- Arg(Pbf)-PTX; e, FA- (NH2)Arg-PTX; f, FA-FITC-Arg-PTX. (B) a, Fmoc-Glu(tBu)-PTX; b, FA-NHS; c, NH2-Glu(tBu)-PTX; d, FA- Glu(tBu)-PTX; e, FA-(COOH) Glu-PTX; f, FA-5AF-Glu-PTX.Abbreviations: 5AF, 5-aminofluorescein; FA, folic acid; FITC, fluorescein isothiocyanate; PTX, paclitaxel; h, hour; min, minute; RT, room temperature.
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f1-ijn-10-5571: Synthetic scheme and structures of FA-FITC-Arg-PTX (A) and FA-5AF-Glu-PTX (B).Notes: (A) a, Fmoc-Arg(Pbf)-PTX; b, FA-NHS; c, NH2- Arg(Pbf)-PTX; d, FA- Arg(Pbf)-PTX; e, FA- (NH2)Arg-PTX; f, FA-FITC-Arg-PTX. (B) a, Fmoc-Glu(tBu)-PTX; b, FA-NHS; c, NH2-Glu(tBu)-PTX; d, FA- Glu(tBu)-PTX; e, FA-(COOH) Glu-PTX; f, FA-5AF-Glu-PTX.Abbreviations: 5AF, 5-aminofluorescein; FA, folic acid; FITC, fluorescein isothiocyanate; PTX, paclitaxel; h, hour; min, minute; RT, room temperature.

Mentions: FA-Arg(Pbf)-PTX and FA-Glu(OtBu)-PTX were synthesized in four steps, as shown in Figure 1Aa–Ad and Ba–Bd, respectively. Briefly, PTX (100 mg, 0.117 mmol, 1 equivalent) and Fmoc-Arg(Pbf)-OH (91.08 mg, 0.14 mmol, 1.2 equivalents) were dissolved in 10 mL of CH2Cl2, and 4-dimethylaminopyridine (14.27 mg, 0.117 mmol, 1 equivalent) was subsequently added (Figure 1Aa). Cold EDC (44.85 mg, 0.234 mmol, 2 equivalent, 5mL of CH2Cl2) was added dropwise to the mixture over 20 minutes and stirred at room temperature for 22 hours. The reaction mixture was diluted further with 15 mL of CH2Cl2. The organic layer was washed with water, saturated aqueous NaHCO3, and dried over MgSO4. The residue obtained after evaporation (vacuum) of the organic solvent was purified by recrystallization from diethyl ether. The purified Fmoc-Arg(Pbf)-PTX was obtained as a white solid in 85% yield. This product was used in the next step without further characterization. Next, 44.14 mg of FA (0.1 mmol) in 5 mL of dimethyl sulfoxide was activated with dicyclohexylcarbodiimide and NHS (molar ratio of FA to dicyclohexylcarbodiimide to NHS, 1:1.5:1.5) at room temperature in the dark for 5 hours (Figure 1Ab). The residue was removed by filtration under reduced pressure, and the activated FA was extracted with anhydrous acetone. Fmoc-Arg(Pbf)-PTX (150.3 mg, 0.1 mmol) was dissolved in 10 mL of CH2Cl2, 2 mL of 2-pipecoline was added to the solvent for deprotection, and the mixture was stirred at room temperature for 10 hours (Figure 1Ac). The residue obtained after evaporation (vacuum) of the organic solvent was purified by recrystallization from diethyl ether. The purified NH2-Arg(Pbf)-PTX was obtained as a white solid in 95% yield. The molecular weight of the purified NH2-Arg(Pbf)-PTX was determined by liquid chromatography mass spectrometry (LC-MS), NH2-Arg[Pbf]-PTX: MS (electrospray ionization [ESI], m/z): 1,279.43 ([M + H]+), NH2-Glu(tBut)-PTX: MS (ESI, m/z): 1,056.15 ([M + H]+). NH2-Arg(Pbf)-PTX (127.94 mg, 0.1 mmol) was dissolved in dimethyl sulfoxide, and the activated FA-NHS was added to the mixture, which was then stirred at room temperature in the dark for 24 hours (Figure 1Ad). The reaction mixture was concentrated in a vacuum and purified on a silica gel column. The purified FA-Arg(Pbf)-PTX was obtained as a yellow solid in 85% yield. A procedure similar to that described above was also applied to synthesize the control FA-Glu(OtBu)-PTX (Figure 1Ba–Bd). These two products were used in the next step without further characterization.


Multi-small molecule conjugations as new targeted delivery carriers for tumor therapy.

Shan L, Liu M, Wu C, Zhao L, Li S, Xu L, Cao W, Gao G, Gu Y - Int J Nanomedicine (2015)

Synthetic scheme and structures of FA-FITC-Arg-PTX (A) and FA-5AF-Glu-PTX (B).Notes: (A) a, Fmoc-Arg(Pbf)-PTX; b, FA-NHS; c, NH2- Arg(Pbf)-PTX; d, FA- Arg(Pbf)-PTX; e, FA- (NH2)Arg-PTX; f, FA-FITC-Arg-PTX. (B) a, Fmoc-Glu(tBu)-PTX; b, FA-NHS; c, NH2-Glu(tBu)-PTX; d, FA- Glu(tBu)-PTX; e, FA-(COOH) Glu-PTX; f, FA-5AF-Glu-PTX.Abbreviations: 5AF, 5-aminofluorescein; FA, folic acid; FITC, fluorescein isothiocyanate; PTX, paclitaxel; h, hour; min, minute; RT, room temperature.
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Related In: Results  -  Collection

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f1-ijn-10-5571: Synthetic scheme and structures of FA-FITC-Arg-PTX (A) and FA-5AF-Glu-PTX (B).Notes: (A) a, Fmoc-Arg(Pbf)-PTX; b, FA-NHS; c, NH2- Arg(Pbf)-PTX; d, FA- Arg(Pbf)-PTX; e, FA- (NH2)Arg-PTX; f, FA-FITC-Arg-PTX. (B) a, Fmoc-Glu(tBu)-PTX; b, FA-NHS; c, NH2-Glu(tBu)-PTX; d, FA- Glu(tBu)-PTX; e, FA-(COOH) Glu-PTX; f, FA-5AF-Glu-PTX.Abbreviations: 5AF, 5-aminofluorescein; FA, folic acid; FITC, fluorescein isothiocyanate; PTX, paclitaxel; h, hour; min, minute; RT, room temperature.
Mentions: FA-Arg(Pbf)-PTX and FA-Glu(OtBu)-PTX were synthesized in four steps, as shown in Figure 1Aa–Ad and Ba–Bd, respectively. Briefly, PTX (100 mg, 0.117 mmol, 1 equivalent) and Fmoc-Arg(Pbf)-OH (91.08 mg, 0.14 mmol, 1.2 equivalents) were dissolved in 10 mL of CH2Cl2, and 4-dimethylaminopyridine (14.27 mg, 0.117 mmol, 1 equivalent) was subsequently added (Figure 1Aa). Cold EDC (44.85 mg, 0.234 mmol, 2 equivalent, 5mL of CH2Cl2) was added dropwise to the mixture over 20 minutes and stirred at room temperature for 22 hours. The reaction mixture was diluted further with 15 mL of CH2Cl2. The organic layer was washed with water, saturated aqueous NaHCO3, and dried over MgSO4. The residue obtained after evaporation (vacuum) of the organic solvent was purified by recrystallization from diethyl ether. The purified Fmoc-Arg(Pbf)-PTX was obtained as a white solid in 85% yield. This product was used in the next step without further characterization. Next, 44.14 mg of FA (0.1 mmol) in 5 mL of dimethyl sulfoxide was activated with dicyclohexylcarbodiimide and NHS (molar ratio of FA to dicyclohexylcarbodiimide to NHS, 1:1.5:1.5) at room temperature in the dark for 5 hours (Figure 1Ab). The residue was removed by filtration under reduced pressure, and the activated FA was extracted with anhydrous acetone. Fmoc-Arg(Pbf)-PTX (150.3 mg, 0.1 mmol) was dissolved in 10 mL of CH2Cl2, 2 mL of 2-pipecoline was added to the solvent for deprotection, and the mixture was stirred at room temperature for 10 hours (Figure 1Ac). The residue obtained after evaporation (vacuum) of the organic solvent was purified by recrystallization from diethyl ether. The purified NH2-Arg(Pbf)-PTX was obtained as a white solid in 95% yield. The molecular weight of the purified NH2-Arg(Pbf)-PTX was determined by liquid chromatography mass spectrometry (LC-MS), NH2-Arg[Pbf]-PTX: MS (electrospray ionization [ESI], m/z): 1,279.43 ([M + H]+), NH2-Glu(tBut)-PTX: MS (ESI, m/z): 1,056.15 ([M + H]+). NH2-Arg(Pbf)-PTX (127.94 mg, 0.1 mmol) was dissolved in dimethyl sulfoxide, and the activated FA-NHS was added to the mixture, which was then stirred at room temperature in the dark for 24 hours (Figure 1Ad). The reaction mixture was concentrated in a vacuum and purified on a silica gel column. The purified FA-Arg(Pbf)-PTX was obtained as a yellow solid in 85% yield. A procedure similar to that described above was also applied to synthesize the control FA-Glu(OtBu)-PTX (Figure 1Ba–Bd). These two products were used in the next step without further characterization.

Bottom Line: In vitro and acute toxicity studies demonstrated the low toxicity of the prodrug formulations compared with the free drug.Notably, compared with free PTX and with PTX-loaded macromolecular carriers from our previous study, this multi-small molecule-conjugated strategy improved the water solubility, loading rate, targeting ability, antitumor activity, and toxicity profile of PTX.These results support the use of multi-small molecules as tumor-targeting drug delivery systems.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pharmaceutical Biotechnology, School of Biology and Food Engineering, Suzhou University, Suzhou, People's Republic of China.

ABSTRACT
In response to the challenges of cancer chemotherapeutics, including poor physicochemical properties, low tumor targeting ability, and harmful side effects, we developed a new tumor-targeted multi-small molecule drug delivery platform. Using paclitaxel (PTX) as a model therapeutic, we prepared two prodrugs, ie, folic acid-fluorescein-5(6)-isothiocyanate-arginine-paclitaxel (FA-FITC-Arg-PTX) and folic acid-5-aminofluorescein-glutamic-paclitaxel (FA-5AF-Glu-PTX), composed of folic acid (FA, target), amino acids (Arg or Glu, linker), and fluorescent dye (fluorescein in vitro or near-infrared fluorescent dye in vivo) in order to better understand the mechanism of PTX prodrug targeting. In vitro and acute toxicity studies demonstrated the low toxicity of the prodrug formulations compared with the free drug. In vitro and in vivo studies indicated that folate receptor-mediated uptake of PTX-conjugated multi-small molecule carriers induced high antitumor activity. Notably, compared with free PTX and with PTX-loaded macromolecular carriers from our previous study, this multi-small molecule-conjugated strategy improved the water solubility, loading rate, targeting ability, antitumor activity, and toxicity profile of PTX. These results support the use of multi-small molecules as tumor-targeting drug delivery systems.

No MeSH data available.


Related in: MedlinePlus