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Activatable photoacoustic-fluorescent dual-mode probe for real-time monitoring of immunotherapy and application of activatable photoacoustic-fluorescent dual-mode probe

A technology for real-time monitoring and immunotherapy, applied in the field of immunomedicine research, which can solve the problems of lack of sensitivity and specificity of early response assessment, heterogeneity of immune response differences, and optimization of unfavorable treatment plans.

Pending Publication Date: 2022-03-01
GUANGDONG GENERAL HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, accurate monitoring of its efficacy remains challenging due to differences in heterogeneous immune responses
Traditional imaging techniques, such as PET / CT and MRI, lack the sensitivity and specificity of early response assessment, which greatly affects the efficacy and prognosis assessment in the early stage of treatment, which is not conducive to the optimization of clinical treatment plans

Method used

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  • Activatable photoacoustic-fluorescent dual-mode probe for real-time monitoring of immunotherapy and application of activatable photoacoustic-fluorescent dual-mode probe
  • Activatable photoacoustic-fluorescent dual-mode probe for real-time monitoring of immunotherapy and application of activatable photoacoustic-fluorescent dual-mode probe
  • Activatable photoacoustic-fluorescent dual-mode probe for real-time monitoring of immunotherapy and application of activatable photoacoustic-fluorescent dual-mode probe

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1 Construction of activatable probes

[0029] The required granzyme B substrate peptide was synthesized by conventional solid-phase synthesis with the following structure: Gly-Lys(Dabcyl)-Ile-Glu-Pro-Asp-Ala-Pro-Cys(FITC), and analyzed by high performance liquid chromatography The successfully labeled peptides were purified using the method for future use.

[0030] Dissolve 1 mg of the above peptide in 1 ml of ultrapure water, add 0.5 mg of NHS and 0.5 mg of EDC and incubate for 30 minutes to activate the carboxyl group in the peptide. Mix the activated peptide with 1 mg of PDL1 antibody evenly, and then magnetically stir at 200 rpm for 12 hours at room temperature, and reflect in the dark during the whole process. After the reaction is complete, use a 1KD dialysis bag to remove impurities such as free polypeptides by dialysis, and recover the antibody.

[0031] Described probe synthesis HPLC report sees attached figure 1 As shown, it can be seen that the ...

Embodiment 2

[0032] Schematic diagram of the working principle of the probe described in embodiment 2

[0033] The activatable photoacoustic-fluorescence dual-mode probe for real-time monitoring of immunotherapy according to the present invention is composed of a PDL1 antibody and a granzyme B substrate peptide, and the surface of the granzyme B substrate peptide is modified with a fluorescent group and A quenching group, the probe is an activatable photoacoustic / fluorescence dual-mode probe based on the PDL1 antibody, and is used for monitoring the activity of granzyme B.

[0034] Such as figure 2 The schematic diagram shown, after intravenous injection of the composite probe, the PDL1 antibody actively targets the tumor surface PDL1 protein (eg figure 2 In a), the anti-tumor immunity of T cells is activated by blocking the interaction between PDL1 and PD1 on the surface of T cells, and the activated T cells release granzyme B to kill tumors (such as figure 2 middle b); at the same t...

Embodiment 3

[0035] Example 3 Probe Functional Activity Characterization

[0036] Incubate the probe-labeled antibody with different concentrations (0-1mg / ml) of recombinant granzyme B for 10 minutes, and detect the fluorescence intensity at 460nm excitation and 480-600nm range by a microplate reader to verify the different probe pairs Signal response characteristics of different concentrations of granzyme B.

[0037] Such as image 3 The changes in fluorescence intensity after incubation with different granzyme concentrations and the probe for the same time indicated, it can be seen that the probe can sensitively detect granzyme B activity and is positively correlated with granzyme B concentration.

[0038] Continue to co-incubate the probe-labeled antibody with 100ng / ml recombinant granzyme B, and detect the signal response characteristics of the probe at different incubation times (0-130min) by a microplate reader.

[0039] Such as Figure 4 The change of the fluorescence intensity a...

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Abstract

The invention relates to the technical field of immunomedical research, in particular to an activatable photoacoustic-fluorescent dual-mode probe for real-time monitoring of immunotherapy, and further discloses a preparation method and application of the activatable photoacoustic-fluorescent dual-mode probe. The activatable photoacoustic-fluorescent dual-mode probe for real-time monitoring of immunotherapy is composed of a PDL1 antibody and granzyme B substrate peptide, the surface of the granzyme B substrate peptide is modified with a fluorescent group and a quenching group, and the probe is an activatable photoacoustic / fluorescent dual-mode probe based on the PDL1 antibody and is used for monitoring the activity of granzyme B.

Description

technical field [0001] The invention relates to the technical field of immunomedicine research, in particular to an activatable photoacoustic-fluorescence dual-mode probe for real-time monitoring of immunotherapy, and further discloses its preparation method and application. Background technique [0002] Tumor immunotherapy is to stimulate and enhance the immune function of the body, and enhance the anti-tumor immunity of the tumor microenvironment, so as to achieve the purpose of controlling and killing tumor cells. However, because immunotherapy can only remove a small amount of disseminated tumor cells, it has limited efficacy for advanced solid tumors, so it is often used as an adjuvant therapy in combination with conventional methods such as surgery, chemotherapy, and radiotherapy. Usually, conventional methods are used to remove a large number of tumor cells, and then immunotherapy is used to remove the remaining tumor cells, which can improve the effect of comprehensi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/37G01N21/64A61K49/00
CPCC12Q1/37G01N21/6428A61K49/0045A61K49/0058G01N2333/96436
Inventor 聂立铭樊志金梁长虹黄国家李澜庆
Owner GUANGDONG GENERAL HOSPITAL
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