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Construction method of double patient-derived tumor xenograft model

A technology of xenotransplantation and construction method, applied in the fields of biochemical equipment and methods, cell dissociation methods, microorganisms, etc., can solve the problems of inapplicable clinical, biological and immunological incompatibility with the human microenvironment, and achieve uniform growth size , the effect of fast growth

Active Publication Date: 2020-12-08
南京普恩瑞生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Immunotherapy has shown high anti-tumor activity and improved survival rate in tumor treatment, but durable immune response only occurs in a small number of patients, and immunotherapy requires the support of a functioning human immune system
The gene-modified immunodeficiency mouse PDTX model is the first-line auxiliary evaluation tool for clinical efficacy, but because its biology and immunology do not conform to the human microenvironment, it has significant immunodeficiency, so it is not suitable in the context of current immunotherapy. Applicable to clinical practice; in addition, in the existing humanized mouse models, most of the CD34+hsc cells and samples inoculated into mice are cultured and expanded in vitro

Method used

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  • Construction method of double patient-derived tumor xenograft model
  • Construction method of double patient-derived tumor xenograft model
  • Construction method of double patient-derived tumor xenograft model

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Such as Figure 1-Figure 12 As shown, a method for constructing a dual humanized tumor xenograft model, including:

[0063] S1: Obtain monocytes:

[0064] Fresh umbilical cord blood should be delivered to the laboratory within 24 hours after blood collection, and then the blood should be diluted 1:3 with phosphate buffer, and mononuclear cells should be separated by density gradient centrifugation on Ficoll medium;

[0065] S2: Prepare 5*10 5 cells / mL of CD34+HSC cell fluid:

[0066] The mononuclear cells were separated by a CD34+ microbead kit to obtain a CD34+HSC cell liquid, and the CD34+HSC cell liquid was adjusted to a concentration of 5*10 5 cells / mL, then the 5*10 5 The cells / mL CD34+HSC cell fluid should be stored at 2-6°C until injection;

[0067] S3: Construction of NSG mouse immune model:

[0068] Get 3 weeks old and do myeloablative NSG mouse, inject described 5*10 through tail vein 5 cells / mL of CD34+HSC cell liquid, and then from the 4th week after ...

Embodiment 2

[0073] Such as Figure 1-Figure 12 As shown, a method for constructing a dual humanized tumor xenograft model, including:

[0074] S1: Obtain monocytes:

[0075] taking fresh umbilical cord blood, and isolating mononuclear cells from the fresh umbilical cord blood;

[0076] S2: Prepare 5*10 5 cells / mL of CD34+HSC cell fluid:

[0077] The mononuclear cells were separated by a CD34+ microbead kit to obtain a CD34+HSC cell liquid, and the CD34+HSC cell liquid was adjusted to a concentration of 5*10 5 cells / mL, then the 5*10 5 The cells / mL CD34+HSC cell fluid should be stored at 4°C until injection;

[0078] S3: Construction of NSG mouse immune model:

[0079] The 3-week-old NSG mice were irradiated with X-rays with a power of 100cGy / min for 2.4min, and the 5*10 mice were injected through the tail vein. 5 cells / mL of CD34+HSC cell liquid, and then detect immunological indicators at 4, 6, 8, 10, and 12 weeks after tail vein injection, and detect the hCD45 in the blood of NSG...

Embodiment 3

[0084] Such as Figure 1-Figure 12 As shown, a method for constructing a dual humanized tumor xenograft model, including:

[0085] S1: Obtain monocytes:

[0086] taking fresh umbilical cord blood, and isolating mononuclear cells from the fresh umbilical cord blood;

[0087] S2: Prepare 5*10 5 cells / mL of CD34+HSC cell fluid:

[0088] The mononuclear cells were separated by a CD34+ microbead kit to obtain a CD34+HSC cell liquid, and the CD34+HSC cell liquid was adjusted to a concentration of 5*10 5 cells / mL, then the 5*10 5 The cells / mL CD34+HSC cell fluid should be stored at 4°C until injection;

[0089] S3: Construction of NSG mouse immune model:

[0090] The 3-week-old NSG mice were irradiated with X-rays with a power of 100cGy / min for 2.4min, and then the NSG mice were treated with myeloablation for 6 hours and then injected with the 5*10 5 cells / mL of CD34+HSC cell fluid, and then detect the immunological indicators at 4, 6, 8, 10, and 12 weeks after tail vein injec...

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PUM

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Abstract

The invention provides a construction method of a double patient-derived tumor xenograft model, and belongs to the technical field of biology. The construction method comprises the steps of S1, obtaining mononuclear cells, S2, preparing CD34+HSC cell sap of 5 * 10<5> cells / mL, S3, constructing an NSG mouse immune model, and S4, obtaining the double patient-derived tumor xenograft model. Accordingto the construction method of the double patient-derived tumor xenograft model, hCD34+ umbilical cord blood hematopoietic stem cells are adopted as an immune reconstruction blood source, the GvHD reaction is small, and the animal survival time is long; and for the standard for screening PDTX samples, the method has the advantages of being high in growth speed and uniform in growth size, and the characteristics of being basically consistent with clinical tumor histological characteristics are achieved.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for constructing a dual humanized tumor xenograft model. Background technique [0002] Cancer, also known as malignant tumor, is a worldwide prevention and control problem, and the incidence of malignant tumors in my country has been increasing year by year in the past two decades. At present, the main treatment methods for tumors include surgery, systemic chemotherapy, radiotherapy, molecular targeted therapy and Immunotherapy, among them, tumor immunotherapy is a new treatment method that can improve the survival of tumor patients after surgery, radiotherapy, chemotherapy and molecular targeted therapy. Normal anti-tumor immune response, thereby controlling and eradicating tumor therapy. [0003] Since the FDA approved the first immune checkpoint inhibitor ipilimumab in the treatment of melanoma in 2011, immune agents have been clinically used in non-small cell ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01K67/027C12N5/0789C12N5/0786
CPCA01K67/0271C12N5/0647C12N5/0645A01K2227/105A01K2267/0387C12N2509/00
Inventor 朱燕萍
Owner 南京普恩瑞生物科技有限公司
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