Patient derived xenograft
Patient derived xenografts (PDX) are models of cancer where the tissue or cells from a patient's tumor are implanted into an immunodeficient or humanized mouse. PDX models are used to create an environment that allows for the natural growth of cancer, its monitoring, and corresponding treatment evaluations for the original patient.
Methods of tumor xenotransplantation
Several types of immunodeficient mice can be used to establish PDX models: athymic nude mice, severely compromised immune deficient (SCID) mice, NOD-SCID mice, and recombination-activating gene 2 (Rag2)-knockout mice. The mice used must be immunocompromised to prevent transplant rejection. The NOD-SCID mouse is considered more immunodeficient than the nude mouse, and therefore is more commonly used for PDX models because the NOD-SCID mouse does not produce natural killer cells.
When human tumors are resected, necrotic tissues are removed and the tumor can be mechanically sectioned into smaller fragments, chemically digested, or physically manipulated into a single-cell suspension. There are advantages and disadvantages in utilizing either discrete tumor fragments or single-cell suspensions. Tumor fragments retain cell-cell interactions as well as some tissue architecture of the original tumor, therefore mimicking the tumor microenvironment. Alternatively, a single-cell suspension enables scientists to collect an unbiased sampling of the whole tumor, eliminating spatially segregate subclones that are otherwise inadvertently selected during analysis or tumor passaging. However, single-cell suspensions subject surviving cells to harsh chemical or mechanical forces that may sensitize cells to anoikis, taking a toll on cell viability and engraftment success.
Heterotopic and orthotopic implantation
Unlike creating xenograft mouse models using existing cancer cell lines, there are no intermediate in vitro processing steps before implanting tumor fragments into a murine host to create a PDX. The tumor fragments are either implanted heterotopically or orthotopically into an immunodeficient mouse. With heterotopical implantantion, the tissue or cells are implanted into an area of the mouse unrelated to the original tumor site, generally subcutaneously or in subrenal capsular sites. The advantages of this method are the direct access for implantation, and ease of monitoring the tumor growth. With orthotopic implantation, scientists transplant the patient’s tumor tissue or cells into the corresponding anatomical position in the mouse. Subcutaneous PDX models rarely produce metastasis in mice, nor do they simulate the initial tumor microenvironment, with engraftment rates of 40-60%. Subrenal capsular PDX maintains the original tumor stroma as well as the equivalent host stroma and has an engraftment rate of 95%. Ultimately, it takes about 2 to 4 months for the tumor to engraft varying by tumor type, implant location, and strain of immunodeficient mice utilized; engraftment failure should not be declared until at least 6 months. Researchers may use heterotopic implantation for the initial engraftment from the patient to the mouse, then use orthotopic implantation to implant the mouse-grown tumor into further generations of mice.
Generations of engraftments
The first generation of mice receiving the patient's tumor fragments are commonly denoted F0. When the tumor-burden becomes too large for the F0 mouse, researchers passage the tumor over to the next generation of mice. Each generation thereafter is denoted F1, F2, F3…Fn. For drug development studies, expansion of mice after the F3 generation is often utilized after ensuring that the PDX has not genetically or histologically diverged from the patient’s tumor.
Advantages over established cancer cell lines
Cancer cell lines are originally derived from patient tumors, but acquire the ability to proliferate within in vitro cell cultures. As a result of in vitro manipulation, cell lines that have been traditionally used in cancer research undergo genetic transformations that are not restored when cells are allowed to grow in vivo. Because of the cell culturing process, which includes enzymatic environments and centrifugation, cells that are better adapted to survive in culture are selected, tumor resident cells and proteins that interact with cancer cells are eliminated, and the culture becomes phenotypically homogeneous.
When implanted into immunodeficient mice, cell lines do not easily develop tumors and the result of any successfully grown tumor is a genetically divergent tumor unlike the heterogeneous patient tumor. Researchers are beginning to attribute the reason that only 5% of anti-cancer agents are approved by the Food and Drug Administration after pre-clinical testing to the lack of tumor heterogeneity and the absence of the human stromal microenvironment. Specifically, cell line-xenografts often are not predictive of the drug response in the primary tumors because cell lines do not follow pathways of drug resistance or the effects of the microenvironment on drug response found in human primary tumors.
Many PDX models have been successfully established for breast, prostate, colorectal, lung, and many other cancers because there are distinctive advantages when using PDX over cell lines for drug safety and efficacy studies as well as predicting patient tumor response to certain anti-cancer agents. Since PDX can be passaged without in vitro processing steps, PDX models allow the propagation and expansion of patient tumors without significant genetic transformation of tumor cells over multiple murine generations. Within PDX models, patient tumor samples grow in physiologically-relevant tumor microenvironments that mimic the oxygen, nutrient, and hormone levels that are found in the patient’s primary tumor site. Furthermore, implanted tumor tissue maintains the genetic and epigenetic abnormalities found in the patient and the xenograft tissue can be excised from the patient to include the surrounding human stroma. As a result, numerous studies have found that PDX models exhibit similar responses to anti-cancer agents as seen in the actual patient who provided the tumor sample.
Humanized xenograft models
One prominent shortcoming of PDX models is that immunodeficient mice must be used to prevent immune attacks against the xenotransplanted tumor. With the immune system incapacitated, a critical component of the known tumor microenvironment interaction is foregone, preventing immunotherapies and anti-cancer agents that target the immune system components from being studied in PDX models. Researchers are beginning to explore the use of humanized-xenograft models to enable immune studies. Humanized-xenograft models are created by co-engrafting the patient tumor fragment and peripheral blood or bone marrow cells into a NOD/SCID mouse. The co-engraftment allows for reconstitution of the murine immune system, giving insight into the interactions between xenogenic human stroma and tumor environments in cancer progression and metastasis. However, these strategies have yet to be validated for most tumor types and there remain questions over whether the reconstituted immune system will behave in the same way as it does in the patient. For example, the immune system could be 'hyper-activated' due to exposure to mouse tissues in a similar fashion to graft versus host disease. Humanized-xenograft models for acute lymphoblastic leukemia and acute myeloid leukemia have been created.
The classification of genetic breast cancer subtypes, including triple-negative and HER2-positive subtypes, have allowed oncologists to use a patient’s breast cancer subtype to personalize cancer therapy schedules. Utilizing PDX triple negative breast cancer models, scientists found that aurora kinase inhibitors slows tumor growth rate and suppresses recurrence in a breast cancer subtype that has a high recurrence rate and poor survivability. Scientists have also found that breast cancer PDX models are capable of predicting the prognosis of newly diagnosed women by observing the rate of tumor engraftment to determine if the patient tumor is aggressive.
Colorectal PDX models are relatively easy to establish and the models maintain genetic similarity of primary patient tumor for about 14 generations. In 2012, a study established 27 colorectal PDX models that did not diverge from their respective human tumors in histology, gene expression, or KRAS/BRAF mutation status. Due to their stability, the 27 colorectal PDX models may be able to serve as pre-clinical models in future drug studies. Drug resistance studies have been conducted using colorectal PDX models. In one study, researchers found that the models predicted patient responsiveness to cetuximab with 90% accuracy. Another study identified the amplification of ERBB2 as another mechanism of resistance, and a putative new actionable target in treatments.
Researchers initially focused on using pancreatic PDX models for drug studies to improve the process to develop predictive and pharmacodynamics end points for several molecularly targeted therapies. Other studies have been conducted to explore if pancreatic PDX models can be used to guide the ongoing treatment of advance pancreatic cancer patient by screening multiple drugs to select the drug with most activity as the next line of treatment. Pancreatic PDX models have shown anti-mesothilin CAR-T cells (T-cells modified with a chimeric antigen receptor) to suppress cancer growth.
Pediatric cancer (neuroblastoma)
Researchers have established neuroblastoma PDXs by orthotopic implantation of patient tumor explants into immunodeficient mice. The PDXs retained the genotype and phenotype of patient tumors, and exhibited substantial infiltrative growth and metastasis to distant organs including the bone marrow. The researchers cultured PDX-derived neuroblastoma cells in vitro and the cells retained tumorigenic and metastatic capacity in vivo.
Challenges with PDX model adaptation
There are several challenges that scientists face when developing or using PDX models in research. For instance, not all tumor samples will successfully engraft in an immunodeficient mouse. When engraftment does occur, clinical study protocols are difficult to standardize if engraftment rates vary. It is also expensive to house mice, maintain histopathological cores for frequent testing, and perform ex vivo passaging of tumors in mice with high tumor burdens. The scientific community is trying to tackle these challenges mutualising efforts and exchanging models and expertise to avoid duplication: academic networks have sprung in Europe and USA, and the Horizon 2020 program is funding a new Research Infrastructure providing standardised services and resources, with the goal of improving reproducibility and open access to resources and services.
With regard to using PDX in personalized medicine, there are financial challenges. In the US, the cost to develop PDX models is can potentially cost a patient thousands of dollars for treatment. PDX models can also take significant time to create, which may pose a challenge to patients with advanced stages of cancer. Despite these setbacks, the PDX market is expected to grow from a market capitalization of $77.4 million in 2017 to $167.6 million in 2022 due to growing demand for personalized medicine.
- Lai, Yunxin; Wei, Xinru; Lin, Shouheng; Qin, Le; Cheng, Lin; Li, Peng (12 May 2017). "Current status and perspectives of patient-derived xenograft models in cancer research". Journal of Hematology & Oncology. 10 (1): 106. doi:10.1186/s13045-017-0470-7. ISSN 1756-8722. PMC 5427553. PMID 28499452.
- Morton CL, Houghton PJ (2007). "Establishment of human tumor xenografts in immunodeficient mice". Nature Protocols (Protocol). 2 (2): 247–50. doi:10.1038/nprot.2007.25. PMID 17406581.
- Siolas D, Hannon GJ (September 2013). "Patient-derived tumor xenografts: transforming clinical samples into mouse models". Cancer Research (Perspective). 73 (17): 5315–9. doi:10.1158/0008-5472.CAN-13-1069. PMC 3766500. PMID 23733750.
- Williams SA, Anderson WC, Santaguida MT, Dylla SJ (September 2013). "Patient-derived xenografts, the cancer stem cell paradigm, and cancer pathobiology in the 21st century". Laboratory Investigation. 93 (9): 970–82. doi:10.1038/labinvest.2013.92. PMID 23917877.
- Zvibel I, Smets F, Soriano H (2002). "Anoikis: roadblock to cell transplantation?". Cell Transplantation. 11 (7): 621–30. doi:10.3727/000000002783985404. PMID 12518889. S2CID 27366723.
- Jin K, Teng L, Shen Y, He K, Xu Z, Li G (July 2010). "Patient-derived human tumour tissue xenografts in immunodeficient mice: a systematic review". Clinical & Translational Oncology (Review). 12 (7): 473–80. doi:10.1007/s12094-010-0540-6. PMID 20615824. S2CID 38277136.
- Cutz JC, Guan J, Bayani J, et al. (July 2006). "Establishment in severe combined immunodeficiency mice of subrenal capsule xenografts and transplantable tumor lines from a variety of primary human lung cancers: potential models for studying tumor progression-related changes". Clinical Cancer Research. 12 (13): 4043–54. doi:10.1158/1078-0432.CCR-06-0252. PMID 16818704.
- Tentler JJ, Tan AC, Weekes CD, et al. (June 2012). "Patient-derived tumour xenografts as models for oncology drug development". Nature Reviews. Clinical Oncology (Review). 9 (6): 338–50. doi:10.1038/nrclinonc.2012.61. PMC 3928688. PMID 22508028. Note: open access via PMC; closed via publisher site.
- Daniel VC, Marchionni L, Hierman JS, et al. (April 2009). "A primary xenograft model of small-cell lung cancer reveals irreversible changes in gene expression imposed by culture in vitro". Cancer Research. 69 (8): 3364–73. doi:10.1158/0008-5472.CAN-08-4210. PMC 2821899. PMID 19351829.
- Hutchinson L, Kirk R (April 2011). "High drug attrition rates--where are we going wrong?". Nature Reviews. Clinical Oncology (Editorial). 8 (4): 189–90. doi:10.1038/nrclinonc.2011.34. PMID 21448176.
- Malaney P, Nicosia SV, Davé V (March 2014). "One mouse, one patient paradigm: New avatars of personalized cancer therapy". Cancer Letters (Mini-review). 344 (1): 1–12. doi:10.1016/j.canlet.2013.10.010. PMC 4092874. PMID 24157811.
- Reyal F, Guyader C, Decraene C, et al. (2012). "Molecular profiling of patient-derived breast cancer xenografts". Breast Cancer Research. 14 (1): R11. doi:10.1186/bcr3095. PMC 3496128. PMID 22247967.
- Richmond A, Su Y (2008). "Mouse xenograft models vs GEM models for human cancer therapeutics". Disease Models & Mechanisms (Editorial). 1 (2–3): 78–82. doi:10.1242/dmm.000976. PMC 2562196. PMID 19048064.
- Kerbel RS (2003). "Human tumor xenografts as predictive preclinical models for anticancer drug activity in humans: better than commonly perceived-but they can be improved". Cancer Biology & Therapy (Review). 2 (4 Suppl 1): S134–9. doi:10.4161/cbt.213. PMID 14508091.
- Talmadge JE, Singh RK, Fidler IJ, Raz A (March 2007). "Murine models to evaluate novel and conventional therapeutic strategies for cancer". The American Journal of Pathology (Review). 170 (3): 793–804. doi:10.2353/ajpath.2007.060929. PMC 1864878. PMID 17322365.
- Cassidy JW, Caldas C, Bruna A (July 2015). "Maintaining tumour heterogeneity in patient derived tumour xenografts". Cancer Research (Review). 75 (15): 2963–2968. doi:10.1158/0008-5472.CAN-15-0727. PMC 4539570. PMID 26180079.
- Meyer LH, Debatin KM (December 2011). "Diversity of human leukemia xenograft mouse models: implications for disease biology". Cancer Research (Review). 71 (23): 7141–4. doi:10.1158/0008-5472.CAN-11-1732. PMID 22088964.
- Romanelli A, Clark A, Assayag F, et al. (December 2012). "Inhibiting aurora kinases reduces tumor growth and suppresses tumor recurrence after chemotherapy in patient-derived triple-negative breast cancer xenografts". Molecular Cancer Therapeutics. 11 (12): 2693–703. doi:10.1158/1535-7163.MCT-12-0441-T. PMID 23012245.
- DeRose YS, Wang G, Lin YC, et al. (2011). "Tumor grafts derived from women with breast cancer authentically reflect tumor pathology, growth, metastasis and disease outcomes". Nature Medicine. 17 (11): 1514–20. doi:10.1038/nm.2454. PMC 3553601. PMID 22019887.
- Guenot D, Guérin E, Aguillon-Romain S, et al. (April 2006). "Primary tumour genetic alterations and intra-tumoral heterogeneity are maintained in xenografts of human colon cancers showing chromosome instability". The Journal of Pathology. 208 (5): 643–52. doi:10.1002/path.1936. PMID 16450341. S2CID 30504094.
- Uronis JM, Osada T, McCall S, et al. (2012). "Histological and molecular evaluation of patient-derived colorectal cancer explants". PLOS ONE. 7 (6): e38422. doi:10.1371/journal.pone.0038422. PMC 3366969. PMID 22675560.
- Krumbach R, Schüler J, Hofmann M, Giesemann T, Fiebig HH, Beckers T (May 2011). "Primary resistance to cetuximab in a panel of patient-derived tumour xenograft models: activation of MET as one mechanism for drug resistance". European Journal of Cancer. 47 (8): 1231–43. doi:10.1016/j.ejca.2010.12.019. PMID 21273060.
- Bertotti A, Migliardi G, Galimi F, et al. (November 2011). "A molecularly annotated platform of patient-derived xenografts ('xenopatients') identifies HER2 as an effective therapeutic target in cetuximab-resistant colorectal cancer". Cancer Discovery. 1 (6): 508–23. doi:10.1158/2159-8290.CD-11-0109. PMID 22586653.
- Hidalgo M, Bruckheimer E, Rajeshkumar NV, et al. (August 2011). "A pilot clinical study of treatment guided by personalized tumorgrafts in patients with advanced cancer". Molecular Cancer Therapeutics. 10 (8): 1311–6. doi:10.1158/1535-7163.MCT-11-0233. PMC 4629061. PMID 21673092.
- Laheru D, Shah P, Rajeshkumar NV, et al. (December 2012). "Integrated preclinical and clinical development of S-trans, trans-Farnesylthiosalicylic Acid (FTS, Salirasib) in pancreatic cancer". Investigational New Drugs. 30 (6): 2391–9. doi:10.1007/s10637-012-9818-6. PMC 3557459. PMID 22547163.
- Jiang, Hua; et al. (8 December 2017). "Efficient growth suppression in pancreatic cancer PDX model by fully human anti-mesothelin CAR-T cells". Protein & Cell. 8 (12): 926–931. doi:10.1007/s13238-017-0472-9. PMC 5712292. PMID 28929447.
- Braekeveldt N, Wigerup C, Gisselsson D, et al. (March 2015). "Neuroblastoma patient-derived orthotopic xenografts retain metastatic patterns and geno- and phenotypes of patient tumours". International Journal of Cancer. 136 (5): E252–61. doi:10.1002/ijc.29217. PMC 4299502. PMID 25220031.
- "EurOPDX - European Oncology PDX network". Retrieved 9 February 2021.
- "PDXNet - PDX Development and Trial Centers Research Network". Retrieved 9 February 2021.
- "Research Infrastructures - European Commission". Retrieved 9 February 2021.
- "EDIReX - the EurOPDX Distributed Infrastructure for Research on patient-derived cancer Xenografts". Retrieved 9 February 2021.
- Pollack A (September 25, 2012). "Seeking cures, patients enlist mice stand-ins". Business Day. The New York Times.
- "Patient Derived Xenograft/PDX Models Market - Global Forecast to 2022 - Growing Demand for Personalized Medicine - Research and Markets". Business Wire. 2017-12-29. Retrieved 11 June 2018.