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Genetically engineered hematopoietic stem cell drug delivery system as well as preparation method and application thereof

A hematopoietic stem cell and genetic engineering technology, applied in the field of genetically engineered hematopoietic stem cell drug delivery system and its preparation, can solve the problems of loss of cell biological activity, CFDV can't specifically target organs, and can't respond to chemical signals, etc., to achieve Reduce immunotoxicity, low immunogenicity, simple purification and culture

Pending Publication Date: 2022-07-26
SUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

CFDV has no key organelles and will lose cell-specific biological activity, so many CFDVs often cannot specifically target organs, nor can they respond to chemical signals in the body

Method used

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  • Genetically engineered hematopoietic stem cell drug delivery system as well as preparation method and application thereof
  • Genetically engineered hematopoietic stem cell drug delivery system as well as preparation method and application thereof
  • Genetically engineered hematopoietic stem cell drug delivery system as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1 Construction of a genetically engineered hematopoietic stem cell drug delivery platform (SB@HSCs-PD-1)

[0047] Configuration of TGF-β inhibitor solution: 10 mg / mL solution was prepared in DMSO, and then diluted in PBS to configure different concentration gradients.

[0048] After incubating the TGF-β inhibitor with a concentration gradient with hematopoietic stem cells for 4 hours, centrifugation at 500 g for 3 minutes, collecting the supernatant, and detecting its loading and drug release by high performance liquid chromatography. The result is as figure 1 As shown, after HSCs were incubated with TGF-β inhibitor at a concentration of 50 μg / mL, the encapsulation efficiency was 14.78 ± 2.55%, and the loading rate was 5.02 ± 0.98 wt%. 7.39±0.49 μg TGF-β inhibition. TGF-[beta] inhibition in phosphate buffer at 37[deg.]C released about 77.24±4.75% in 48 hours.

[0049] The purified HSCs were co-incubated with the lentiviral vector (pRLenti-EF1-EGFP-P2A-Puro-CM...

Embodiment 2

[0051] Example 2 Correlation characterization of SB@HSCs-PD-1

[0052] (1) Binding ability of HSCs-PD-1 to PD-L1 antibody

[0053] The melanomas with high PD-L1 expression were prepared by transfecting lentivirus (HBLV-m-CD274-3xflag-ZsGreen-PURO) to detect the PD-L1 binding ability of HSCs-PD-1. The results are as follows Figure 4 As shown, the transformed melanoma cells not only stably express ZsGreen signal, but also highly express PD-L1. Suspended HSCs-PD-1 and HSCs were separately co-cultured with adherent PD-L1-expressing melanoma cells B16F10 for 3 hours, and unbound supernatant cells were removed by washing with PBS. The remaining cells were stained with DAPI and PE-CD44 antibodies and the results were as follows Figure 5 shown, HSCs-PD-1 co-incubation showed a significant increase in CD44 signal compared to cancer cells co-cultured with HSCs, suggesting that HSCs-PD-1 can adhere to B16F10-PD-L1 via PD-1 / PD-L1 binding on cells.

[0054] (2) Bone homing ability of...

Embodiment 3

[0058] Example 3 Therapeutic effect of SB@HSCs-PD-1 on bone metastases

[0059] The mice were administered with PBS (200 μL) and HSCs (1 × 10 via tail vein) on the third day after the establishment of the bone metastases model. 5 cells), SB (7.5 μg), HSC-PD-1 (1×10 5 cells), SB@HSCs (1×10 5 cells) and SB@HSCs-PD-1 (1×10 5 cells), injected every three days for a total of 4 treatments. The size of bone metastases was measured by small animal imaging at specific time points and the survival of mice in each group was recorded.

[0060] The result is as Figure 8 As shown, in mice receiving PBS, HSC, SB alone, the bioluminescence signal of bone metastases increased rapidly, indicating that bone metastases do not respond to antibodies to PD-L1. Bone metastatic burden was slightly reduced in mice treated with SB@HSCs or HSCs-PD-1 compared with control mice, and after treatment with the genetically engineered hematopoietic cell delivery system (SB@HSCs-PD-1), bone metastases were...

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Abstract

The invention discloses a genetically engineered hematopoietic stem cell drug delivery system which is used for treating bone metastatic tumor. The immunotherapy substance is over-expressed by mediating hematopoietic stem cells, the drug TGF-beta inhibitor is loaded, the bone marrow site is targeted, and the immunotherapy substance is used for treating the bone marrow metastasis tumor. The genetically engineered hematopoietic stem cell drug delivery system disclosed by the invention has natural attributes of targeted bone marrow, low immunogenicity, good biological safety, expanded production through cell culture after gene modification, convenience in preparation and wide application prospect.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a genetically engineered hematopoietic stem cell drug delivery system and a preparation method and application thereof. Background technique [0002] Bone is one of the most common metastatic destinations in breast, prostate and lung cancers. More than two-thirds of bone metastases are not confined to the bone, but metastasize further to systemic metastases, ultimately resulting in patient death. In addition, bone metastases are often accompanied by bone destruction, which can lead to severe fracture destruction and severe bone pain, seriously affecting the quality of life of patients. The current clinical treatments for bone metastases include palliative radiotherapy, systemic chemotherapy, osteoclast blocker therapy, and adjuvant therapy with analgesics. However, all of these are palliative rather than curative treatments. Therefore, there is an urgent need to develop ne...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K38/17A61K45/06A61K47/46A61P19/08A61P35/04A61K31/4439
CPCA61K48/0058A61K48/0008A61K38/1774A61K45/06A61K31/4439A61K47/46A61P35/04A61P19/08A61K2300/00
Inventor 汪超王蓓蕾
Owner SUZHOU UNIV
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