The patient-derived tumor xenograft (PDX) model is a transplanted tumor model constructed by transplanting fresh tumor tissue from patients into immunodeficient mice. This model simulates the growth of tumors in patients and can accurately reflect the biological characteristics of human tumors. It provides a superior in vivo model for tumor biological research, the search for diagnostic markers and drug screening, and has important translational significance for the preclinical evaluation, treatment and prognosis of tumors, especially for the personalized diagnosis and treatment of tumors.
1. Experimental principle
Fresh tumor tissue from patients is transplanted into immunodeficient mice, and the first generation of transplanted tumors is grown in the microenvironment provided by the mice. When it grows to a certain size, the transplanted tumor tissue is removed and transplanted into a new batch of mice under sterile conditions to form the second generation of transplanted tumors. Repeating this operation forms the third and fourth generations of transplanted tumors. This model can not only simulate the human microenvironment of the primary tumor, well retain the molecular biology and histopathological characteristics of the patient's primary tumor, but also better simulate the distant metastasis of the patient's primary tissue. PDX models can better preserve the heterogeneity of primary tumors than traditional transplant models, and can accurately reflect the drug responsiveness of tumors and explore the mechanism of drug resistance.
2. Experimental steps
2.1 Collection of fresh tumor tissue
Fresh tumor tissue specimens obtained from the operating room were collected immediately after ex vivo, with a size of about 0.5-1 cm3, and divided into two equal parts, one of which was kept in serum-free culture medium (containing penicillin-streptomycin) in an ice bath, and the other was quickly frozen for pathological examination. When the frozen pathological results indicate a malignant tumor, the xenotransplantation experiment will continue.
2.2 Tissue transportation
The fresh tissue specimens kept in serum-free culture medium (containing penicillin-streptomycin) in an ice bath were quickly transported to the clean bench in the animal room.
2.3 Xenotransplantation of tumor tissue
Immune-deficient mice aged 4-6 weeks were selected, and the tissue blocks were first rinsed three times in culture medium, and necrotic tissue components were removed in the cell culture dish. Three different methods of tissue transplantation are as follows:
A (homogenate injection method): After cutting the tissue into small pieces, place it in an EP tube containing 300 μL PBS, and repeatedly cut it with ophthalmic scissors until a tissue homogenate is formed. Then use a 1mL syringe to extract about 100-150 μL of the homogenate and inject it directly into the subcutaneous part of the right hypochondrium of the mouse.
B (whole piece embedding method): Cut the specimen into small pieces with a diameter of about 3 mm. After the experimental mouse is anesthetized, disinfect the skin of the right hypochondrium of the mouse, make an incision of about 3 mm in length, use separation forceps to free the subcutaneous tissue to form a small pouch, and then directly embed two small pieces of tumor tissue into the subcutaneous part of the mouse, and finally suture the skin incision with absorbable sutures.
C (puncture needle method): First cut the specimen into small pieces with a diameter of about 2 mm. Pre-fill an appropriate amount of tissue small pieces into the bone puncture needle cannula. After the experimental mouse is anesthetized, disinfect the skin of the right hypochondrium of the mouse, puncture the right hypochondrium of the mouse with an 8# needle, and expand a tunnel along the subcutaneous part. Then, insert the prepared tumor tissue into the subcutaneous tissue of the right chest and back with a bone puncture needle along the tunnel. Withdraw the puncture needle and disinfect the needle inlet. Complete the tumor transplantation operation.
2.4 Tumor observation after transplantation
After tumor transplantation, regularly observe the mouse status and tumor growth, measure the tumor size with a vernier caliper 1-2 times/week, and record the tumor volume and mouse weight change curve.
2.5 PDX passage
The successfully transplanted tumor continues to be passaged in immunodeficient mice for 2 times. The successfully transplanted tumor continues to be passaged in immunodeficient mice for 2 times. The mouse model of the third generation and later can be used for experiments.
3. Example of experimental results
Cervical cancer tissues obtained from surgical operations were transplanted into nude mice. After 6 weeks of successful establishment of the PDX model, drug treatment was given, the tumor size was measured, and tumor metastasis was observed through dissection.
4. References
[1] Ji, X. , Chen, S. , Guo, Y. , Li, W. , Qi, X. , & Yang, H. , et al. (2017). Establishment and evaluation of four different types of patient-derived xenograft models. Cancer Cell International, 17(1), 122.
[2] Yukihiko, H. , Ali, M. , Zhang, Y. , Nan, Z. , Takashi, M. , & Takashi, C. , et al. (2016). Patient-derived mouse models of cancer need to be orthotopic in order to evaluate targeted anti-metastatic therapy. Oncotarget, 7(44), 71696-71702.
[3] Yang, H. , Tong, Z. , Shen, L. , Sun, Y. U. , & Huang, J. . (2020). Brucea javanica increases survival and enhances gemcitabine efficacy in a patient-derived orthotopic xenograft (PDOX) mouse model of pancreatic cancer. Anticancer Research, 40(9), 4969-4978.