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Targeting nanoprobe combined with NK92 cells as well as preparation method and application of targeting nanoprobe

A nanoprobe, NK cell technology, applied in the field of medical imaging and nanomedicine, to achieve the effect of being conducive to long-term monitoring and mechanism research, improving targeting, and enhancing lethality

Pending Publication Date: 2020-11-27
SHANGHAI FIRST PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, there is no Fc binding to NK92 cells as in the present application 3 o 4 -PEG-CD56 / Avastin@Ce6 nanoprobe

Method used

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  • Targeting nanoprobe combined with NK92 cells as well as preparation method and application of targeting nanoprobe
  • Targeting nanoprobe combined with NK92 cells as well as preparation method and application of targeting nanoprobe
  • Targeting nanoprobe combined with NK92 cells as well as preparation method and application of targeting nanoprobe

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1 Synthesis and characterization of Fe3O4-PEG-CD56 / Avastin@Ce6 nanoprobe; binding to NK-92 cells and its effect on NK-92 cells

[0039] 1 Materials and methods

[0040] 1.1 Materials

[0041] 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC, Aladdin), N-hydroxysuccinimide (NHS, Aladdin), paramethoxy Carboxylated polyethylene glycol (mPEG-COOH, Shanghai Yaer Technology Co., Ltd.), sodium borate buffer (SBB), α-MEM medium (Gibco company), fetal bovine serum (FBS, Gibco company), horse serum ( Gibco Company), recombinant human IL-2 (rhIL-2, PeproTech Company), bevacizumab injection (Avastin, Roche Company), anti-CD56 antibody (BD Company), penicillin-streptomycin solution and trypsin ( Xinsaimei Biotechnology Co., Ltd.), UV-visible spectrophotometer (PerkinElmer, USA), JEOL 2010F transmission electron microscope (TEM, JEOL Ltd.), flow cytometer (BD Company).

[0042] 1.2 Synthesis of targeted NPs

[0043] (1) Dissolve 2mg EDC and 2mg NHS in 0.64...

Embodiment 2

[0070] Example 2 incorporates Fe 3 o 4 -The killing effect of PEG-CD56 / Avastin@Ce6 nanoprobe on NK-92 cells against breast cancer cells

[0071] 1 Experimental method

[0072] Take breast cancer cells (4T1 cells and MCF-7 cells) growing in the logarithmic phase, and adjust the cell concentration to 5×10 5 / mL, added to a six-well plate for culture, after the cells adhered to the wall, the old medium was discarded, and the pre-treated NK-92 cells combined with Fe 3 o 4 - NK-92 cells (NK+nNPs) of PEG-CD56@Ce6 (20 μg / mL Fe 3 o 4 -PEG-CD56@Ce6, co-cultivated for 24h), combined with Fe 3 o 4 - NK-92 cells (NK+NPs) of PEG-CD56 / Avastin@Ce6 (20 μg / mL Fe 3 o 4 -PEG-CD56 / Avastin@Ce6, co-cultivated for 24 hours) Add the transwell chamber at 10 times the density of breast cancer cells, put the transwell chamber into a six-well plate, and continue to culture for 24 hours. After the co-cultivation, collect the supernatant in the six-well plate, The adherent breast cancer cells wer...

Embodiment 3

[0075] Example 3 NK+NPs therapeutic effect in vivo and MRI and fluorescence dual-modal imaging

[0076] 1 Experimental method

[0077] 1.1 Establishment of nude mouse mammary gland model

[0078] (1) Feeding of nude mice: 5-week-old BALB / c nude mice with a body weight of 15-22 grams were raised in an SPF animal room, the indoor temperature was kept at 25-28°C, and the relative humidity was controlled at 40%-60 %, the air is filtered through a high-efficiency air filter. 4-5 nude mice were raised in each cage, and the litter, feed and water used were all autoclaved, replaced every three days, cleaned on time, and the environment was kept sterile and dry.

[0079] (2) Transfer the 4T1 cells in the logarithmic growth phase and in a good growth state to a centrifuge tube after trypsinization, centrifuge at 1000 rpm for 3 minutes, discard the supernatant, and wash twice with pre-cooled sterile PBS to ensure that no After residual serum and trypsin, resuspend with PBS, count the ...

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Abstract

The invention relates to a targeting nanoprobe combined with NK92 cells as well as a preparation method and application of the targeting nanoprobe. According to the invention, the Fe3O4-PEG-CD56 / Avastin-@ Ce6 nanoprobe targeting NK92 cells is synthesized, an anti-CD56 antibody targeting NK cells and an antitumor drug bevacizumab are covalently connected to the surfaces of superparamagnetic Fe3O4 nanoparticles, and then a photosensitizer Ce6 is physically adsorbed to the surfaces of the nanoparticles. The probe can be used for fluorescence and magnetic resonance bimodal imaging, is good in biocompatibility and stability, can be used for long-time monitoring of living bodies, and shows excellent anti-tumor effect and biological safety in vivo and in vitro. The invention lays a foundation forresearch and immunotherapy of the action mechanism of the NK cells.

Description

technical field [0001] The invention relates to the fields of medical imaging and nanomedicine, in particular to a targeted nanoprobe combined with NK92 cells and its preparation method and application. Background technique [0002] Natural killer (natural kill, NK) cells have been paid more and more attention as a treatment mode of tumor immunotherapy. Usually, the effect of immunotherapy is evaluated by indicators such as the reduction of tumor markers, the reduction of tumor volume, and the improvement of survival rate, which often require invasive biopsy pathology and are time-consuming. Molecular imaging probes can perform real-time non-invasive imaging of tumor occurrence and development at the molecular level, and can also guide and monitor targeted therapy, thereby realizing the integration of tumor diagnosis and treatment. Therefore, the application of molecular imaging technology to label immune cells with nanotechnology has led to the development of many immune c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K47/60A61K47/68A61K49/00A61K49/12A61K49/16A61K39/395A61P35/00B82Y5/00
CPCA61K49/0002A61K49/0036A61K49/0058A61K49/0054A61K49/16A61K49/126A61K47/6849A61K47/60C07K16/22A61P35/00B82Y5/00A61K2039/505A61K2039/585
Inventor 李康安连婧阁高国段猛
Owner SHANGHAI FIRST PEOPLES HOSPITAL
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