A novel peptide with active tumor targeting and pH-sensitive cell membrane penetration

Abstract

The invention provides a novel polypeptide which not only can be actively targeted to highly expressed integrin receptor of tumor cells, but also can efficiently mediate cell entry. The novel polypeptide TR(c(RGDfK)-AGYLLGHINLHHLAHL(Aib)HHIL-Cys) is formed by connecting cyclic RGD peptide (c(RGDfK)) of specificity-targeted integrin receptor and non-specific pH sensitive cell penetrating peptide TH(AGYLLGHINLHHLAHL(Aib)HHIL-Cys) through covalent bonds. The novel polypeptide TR is modified onto the surface of a drug carrier system, and can mediate the drug carrier system to more efficiently deliver to the inside of the tumor, so that the tumor can be more effectively treated.

Description

technical field

[0001] The invention relates to the technical field of pharmaceutical preparations. Specifically, the present invention relates to a novel polypeptide that has the ability to actively target integrin receptors highly expressed in tumor cells and has the ability to penetrate pH-sensitive cells. The novel polypeptide is modified on the surface of the drug-carrying system to mediate the efficient and active targeting of the drug-carrying system to tumors with high expression of integrin receptors and efficiently mediate its entry into cells, so as to achieve the purpose of improving the therapeutic effect of tumors. Background technique

[0002] 1. The morbidity and mortality of cancer have been increasing year by year in recent years, which has seriously threatened human health. Although there are many ways to treat cancer, chemotherapy is still the most commonly used. Because chemotherapy drugs often lack selectivity between normal tissue and tumor tissue, t...

Examples

Embodiment 1

[0019] Preparation and Characterization of TR Peptide Modified Liposomes (TR-Lip)

[0020] Liposomes were prepared by thin film dispersion method. Weigh 1.2 mg soybean lecithin, 0.4 mg cholesterol, 0.05 mg monomethoxy-modified polyethylene glycol and 1 mg TR peptide-modified polyethylene glycol, dissolve them all in chloroform / methanol mixture, rotate and evaporate , form a thin film layer, and place it in a vacuum desiccator overnight, so that the organic solvent is fully evaporated. Add 1ml of Hepes buffer solution (pH 7.4), shake on a shaker (180 rpm×30 min, 37°C), and ultrasonicate (5 times×5 s×15 s) to obtain TR with uniform particle size. -Lip. Take 100 μl of TR-Lip and dilute it 10 times, and measure its particle size distribution and charge distribution with a Zetanano particle size analyzer. The particle size of TR-Lip is around 150 nm. At pH 7.4, the Zeta potential of TR-Lip was around -7 mV; at pH 6.5, the Zeta potential of TR-Lip was around +5 mV. Surface plas...

Embodiment 2

[0022] Preparation and characterization of TR peptide-modified nanoparticles (TR-NPs)

[0023] TR peptide-modified nanoparticles were prepared by W / O / W emulsification-solvent evaporation method. Dissolve a certain amount of PLA-PLL-TR in dichloromethane to form an organic phase, use pure water as the water phase, mix the organic phase and the water phase in a certain proportion, and ultrasonically form colostrum. Pour the colostrum into the external water phase containing the poloxamer F-18 emulsifier, continue to sonicate to obtain the double emulsion, put it in a beaker, stir and evaporate the organic solvent to obtain a nanoparticle colloidal dispersion system. Zetanano particle size analyzer was used to measure its particle size distribution and charge distribution; SPR instrument was used to measure its binding force with integrin. The particle size of TR-NPs is about 90 nm. At pH 7.4, the Zeta potential of TR-NPs was about -7.5 mV; at pH 6.5, the Zeta potential of TR-N...

Embodiment 3

[0025] Preparation and characterization of TR peptide-modified micelles (TR-micelle)

[0026]Take 10 mg DSPE-PEG-TR and 20 mg DSPE-PEG-OMe respectively and dissolve them in anhydrous chloroform, rotary evaporate to form a film, dry under reduced pressure overnight, add 5 ml of 20 mM Hepes buffer (pH 7.4), shake in a water bath at 37 °C After 20 min, filter with a 0.22 μm microporous membrane to obtain TR peptide-modified polymer micelles (TR-micelle). Zetanano particle size analyzer was used to measure its particle size distribution and charge distribution; SPR instrument was used to measure its binding force with integrin. The particle size of TR-micelle is about 112 nm. At pH 7.4, the Zeta potential of TR-micelle was about -8 mV; at pH 6.5, the Zeta potential of TR-micelle was about +4 mV. Surface plasmon resonance (SPR) was used to measure the binding ability of micelles modified by different polypeptides to integrin, and the binding ability of TR-micelle to integrin was ...

technical field

[0001] The invention relates to the technical field of pharmaceutical preparations. Specifically, the present invention relates to a novel polypeptide that has the ability to actively target integrin receptors highly expressed in tumor cells and has the ability to penetrate pH-sensitive cells. The novel polypeptide is modified on the surface of the drug-carrying system to mediate the efficient and active targeting of the drug-carrying system to tumors with high expression of integrin receptors and efficiently mediate its entry into cells, so as to achieve the purpose of improving the therapeutic effect of tumors. Background technique

[0002] 1. The morbidity and mortality of cancer have been increasing year by year in recent years, which has seriously threatened human health. Although there are many ways to treat cancer, chemotherapy is still the most commonly used. Because chemotherapy drugs often lack selectivity between normal tissue and tumor tissue, t...

Examples

Embodiment 1

[0019] Preparation and Characterization of TR Peptide Modified Liposomes (TR-Lip)

[0020] Liposomes were prepared by thin film dispersion method. Weigh 1.2 mg soybean lecithin, 0.4 mg cholesterol, 0.05 mg monomethoxy-modified polyethylene glycol and 1 mg TR peptide-modified polyethylene glycol, dissolve them all in chloroform / methanol mixture, rotate and evaporate , form a thin film layer, and place it in a vacuum desiccator overnight, so that the organic solvent is fully evaporated. Add 1ml of Hepes buffer solution (pH 7.4), shake on a shaker (180 rpm×30 min, 37°C), and ultrasonicate (5 times×5 s×15 s) to obtain TR with uniform particle size. -Lip. Take 100 μl of TR-Lip and dilute it 10 times, and measure its particle size distribution and charge distribution with a Zetanano particle size analyzer. The particle size of TR-Lip is around 150 nm. At pH 7.4, the Zeta potential of TR-Lip was around -7 mV; at pH 6.5, the Zeta potential of TR-Lip was around +5 mV. Surface plas...

Embodiment 2

[0022] Preparation and characterization of TR peptide-modified nanoparticles (TR-NPs)

[0023] TR peptide-modified nanoparticles were prepared by W / O / W emulsification-solvent evaporation method. Dissolve a certain amount of PLA-PLL-TR in dichloromethane to form an organic phase, use pure water as the water phase, mix the organic phase and the water phase in a certain proportion, and ultrasonically form colostrum. Pour the colostrum into the external water phase containing the poloxamer F-18 emulsifier, continue to sonicate to obtain the double emulsion, put it in a beaker, stir and evaporate the organic solvent to obtain a nanoparticle colloidal dispersion system. Zetanano particle size analyzer was used to measure its particle size distribution and charge distribution; SPR instrument was used to measure its binding force with integrin. The particle size of TR-NPs is about 90 nm. At pH 7.4, the Zeta potential of TR-NPs was about -7.5 mV; at pH 6.5, the Zeta potential of TR-N...

Embodiment 3

[0025] Preparation and characterization of TR peptide-modified micelles (TR-micelle)

[0026]Take 10 mg DSPE-PEG-TR and 20 mg DSPE-PEG-OMe respectively and dissolve them in anhydrous chloroform, rotary evaporate to form a film, dry under reduced pressure overnight, add 5 ml of 20 mM Hepes buffer (pH 7.4), shake in a water bath at 37 °C After 20 min, filter with a 0.22 μm microporous membrane to obtain TR peptide-modified polymer micelles (TR-micelle). Zetanano particle size analyzer was used to measure its particle size distribution and charge distribution; SPR instrument was used to measure its binding force with integrin. The particle size of TR-micelle is about 112 nm. At pH 7.4, the Zeta potential of TR-micelle was about -8 mV; at pH 6.5, the Zeta potential of TR-micelle was about +4 mV. Surface plasmon resonance (SPR) was used to measure the binding ability of micelles modified by different polypeptides to integrin, and the binding ability of TR-micelle to integrin was ...

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