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1.
China Pharmacy ; (12): 1431-1436, 2024.
Article Dans Chinois | WPRIM | ID: wpr-1032288

Résumé

OBJECTIVE To study the effects of transferrin-targeting peptide T7 (7pep) on intracellular transportation of polyethylene glycol-polycaprolactone (PEG-PCL) micelles in human cervical cancer HeLa cells. METHODS Using coumarin-6 (C6) as fluorescent indicator probe, both coumarin-6 (C6)-loaded PEG-PCL (PEG-PCL-C6) micelles and 7pep-modified PEG- PCL (7pep-PEG-PCL-C6) micelles were prepared by film-dispersion method. The particle size, polydispersity index and appearance morphology were compared between two types of micelles; the real-time uptake of two types of micelles by HeLa cells was compared, and the colocalization of two types of micelles with early endosomes (EE), endocytic recycling compartments (ERC) and late endosomes (LE) after entry into the cells was observed. RESULTS The particle sizes of PEG-PCL-C6 and 7pep-PEG-PCL- C6 micelles were(75.0±2.3)and(82.0±1.5)nm; the polymer dispersity indexes were 0.17±0.20 and 0.17±0.32, respectively, with a regular spherical appearance. The colocalization results showed that entry speed and amount of 7pep-PEG-PCL-C6 micelles were significantly faster/more than those of PEG-PCL-C6 micelles. 7pep-PEG-PCL-C6 micelles entered EE faster than PEG-PCL-C6 micelles, while PEG-PCL-C6 micelles entered ERC at a faster rate than 7pep-PEG-PCL-C6 micelles, and both PEG-PCL-C6 micelles and 7pep-PEG-PCL-C6 micelles tended to accumulate gradually in LE; Pearson coefficient, signal overlap ratio, and colocalization ratio of 7pep-PEG-PCL-C6 micelles with LE were significantly lower 60 minutes after entering the cell than those 30 minutes after entering the cell (P<0.05 or P<0.01). CONCLUSIONS Targeting 7pep modification can increase the entry speed and amount of PEG-PCL-C6 micelles, and also alter their intracellular transportation behavior.

2.
Chinese Journal of Biotechnology ; (12): 1138-1148, 2022.
Article Dans Chinois | WPRIM | ID: wpr-927769

Résumé

Loofah seeds ribosome inactivating protein luffin-α was fused with a tumor-targeting peptide NGR to create a recombinant protein, and its inhibitory activity on tumor cells and angiogenesis were assessed. luffin-α-NGR fusion gene was obtained by PCR amplification. The fusion gene was ligated with pGEX-6p-1 vector to create a recombinant plasmid pGEX-6p-1/luffin-α-NGR. The plasmid was transformed into E. coli BL21, and the target protein was isolated and purified by GST affinity chromatography. The luffin-α-NGR fusion gene with a full length of 849 bp was successfully obtained, and the optimal soluble expression of the target protein was achieved under the conditions of 16 ℃, 0.5 mmol/L IPTG after 16 h induction. SDS-PAGE and Western blotting confirmed the recombinant protein has an expected molecular weight of 56.6 kDa. Subsequently, the recombinant protein was de-tagged by precision protease digestion. The inhibitory effects of the recombinant protein on liver tumor cells HepG2 and breast cancer cells MDA-MB-231 were significantly stronger than that of luffin-α. The Transwell and CAM experiment proved that the recombinant protein luffin-α-NGR also had a significant inhibitory effect on tumor cells migration and neovascularization. The inhibitory activity on tumor cells and angiogenesis of the recombinant luffin-α-NGR protein lays a foundation for the development of subsequent recombinant tumor-targeting drugs.


Sujets)
Électrophorèse sur gel de polyacrylamide , Escherichia coli/métabolisme , Plasmides , Protéines recombinantes/pharmacologie , Saporines/métabolisme
3.
Cancer Research on Prevention and Treatment ; (12): 24-30, 2021.
Article Dans Chinois | WPRIM | ID: wpr-988320

Résumé

Objective To investigate the specificity and targeting abilities of FITC-CSPLNTRFC peptide FITC-BCSP1 optical molecular probe on breast cancer cells Bcap-37. Methods Probe FITC-BCSP1 and negative control probe FITC-svBCSP1 were prepared by solid phase synthesis. MTT assay was used to determine the toxicity of the two probes on breast cancer cells Bcap-37. The specificity of the binding of FITC-BCSP1 probe to Bcap-37 cells was identified by flow cytometry and fluorescent inversed microscopy. The specificity and targeting abilities of FITC-BCSP1 probe for transplantation tumor in Bcap-37 cells tumor-bearing nude mice model were tested by optical molecular imager. Results The purity of the synthesized probe was more than 98%, identified by mass spectrometry and high performance liquid chromatography. FITC-BCSP1 and FITC-svBCSP1 probes had no effect on proliferation and activity of Bcap-37 cells at the concentrations of 50-300 mol/L (IR%≤30%). FCM results showed that the percentage of FITC-BCSP1-labeled cells in Bcap-37 cells was significantly higher than that in other cells (all P < 0.001), and the percentage of FITC-BCSP1-labeled Bcap-37 cells was significantly higher than that of the control group (P < 0.001). It was observed under inverted fluorescence microscope that there were a large number of fluorescent cells in FITC-BCSP1-labeled Bcap-37 cells, with a positive rate of 100%, while the positive rate of FITC-svBCSP1 group was only 1%. In vivo assay with Bcap-37 cells tumor-bearing nude mice model showed that FITC-BCSP1 probe could be specifically enriched in the transplantation tumor tissue. Conclusion The optical molecular probe FITC-BCSP1 has good specificity and targeting abilities on breast cancer cells and can be used in the early diagnosis of breast cancer.

4.
Acta Pharmaceutica Sinica B ; (6): 529-545, 2020.
Article Dans Anglais | WPRIM | ID: wpr-792991

Résumé

The limited penetration of nanoparticles and their poor accessibility to cancer cell fractions in tumor remain essential challenges for effective anticancer therapy. Herein, we designed a targeting peptide-decorated biomimetic lipoprotein (termed as BL-RD) to enable their deep penetration and efficient accessibility to cancer cell fractions in a tumor, thereby improving the combinational chemo-photodynamic therapy of triple negative breast cancer. BL-RD was composed of phospholipids, apolipoprotein A1 mimetic peptide (PK22), targeting peptide-conjugated cytotoxic mertansine (RM) and photodynamic agents of DiIC18(5) (DiD). The counterpart biomimetic lipoprotein system without RM (termed as BL-D) was fabricated as control. Both BL-D and BL-RD were nanometer-sized particles with a mean diameter of less than 30 nm and could be efficiently internalized by cancer cells. After intravenous injection, they can be specifically accumulated at tumor sites. When comparing to the counterpart BL-D, BL-RD displayed superior capability to permeate across the tumor mass, extravasate from tumor vasculature to distant regions and efficiently access the cancer cell fractions in a solid tumor, thus producing noticeable depression of the tumor growth. Taken together, BL-RD can be a promising delivery nanoplatform with prominent tumor-penetrating and cancer cells-accessing capability for effective tumor therapy.

5.
Protein & Cell ; (12): 787-807, 2019.
Article Dans Anglais | WPRIM | ID: wpr-757871

Résumé

Recently, phage display technology has been announced as the recipient of Nobel Prize in Chemistry 2018. Phage display technique allows high affinity target-binding peptides to be selected from a complex mixture pool of billions of displayed peptides on phage in a combinatorial library and could be further enriched through the biopanning process; proving to be a powerful technique in the screening of peptide with high affinity and selectivity. In this review, we will first discuss the modifications in phage display techniques used to isolate various cancer-specific ligands by in situ, in vitro, in vivo, and ex vivo screening methods. We will then discuss prominent examples of solid tumor targeting-peptides; namely peptide targeting tumor vasculature, tumor microenvironment (TME) and over-expressed receptors on cancer cells identified through phage display screening. We will also discuss the current challenges and future outlook for targeting peptide-based therapeutics in the clinics.

6.
Chinese Pharmacological Bulletin ; (12): 204-210, 2016.
Article Dans Chinois | WPRIM | ID: wpr-492007

Résumé

Aim To study the growth inhibitory effect of the conjugate ( ovarian cancer specific targeting peptide and cisplatin, OSTP-DDP ) that targeting ovarian cancer cells A2780. Methods Using chemical method to syn-thesize OSTP-DDP, ovarian cancer cells A2780 were cul-tured in vitro, using CCK-8 method ( Cell Counting Kit-8) to detect the growth inhibitory effect of ovarian cancer A2780 cells, which were disposed by OSTP-DDP and DDP. Annexin V-FITC was used to detect the cycle and apoptosis effect of ovarian cancer A2780 cells which were disposed by OSTP-DDP and DDP. Results According to the mass spectrometry and the high performance liquid chromatography ( HPLC ) analysis, OSTP-DDP was proved to synthesize successfully. CCK-8 assay showed that both OSTP-DDP and DDP could play the growth in-hibitory effect and showed a concentration-dependent manner when cells were treated in different concentrations (10,20,40,80,160,320μmol·L-1 ) respectively after 24 h, 48 h, 72 h. And the effect of OSTP-DDP was stronger than DDP (P<0. 05), indicated OSTP-DDP had targeted cytostatic effect. The result of the flow cytometry showed that cell cycle was mostly arrested in G1 phase after 72h treated by OSTP-DDP and DDP, the inhibitory effect of OSTP-DDP was stronger than DDP (P<0. 05). The apop-tosis effect of OSTP-DDP was stronger than DDP ( P <0. 01),suggested that OSTP-DDP had a stronger targeting apoptosis-inducing effect. Conclusion OSTP-DDP has the targeting growth inhibitory effect on the ovarian cancer cell A2780, OSTP as a chemotherapeutic drug targeting vector has a great prospect to treat ovarian cancer.

7.
Chinese Journal of Biotechnology ; (12): 1715-1726, 2016.
Article Dans Chinois | WPRIM | ID: wpr-243686

Résumé

To obtain sufficient purified and active fusion protein-hepatocyte-targeting peptide-human endostatin (HTP-rES), we studied the growth curve and the optimal induction timing of BL21/pET21b-HTP-rES. Different conditions of pH value, induction time, induction concentration and induction temperature were optimized by univariate analysis. After washing, refolding and purifying, the activity of fusion protein was identified by flow cytometry and 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT). Results show that the logarithmic growth phase of BL21/pET21b-HTP-rES was from 1.5 h to 3.5 h, the optimum expression conditions were pH 8.0, 0.06 mmol/L IPTG, at 42 ℃ for 5 h. The purity of inclusion bodies was up to 60% after washing. The purity of target protein was more than 95% after refolding and purification. Our findings provide the foundation for further biological activity and drug development.


Sujets)
Humains , Systèmes de délivrance de médicaments , Endostatines , Pharmacologie , Escherichia coli , Hépatocytes , Corps d'inclusion , Peptides , Pharmacologie , Protéines de fusion recombinantes
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