Anal. Chem. Cover article | new targeted polypeptide probe accurately recognizes and targets to penetrate HER2 positive cells
Original English Title: synergetic tumor probes for facilitating theoretical delivery by combined functional peptide talents
Corresponding author: Wang Weizhi, Beijing University of Technology
First author: Xiangqian Jia and Mingmei Guo
Wang Weizhi's team of researchers from Beijing University of science and technology used the new HER2 targeted peptide and membrane penetrating peptide to carry out functional assembly of liposomes, constructed a kind of collaborative nano probe and realized in vivo transport and targeted diagnosis and treatment. In this work, the new probe is not only a "radar" to identify and anchor HER2 cancer cells, but also a "bullet" to penetrate and strike accurately. The work was recently published as a cover article in the journal Analytical Chemistry.
Molecular recognition is a special and specific interaction between molecules formed by noncovalent bonds. Compared with antigen antibody recognition, small molecule recognition units have unique advantages in targeting peptides, especially peptides with specific affinity for specific biological targets. Compared with the antibody recognition, the targeted peptide has strong penetrability, low immunogenicity, easy chemical synthesis and modification, and high batch repeatability; compared with the nucleic acid recognition, the peptide has a higher diversity and a greater amount of information due to the more kinds of amino acids, the different hydrophobicity and charge. However, the target polypeptide faces the challenge of commonness with other small molecule probes, which is mainly manifested in: 1) it does not have stable spatial structure and target complementary, resulting in the lack of quality improvement of affinity; 2) it is easy to produce false positive when combined with the target unit price, and the specificity is not enough; 3) it is easy to be degraded and cleared by enzyme in vivo, and the biological stability is relatively low.
Recently, Wang Weizhi's team from Beijing University of science and technology reported that using polypeptides with different functions to couple with long-chain polymers, a synergistic liposome nanoprobe was constructed by self-assembly. By rational design and preparation, the probe has the advantages of orderly structure, controllable conformation, multivalent recognition and stable properties.
Figure 1. The design of polypeptide functionalized collaborative liposome probe
Firstly, a novel peptide targeting for HER2 was screened by high throughput peptide platform of microfluidic chip. HER2 (human environmental growth factor receptor-2) is a tyrosine kinase receptor, which is one of the most important molecular targets of breast cancer. HER2 is the only receptor in the family member that has not been found to be highly affinity and force specific ligands. Based on the microchannel array chip, 108 libraries of combinatorial chemical peptide library and matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS) in situ sequencing were realized. The whole process can be completed in a few hours, and the efficiency is significantly improved compared with the screening cycle of the conventional method in a few days. Based on the high efficiency screening, further development of high content screening, while obtaining more parameters. The peptides in the microcavity array are transferred to the surface plasmon resonance imaging (spri) chip in the form of in-situ "printing" to realize real-time, online and label free detection. Two groups of arrays were analyzed qualitatively (in situ sequencing) and quantitatively (in situ affinity characterization) at the same time. The affinity dissociation constant between HP2 and HER2 is 3.9 × 10? 8mol / L, which shows a good ligand / receptor interaction.
Figure 2. Cell level recognition and delivery of novel targeted polypeptides and their functionalized liposomes
Then, the cell level was verified. On the one hand, the specific affinity of HP2 to HER2 positive tumor cells was investigated. SKBR3, a human breast cancer cell line with high expression of HER2, was selected as a positive cell model to measure the affinity of living cells. FITC labeled HP2 was incubated with SKBR3 and 293T cells respectively. From Figure 2, it can be seen that HP2 binds to the cell membrane of SKBR3, showing a strong fluorescence signal. However, HP2 hardly binds to 293T cells. At the same time, HP2 is no longer combined after the HER2 expression in SKBR3 is interfered. The co localization experiment with antibody also proved that the binding site of HP2 is HER2. Based on the good affinity and specificity of HP2, we used it to connect with polymer to construct liposome probe. At the same time of HP2 functionalization, another functional peptide, the membrane penetrating peptide tat, was introduced to make the liposome probe have the function of multivalent recognition and deep penetration. By precisely controlling the distance between ligands, HP2 and Tat can play their respective roles. To construct a multi-functional vector for target recognition and deliver the drug (adriamycin shown in the red channel in the figure) to the cell.
Figure 3. Collaborative peptide functional probe to achieve diagnosis and treatment effect
Figure 4. RNA SEQ analysis of related homologous pathways by collaborative liposomes
Finally, after delivering at the cellular level, we have also developed imaging, therapy and gene analysis at the living level. In imaging, the collaborative liposome probe can more efficiently bring signal molecules into the focus to achieve accurate diagnosis; in vivo treatment, the collaborative liposome can bring more drugs to the target site and maintain good efficacy to achieve accurate treatment; and through gene analysis, we found that the targeted liposome has more tumor signal pathways related to HER2 The deep and further influence reveals the internal mechanism of targeted liposomes to achieve diagnosis and treatment.
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Anal. Chem. 2020, 92, 8, 5650-5655
Publication Date: February 28, 2020
Copyright ? 2020 American Chemical Society
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