Patient derived xenograft: Difference between revisions
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==Methods of tumor xenotransplantation== |
==Methods of tumor xenotransplantation== |
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Several types of immunodeficient mice can be used to establish PDTX models: athymic [[Nude mouse|nude mice]], [[SCID mouse#Mice|severe combined immune deficient (SCID) mice]], [[NOD mice|nonobese diabetic (NOD)-SCID mice]], and recombination-activating gene 2 (Rag2)-knockout mice.<ref>Morton CL, Houghton PJ |
Several types of immunodeficient mice can be used to establish PDTX models: athymic [[Nude mouse|nude mice]], [[SCID mouse#Mice|severe combined immune deficient (SCID) mice]], [[NOD mice|nonobese diabetic (NOD)-SCID mice]], and recombination-activating gene 2 (Rag2)-knockout mice.<ref>{{cite journal |author=Morton CL, Houghton PJ |title=Establishment of human tumor xenografts in immunodeficient mice |journal=Nature Protocols |volume=2 |issue=2 |pages=247–50 |year=2007 |pmid=17406581 |doi=10.1038/nprot.2007.25}}</ref> The mice used must be immunocompromised to prevent transplant rejection or other transplant complications such as graft-versus-host-disease. The NOD-SCID mouse is considered more immunodeficient than the nude mouse, and therefore is more commonly used for PDTX models because the NOD-SCID mouse does not produce Natural Killer cells.<ref name="Siolas">{{cite journal |author=Siolas D, Hannon GJ |title=Patient-derived tumor xenografts: transforming clinical samples into mouse models |journal=Cancer Research |volume=73 |issue=17 |pages=5315–9 |year=2013 |month=September |pmid=23733750 |pmc=3766500 |doi=10.1158/0008-5472.CAN-13-1069}}</ref> |
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When human tumors are resected, necrotic tissues is removed and the tumor is either mechanically sectioned into smaller fragments or chemically digested or physically manipulated into a single-cell suspension. There are both 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, 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<ref>Williams |
When human tumors are resected, necrotic tissues is removed and the tumor is either mechanically sectioned into smaller fragments or chemically digested or physically manipulated into a single-cell suspension. There are both 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, 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<ref>{{cite journal |author=Williams SA, Anderson WC, Santaguida MT, Dylla SJ |title=Patient-derived xenografts, the cancer stem cell paradigm, and cancer pathobiology in the 21st century |journal=Laboratory Investigation |volume=93 |issue=9 |pages=970–82 |year=2013 |month=September |pmid=23917877 |doi=10.1038/labinvest.2013.92}}</ref> 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.<ref>{{cite journal |author=Zvibel I, Smets F, Soriano H |title=Anoikis: roadblock to cell transplantation? |journal=Cell Transplantation |volume=11 |issue=7 |pages=621–30 |year=2002 |pmid=12518889 |doi=10.3727/000000002783985404}}</ref> |
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Unlike creating xenograft mouse models using existing cancer cell lines, there are no intermediate ''in vitro'' processing steps before implanting tumor fragments murine host to create a PDTX. The tumor fragments are either be implanted heterotopically or orthotopically of an immunodeficient mouse. Heterotopical implants occur when the tumor fragment is implanted into an area of the mouse unrelated to the original tumor site, generally subcutaneously or subrenal capsular sites.<ref name="Jin K 2010">Jin K, Teng L, Shen Y, He K, Xu Z, Li G |
Unlike creating xenograft mouse models using existing cancer cell lines, there are no intermediate ''in vitro'' processing steps before implanting tumor fragments murine host to create a PDTX. The tumor fragments are either be implanted heterotopically or orthotopically of an immunodeficient mouse. Heterotopical implants occur when the tumor fragment is implanted into an area of the mouse unrelated to the original tumor site, generally subcutaneously or subrenal capsular sites.<ref name="Jin K 2010">{{cite journal |author=Jin K, Teng L, Shen Y, He K, Xu Z, Li G |title=Patient-derived human tumour tissue xenografts in immunodeficient mice: a systematic review |journal=Clinical & Translational Oncology |volume=12 |issue=7 |pages=473–80 |year=2010 |month=July |pmid=20615824 |doi=10.1007/s12094-010-0540-6}}</ref> Whereas, scientists tranplant the patient’s tumor tissue and implants the fragments into the corresponding anatomical position in the mouse in an orthotopic transplant. Subcutaneous PDTX rarely produce metastasis in mice, does not simulate the initial tumor microenvironment, and has engraftment rates 40-60%.<ref name="Jin K 2010" /> Subrenal capsular PDTX maintains the original tumor stroma as well as the equivalent host stroma and has an engraftment rate of 95%.<ref>{{cite journal |author=Cutz JC, Guan J, Bayani J, ''et al.'' |title=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 |journal=Clinical Cancer Research |volume=12 |issue=13 |pages=4043–54 |year=2006 |month=July |pmid=16818704 |doi=10.1158/1078-0432.CCR-06-0252}}</ref> Ultimately, the time 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.<ref>{{cite journal |author=Morton CL, Houghton PJ |title=Establishment of human tumor xenografts in immunodeficient mice |journal=Nature Protocols |volume=2 |issue=2 |pages=247–50 |year=2007 |pmid=17406581 |doi=10.1038/nprot.2007.25}}</ref> |
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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 PDTX has not genetically or histologically diverged from the patient’s tumor.<ref name="Tentler 2012">Tentler JJ, Tan AC, Weekes CD, |
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 PDTX has not genetically or histologically diverged from the patient’s tumor.<ref name="Tentler 2012">{{cite journal |author=Tentler JJ, Tan AC, Weekes CD, ''et al.'' |title=Patient-derived tumour xenografts as models for oncology drug development |journal=Nature Reviews. Clinical Oncology |volume=9 |issue=6 |pages=338–50 |year=2012 |month=June |pmid=22508028 |pmc=3928688 |doi=10.1038/nrclinonc.2012.61}}</ref> |
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==Advantages over cancer cell lines== |
==Advantages over cancer cell lines== |
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[[Immortalised cell line|Cancer cell lines (CCL)]] are originally derived from patient tumors, but acquire the ability to proliferate within ''in vitro'' [[cell culture]]s. As a result of ''in vitro'' manipulation, CCL that have been traditionally used in cancer research undergo genetic transformations that are not restored when cells are allowed to grow ''in vivo''.<ref>Daniel VC, Marchionni L, Hierman JS, |
[[Immortalised cell line|Cancer cell lines (CCL)]] are originally derived from patient tumors, but acquire the ability to proliferate within ''in vitro'' [[cell culture]]s. As a result of ''in vitro'' manipulation, CCL that have been traditionally used in cancer research undergo genetic transformations that are not restored when cells are allowed to grow ''in vivo''.<ref>{{cite journal |author=Daniel VC, Marchionni L, Hierman JS, ''et al.'' |title=A primary xenograft model of small-cell lung cancer reveals irreversible changes in gene expression imposed by culture in vitro |journal=Cancer Research |volume=69 |issue=8 |pages=3364–73 |year=2009 |month=April |pmid=19351829 |pmc=2821899 |doi=10.1158/0008-5472.CAN-08-4210}}</ref> 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.<ref name="William 2013">{{cite journal |author=Williams SA, Anderson WC, Santaguida MT, Dylla SJ |title=Patient-derived xenografts, the cancer stem cell paradigm, and cancer pathobiology in the 21st century |journal=Laboratory Investigation |volume=93 |issue=9 |pages=970–82 |year=2013 |month=September |pmid=23917877 |doi=10.1038/labinvest.2013.92}}</ref> |
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When implanted into immunodeficient mice, CCL do not easily develop tumors and the result of any successfully grown tumor is a genetically divergent tumor unlike the heterogeneous patient tumor.<ref name="William 2013" /> 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.<ref name="Hutchinson">Hutchinson L, Kirk R |
When implanted into immunodeficient mice, CCL do not easily develop tumors and the result of any successfully grown tumor is a genetically divergent tumor unlike the heterogeneous patient tumor.<ref name="William 2013" /> 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.<ref name="Hutchinson">{{cite journal |author=Hutchinson L, Kirk R |title=High drug attrition rates--where are we going wrong? |journal=Nature Reviews. Clinical Oncology |volume=8 |issue=4 |pages=189–90 |year=2011 |month=April |pmid=21448176 |doi=10.1038/nrclinonc.2011.34}}</ref> Specifically, CCL-xenografts often are not predictive of the drug response in the primary tumors because CCL do not follow pathways of drug resistance or the effects of the microenvironment on drug response found in human primary tumors.<ref name="Hutchinson" /> |
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Many PDTX models have been successfully established for breast, prostate, colorectal, lung, and many other cancers because there are distinctive advantages when using PDTX over CCL for drug safety and efficacy studies as well as predicting patient tumor response to certain anti-cancer agents.<ref name="one mouse">Malaney P, Nicosia SV, Davé V |
Many PDTX models have been successfully established for breast, prostate, colorectal, lung, and many other cancers because there are distinctive advantages when using PDTX over CCL for drug safety and efficacy studies as well as predicting patient tumor response to certain anti-cancer agents.<ref name="one mouse">{{cite journal |author=Malaney P, Nicosia SV, Davé V |title=One mouse, one patient paradigm: New avatars of personalized cancer therapy |journal=Cancer Letters |volume=344 |issue=1 |pages=1–12 |year=2014 |month=March |pmid=24157811 |doi=10.1016/j.canlet.2013.10.010}}</ref> Since PDTX can be passaged without ''in vitro'' processing steps, PDTX models allow the propagation and expansion of patient tumors without significant genetic transformation of tumor cells over multiple murine generations.<ref>{{cite journal |author=Reyal F, Guyader C, Decraene C, ''et al.'' |title=Molecular profiling of patient-derived breast cancer xenografts |journal=Breast Cancer Research |volume=14 |issue=1 |pages=R11 |year=2012 |pmid=22247967 |pmc=3496128 |doi=10.1186/bcr3095}}</ref> Within PDTX models, patient tumor samples grow in physiological relevant tumor microenvironments that mimic the oxygen, nutrient, and hormone levels that are found in the patient’s primary tumor site.<ref name="Tentler 2012" /> 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.<ref name="xeno v gem">{{cite journal |author=Richmond A, Su Y |title=Mouse xenograft models vs GEM models for human cancer therapeutics |journal=Disease Models & Mechanisms |volume=1 |issue=2-3 |pages=78–82 |year=2008 |pmid=19048064 |pmc=2562196 |doi=10.1242/dmm.000976}}</ref> As a result, numerous studies have found that PDTX models exhibit similar responses to anti-cancer agents as seen in the actual patient who provided the tumor sample.<ref>{{cite journal |author=Kerbel RS |title=Human tumor xenografts as predictive preclinical models for anticancer drug activity in humans: better than commonly perceived-but they can be improved |journal=Cancer Biology & Therapy |volume=2 |issue=4 Suppl 1 |pages=S134–9 |year=2003 |pmid=14508091 |url=http://www.landesbioscience.com/journals/cbt/abstract.php?id=213}}</ref> PDTX models are beneficial to use to study therapeutic responses to drugs because multiple therapies can be tested against one biopsy and pre- and post-treatment data can be acquired from the human biopsy and xenograft tissues, potentially sparing a patient from therapies that may not work.<ref name="xeno v gem" /> |
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===Humanized-xenograft models=== |
===Humanized-xenograft models=== |
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One prominent shortcoming of PDTX models is that immunodeficient mice must be used to prevent immune attacks against the xenotransplanted tumor. Therefore, a critical component of the known tumor microenvironment interaction is foregone. As a result, immunotherapies and anti-cancer agents that target the immune system components cannot be studied using PDTX models. Consequently, researchers are beginning to explore the use of humanized-xenograft models. Humanized-xenograft models are created by co-engrafting the patient tumor fragment and peripheral blood or bone marrow cells into a NOD/SCID mouse.<ref name="Siolas" /> The co-engraftment allows for reconstitution of the murine immune system enabling researchers to study the interactions between xenogenic human stroma and tumor environments in cancer progression and metastasis.<ref>Talmadge JE, Singh RK, Fidler IJ, Raz A |
One prominent shortcoming of PDTX models is that immunodeficient mice must be used to prevent immune attacks against the xenotransplanted tumor. Therefore, a critical component of the known tumor microenvironment interaction is foregone. As a result, immunotherapies and anti-cancer agents that target the immune system components cannot be studied using PDTX models. Consequently, researchers are beginning to explore the use of humanized-xenograft models. Humanized-xenograft models are created by co-engrafting the patient tumor fragment and peripheral blood or bone marrow cells into a NOD/SCID mouse.<ref name="Siolas" /> The co-engraftment allows for reconstitution of the murine immune system enabling researchers to study the interactions between xenogenic human stroma and tumor environments in cancer progression and metastasis.<ref>{{cite journal |author=Talmadge JE, Singh RK, Fidler IJ, Raz A |title=Murine models to evaluate novel and conventional therapeutic strategies for cancer |journal=The American Journal of Pathology |volume=170 |issue=3 |pages=793–804 |year=2007 |month=March |pmid=17322365 |pmc=1864878 |doi=10.2353/ajpath.2007.060929}}</ref> Humanized-xenograft models for acute lymphoblastic leukemia and acute myeloid leukemia have been created.<ref>{{cite journal |author=Meyer LH, Debatin KM |title=Diversity of human leukemia xenograft mouse models: implications for disease biology |journal=Cancer Research |volume=71 |issue=23 |pages=7141–4 |year=2011 |month=December |pmid=22088964 |doi=10.1158/0008-5472.CAN-11-1732}}</ref> |
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==Clinical Relevance== |
==Clinical Relevance== |
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===Breast Cancer=== |
===Breast Cancer=== |
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There have been many advances in breast cancer biology resulting in the classification of different molecular and [[Breast cancer classification#DNA classification|genetic breast cancer subtypes]] including triple-negative and [[HER2/neu|HER2-positive]] subtypes.<ref name="Tentler 2012" /> Oncologist can use a patient’s breast cancer subtype to personalize cancer therapy schedules to better address the patient’s tumor distinct gene-expression profile. Utilizing PDTX [[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.<ref>Romanelli A, Clark A, Assayag F, |
There have been many advances in breast cancer biology resulting in the classification of different molecular and [[Breast cancer classification#DNA classification|genetic breast cancer subtypes]] including triple-negative and [[HER2/neu|HER2-positive]] subtypes.<ref name="Tentler 2012" /> Oncologist can use a patient’s breast cancer subtype to personalize cancer therapy schedules to better address the patient’s tumor distinct gene-expression profile. Utilizing PDTX [[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.<ref>{{cite journal |author=Romanelli A, Clark A, Assayag F, ''et al.'' |title=Inhibiting aurora kinases reduces tumor growth and suppresses tumor recurrence after chemotherapy in patient-derived triple-negative breast cancer xenografts |journal=Molecular Cancer Therapeutics |volume=11 |issue=12 |pages=2693–703 |year=2012 |month=December |pmid=23012245 |doi=10.1158/1535-7163.MCT-12-0441-T}}</ref> Scientist have also found that breast cancer PDTX 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.<ref>{{cite journal |author=DeRose YS, Wang G, Lin YC, ''et al.'' |title=Tumor grafts derived from women with breast cancer authentically reflect tumor pathology, growth, metastasis and disease outcomes |journal=Nature Medicine |volume=17 |issue=11 |pages=1514–20 |year=2011 |pmid=22019887 |pmc=3553601 |doi=10.1038/nm.2454}}</ref> |
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===Colorectal Cancer=== |
===Colorectal Cancer=== |
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Colorectal PDTX models are relatively easy to establish and the models maintain genetic similarity of primary patient tumor for about 14 generations.<ref>Guenot D, et al. |
Colorectal PDTX models are relatively easy to establish and the models maintain genetic similarity of primary patient tumor for about 14 generations.<ref>{{cite journal |author=Guenot D, Guérin E, Aguillon-Romain S, ''et al.'' |title=Primary tumour genetic alterations and intra-tumoral heterogeneity are maintained in xenografts of human colon cancers showing chromosome instability |journal=The Journal of Pathology |volume=208 |issue=5 |pages=643–52 |year=2006 |month=April |pmid=16450341 |doi=10.1002/path.1936}}</ref> In 2012, a study established 27 colorectal PDTX models that did not diverge from their respective human tumors in histology, gene expression, or KRAS/BRAF mutation status.<ref>{{cite journal |author=Uronis JM, Osada T, McCall S, ''et al.'' |title=Histological and molecular evaluation of patient-derived colorectal cancer explants |journal=Plos One |volume=7 |issue=6 |pages=e38422 |year=2012 |pmid=22675560 |pmc=3366969 |doi=10.1371/journal.pone.0038422}}</ref> Due to their stability, the 27 colorectal PDTX models may be able to serve as pre-clinical models in future drug studies. Drug resistance studies have been conducted using colorectal PDTX models. In one study, researchers found that the models predicted patient responsiveness to [[cetuximab]] with 90% accuracy.<ref>{{cite journal |author=Krumbach R, Schüler J, Hofmann M, Giesemann T, Fiebig HH, Beckers T |title=Primary resistance to cetuximab in a panel of patient-derived tumour xenograft models: activation of MET as one mechanism for drug resistance |journal=European Journal of Cancer |volume=47 |issue=8 |pages=1231–43 |year=2011 |month=May |pmid=21273060 |doi=10.1016/j.ejca.2010.12.019}}</ref> Another study identified the amplification of [[ERBB2]] as another mechanism of resistance, and a putative new actionable target in treatments.<ref>{{cite journal |author=Bertotti A, Migliardi G, Galimi F, ''et al.'' |title=A molecularly annotated platform of patient-derived xenografts ('xenopatients') identifies HER2 as an effective therapeutic target in cetuximab-resistant colorectal cancer |journal=Cancer Discovery |volume=1 |issue=6 |pages=508–23 |year=2011 |month=November |pmid=22586653 |doi=10.1158/2159-8290.CD-11-0109}}</ref> |
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''PLoS ONE'' 7:e38422. [http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0038422/ article] {{DOI|10.1371/journal.pone.0038422}}</ref> Due to their stability, the 27 colorectal PDTX models may be able to serve as pre-clinical models in future drug studies. Drug resistance studies have been conducted using colorectal PDTX models. In one study, researchers found that the models predicted patient responsiveness to [[cetuximab]] with 90% accuracy.<ref>R. Krumbach, J. Schuler, M. Hofmann, T. Giesemann, H.H. Fiebig, T. Beckers, 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 (2011) 1231–1243. [http://www.ncbi.nlm.nih.gov/pubmed/21273060/ article] {{DOI|10.1016/j.ejca.2010.12.019}}</ref> Another study identified the amplification of [[ERBB2]] as another mechanism of resistance, and a putative new actionable target in treatments.<ref>Andrea Bertotti, Giorgia Migliardi, Francesco Galimi, Francesco Sassi, Davide Torti, Claudio Isella, Davide Corà, Federica Di Nicolantonio, Michela Buscarino, Consalvo Petti, Dario Ribero, Nadia Russolillo, Andrea Muratore, Paolo Massucco, Alberto Pisacane, Luca Molinaro, Emanuele Valtorta, Andrea Sartore-Bianchi, Mauro Risio, Lorenzo Capussotti, Marcello Gambacorta, Salvatore Siena, Enzo Medico, Anna Sapino, Silvia Marsoni, Paolo M. Comoglio, Alberto Bardelli, and Livio Trusolino, A molecularly annotated platform of patient-derived xenografts ("xenopatients") identifies HER2 as an effective therapeutic target in cetuximab-resistant colorectal cancer.[http://www.ncbi.nlm.nih.gov/pubmed/22586653/ article] {{DOI| 10.1158/2159-8290.CD-11-0109}}</ref> |
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===Pancreatic Cancer=== |
===Pancreatic Cancer=== |
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Researchers initially focused on using pancreatic PDTX models for drug studies to improve the process to develop predictive and pharmacodynamics end points for several molecularly targeted therapies.<ref name="Tentler 2012" /> Other studies have been conducted to explore if pancreatic PDTX 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.<ref>Hidalgo M, et al. |
Researchers initially focused on using pancreatic PDTX models for drug studies to improve the process to develop predictive and pharmacodynamics end points for several molecularly targeted therapies.<ref name="Tentler 2012" /> Other studies have been conducted to explore if pancreatic PDTX 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.<ref>{{cite journal |author=Hidalgo M, Bruckheimer E, Rajeshkumar NV, ''et al.'' |title=A pilot clinical study of treatment guided by personalized tumorgrafts in patients with advanced cancer |journal=Molecular Cancer Therapeutics |volume=10 |issue=8 |pages=1311–6 |year=2011 |month=August |pmid=21673092 |doi=10.1158/1535-7163.MCT-11-0233}}</ref> Pancreatic PDTX models have enabled researchers to simultaneously run pre-clinical and [[clinical trial]] to test the effectiveness of drugs in cancer patients.<ref>{{cite journal |author=Laheru D, Shah P, Rajeshkumar NV, ''et al.'' |title=Integrated preclinical and clinical development of S-trans, trans-Farnesylthiosalicylic Acid (FTS, Salirasib) in pancreatic cancer |journal=Investigational New Drugs |volume=30 |issue=6 |pages=2391–9 |year=2012 |month=December |pmid=22547163 |pmc=3557459 |doi=10.1007/s10637-012-9818-6}}</ref> |
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===Pediatric Preclinical Testing Program=== |
===Pediatric Preclinical Testing Program=== |
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The [http://pptp.nchresearch.org/ Pediatric Preclinical Testing Program (PPTP)] is operated by St. Jude Children’s Research Hospital sponsored through a [[National Cancer Institute]] research contract. The purpose of the PPTP is to identify more effective treatments for childhood cancers through systematic evaluation of cancer drugs in childhood solid tumors and leukemia models.<ref name="PPTP">"Pediatric Preclinical Testing Program (PPTP)." Pediatric Preclinical Testing Program. Nationwide Children's Hospital, 2009. [http://pptp.nchresearch.org/index.html/ http://pptp.nchresearch.org/index.html]</ref> The PPTP is a collaborative effort between research institutions and pharmaceutical companies to test novel agents against PPTP developed pre-clinical cancer models.<ref> |
The [http://pptp.nchresearch.org/ Pediatric Preclinical Testing Program (PPTP)] is operated by St. Jude Children’s Research Hospital sponsored through a [[National Cancer Institute]] research contract. The purpose of the PPTP is to identify more effective treatments for childhood cancers through systematic evaluation of cancer drugs in childhood solid tumors and leukemia models.<ref name="PPTP">"Pediatric Preclinical Testing Program (PPTP)." Pediatric Preclinical Testing Program. Nationwide Children's Hospital, 2009. [http://pptp.nchresearch.org/index.html/ http://pptp.nchresearch.org/index.html]</ref> The PPTP is a collaborative effort between research institutions and pharmaceutical companies to test novel agents against PPTP developed pre-clinical cancer models.<ref>{{cite web |title=Pediatric Preclinical Testing Program (PPTP) |publisher=[[CTEP]] |date=August 4, 2014 |accessdate=August 31, 2014 |url=http://ctep.cancer.gov/MajorInitiatives/Pediatric_Preclinical_Testing_Program.htm}}</ref> The has program established panels of childhood cancer xenografts as well as cell lines models to be used for ''in vivo'' and ''in vitro'' testing, including models for Ewing sarcoma, Wilms tumor, neuroblastoma, glioblastoma, and acute lymphoblastic leukemia. In 2007, ''in vivo'' testing on 51 solid tumor and 10 acute lymphoblastic leukemia models established the broad-spectrum activity of vincristine and cyclophosphamide in treating childhood cancers.<ref>{{cite journal |author=Houghton PJ, Morton CL, Tucker C, ''et al.'' |title=The pediatric preclinical testing program: description of models and early testing results |journal=Pediatric Blood & Cancer |volume=49 |issue=7 |pages=928–40 |year=2007 |month=December |pmid=17066459 |doi=10.1002/pbc.21078}}</ref> As of April 2014, there has been 62 peer-review publications releasing data from the project.<ref name="PPTP" /> |
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====Principal Investigators==== |
====Principal Investigators==== |
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==Challenges with PDTX model adaptation== |
==Challenges with PDTX model adaptation== |
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There are several challenges that scientists face when developing or using PDTX models in research. For instance not all tumor samples will successfully engraft in the immunodeficient mouse.<ref name="one mouse" /> When engraftment does occur, clinical study protocols are difficult to standardize if engraftment rates vary.<ref name="one mouse" /> There are financial challenges that slow the adaption of PDTX models as the standard for drug development. The cost to develop PDTX models is not covered by insurance and can potentially cost a patient $25,500 just for doctors to have access to the technology to potentially guide the patient's treatment.<ref>Pollack |
There are several challenges that scientists face when developing or using PDTX models in research. For instance not all tumor samples will successfully engraft in the immunodeficient mouse.<ref name="one mouse" /> When engraftment does occur, clinical study protocols are difficult to standardize if engraftment rates vary.<ref name="one mouse" /> There are financial challenges that slow the adaption of PDTX models as the standard for drug development. The cost to develop PDTX models is not covered by insurance and can potentially cost a patient $25,500 just for doctors to have access to the technology to potentially guide the patient's treatment.<ref>{{cite news |last=Pollack |first=Andrew |year=2012 |url=http://www.nytimes.com/2012/09/26/business/mice-as-stand-ins-in-the-fight-against-disease.html |title=Seeking cures, patients enlist ice stand-ins |work=The New York Times |date=September 25, 2012}}</ref> Furthermore, the cost just for researchers rise steeply when considering the resources one must utilize to house mice, maintain histopatholigcal cores for frequent testing,<ref name="one mouse" /> and pay for the expertise necessary to perform ''ex vivo'' passaging of tumors in mice with high tumor burdens.<ref name="Siolas" /> Therefore, many institutions cannot afford to run PDTX labs because of limited funds. |
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==Commercial Companies and Contract Research Organizations== |
==Commercial Companies and Contract Research Organizations== |
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Revision as of 12:15, 31 August 2014
Patient derived tumor xenografts (PDTX) are created when cancerous tissue from a patient’s primary tumor is implanted directly into an immunodeficient mouse. PDTX models are providing solutions to the challenges that researchers face in cancer drug research such as positive tumor responses in mouse models but not translating over when the study is implemented in humans. As a result, PDTX cancer models are becoming popular models to use in cancer drug research.
Methods of tumor xenotransplantation
Several types of immunodeficient mice can be used to establish PDTX models: athymic nude mice, severe combined immune deficient (SCID) mice, nonobese diabetic (NOD)-SCID mice, and recombination-activating gene 2 (Rag2)-knockout mice.[1] The mice used must be immunocompromised to prevent transplant rejection or other transplant complications such as graft-versus-host-disease. The NOD-SCID mouse is considered more immunodeficient than the nude mouse, and therefore is more commonly used for PDTX models because the NOD-SCID mouse does not produce Natural Killer cells.[2]
When human tumors are resected, necrotic tissues is removed and the tumor is either mechanically sectioned into smaller fragments or chemically digested or physically manipulated into a single-cell suspension. There are both 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, 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[3] 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.[4]
Unlike creating xenograft mouse models using existing cancer cell lines, there are no intermediate in vitro processing steps before implanting tumor fragments murine host to create a PDTX. The tumor fragments are either be implanted heterotopically or orthotopically of an immunodeficient mouse. Heterotopical implants occur when the tumor fragment is implanted into an area of the mouse unrelated to the original tumor site, generally subcutaneously or subrenal capsular sites.[5] Whereas, scientists tranplant the patient’s tumor tissue and implants the fragments into the corresponding anatomical position in the mouse in an orthotopic transplant. Subcutaneous PDTX rarely produce metastasis in mice, does not simulate the initial tumor microenvironment, and has engraftment rates 40-60%.[5] Subrenal capsular PDTX maintains the original tumor stroma as well as the equivalent host stroma and has an engraftment rate of 95%.[6] Ultimately, the time 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.[7]
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 PDTX has not genetically or histologically diverged from the patient’s tumor.[8]
Advantages over cancer cell lines
Cancer cell lines (CCL) are originally derived from patient tumors, but acquire the ability to proliferate within in vitro cell cultures. As a result of in vitro manipulation, CCL that have been traditionally used in cancer research undergo genetic transformations that are not restored when cells are allowed to grow in vivo.[9] 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.[10]
When implanted into immunodeficient mice, CCL do not easily develop tumors and the result of any successfully grown tumor is a genetically divergent tumor unlike the heterogeneous patient tumor.[10] 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.[11] Specifically, CCL-xenografts often are not predictive of the drug response in the primary tumors because CCL do not follow pathways of drug resistance or the effects of the microenvironment on drug response found in human primary tumors.[11]
Many PDTX models have been successfully established for breast, prostate, colorectal, lung, and many other cancers because there are distinctive advantages when using PDTX over CCL for drug safety and efficacy studies as well as predicting patient tumor response to certain anti-cancer agents.[12] Since PDTX can be passaged without in vitro processing steps, PDTX models allow the propagation and expansion of patient tumors without significant genetic transformation of tumor cells over multiple murine generations.[13] Within PDTX models, patient tumor samples grow in physiological relevant tumor microenvironments that mimic the oxygen, nutrient, and hormone levels that are found in the patient’s primary tumor site.[8] 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.[14] As a result, numerous studies have found that PDTX models exhibit similar responses to anti-cancer agents as seen in the actual patient who provided the tumor sample.[15] PDTX models are beneficial to use to study therapeutic responses to drugs because multiple therapies can be tested against one biopsy and pre- and post-treatment data can be acquired from the human biopsy and xenograft tissues, potentially sparing a patient from therapies that may not work.[14]
Humanized-xenograft models
One prominent shortcoming of PDTX models is that immunodeficient mice must be used to prevent immune attacks against the xenotransplanted tumor. Therefore, a critical component of the known tumor microenvironment interaction is foregone. As a result, immunotherapies and anti-cancer agents that target the immune system components cannot be studied using PDTX models. Consequently, researchers are beginning to explore the use of humanized-xenograft models. Humanized-xenograft models are created by co-engrafting the patient tumor fragment and peripheral blood or bone marrow cells into a NOD/SCID mouse.[2] The co-engraftment allows for reconstitution of the murine immune system enabling researchers to study the interactions between xenogenic human stroma and tumor environments in cancer progression and metastasis.[16] Humanized-xenograft models for acute lymphoblastic leukemia and acute myeloid leukemia have been created.[17]
Clinical Relevance
Breast Cancer
There have been many advances in breast cancer biology resulting in the classification of different molecular and genetic breast cancer subtypes including triple-negative and HER2-positive subtypes.[8] Oncologist can use a patient’s breast cancer subtype to personalize cancer therapy schedules to better address the patient’s tumor distinct gene-expression profile. Utilizing PDTX 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.[18] Scientist have also found that breast cancer PDTX 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.[19]
Colorectal Cancer
Colorectal PDTX models are relatively easy to establish and the models maintain genetic similarity of primary patient tumor for about 14 generations.[20] In 2012, a study established 27 colorectal PDTX models that did not diverge from their respective human tumors in histology, gene expression, or KRAS/BRAF mutation status.[21] Due to their stability, the 27 colorectal PDTX models may be able to serve as pre-clinical models in future drug studies. Drug resistance studies have been conducted using colorectal PDTX models. In one study, researchers found that the models predicted patient responsiveness to cetuximab with 90% accuracy.[22] Another study identified the amplification of ERBB2 as another mechanism of resistance, and a putative new actionable target in treatments.[23]
Pancreatic Cancer
Researchers initially focused on using pancreatic PDTX models for drug studies to improve the process to develop predictive and pharmacodynamics end points for several molecularly targeted therapies.[8] Other studies have been conducted to explore if pancreatic PDTX 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.[24] Pancreatic PDTX models have enabled researchers to simultaneously run pre-clinical and clinical trial to test the effectiveness of drugs in cancer patients.[25]
Pediatric Preclinical Testing Program
The Pediatric Preclinical Testing Program (PPTP) is operated by St. Jude Children’s Research Hospital sponsored through a National Cancer Institute research contract. The purpose of the PPTP is to identify more effective treatments for childhood cancers through systematic evaluation of cancer drugs in childhood solid tumors and leukemia models.[26] The PPTP is a collaborative effort between research institutions and pharmaceutical companies to test novel agents against PPTP developed pre-clinical cancer models.[27] The has program established panels of childhood cancer xenografts as well as cell lines models to be used for in vivo and in vitro testing, including models for Ewing sarcoma, Wilms tumor, neuroblastoma, glioblastoma, and acute lymphoblastic leukemia. In 2007, in vivo testing on 51 solid tumor and 10 acute lymphoblastic leukemia models established the broad-spectrum activity of vincristine and cyclophosphamide in treating childhood cancers.[28] As of April 2014, there has been 62 peer-review publications releasing data from the project.[26]
Principal Investigators
| Name | Institution |
|---|---|
| Dr. Peter Houghton, PhD | The Research Institute Nationwide Children's Hospital |
| Dr. E. Anders Kolb, MD | A.I duPont Children’s Hospital Wilmington |
| Dr. Richard Gorlick, MD | Children's Hospital at Montefiore Albert Einstein College of Medicine |
| Dr. Stephen Keir, PhD | Preston Robert Tisch Brain Tumor Center Duke University |
| Dr. John Maris, MD | Children's Hospital of Philadelphia |
| Dr. Richard Lock, PhD | Children's Cancer Institute Randwick, Australia |
| Dr. C. Patrick Reynolds, MD, PhD | Texas Tech University Health Science Center Lubbock |
Challenges with PDTX model adaptation
There are several challenges that scientists face when developing or using PDTX models in research. For instance not all tumor samples will successfully engraft in the immunodeficient mouse.[12] When engraftment does occur, clinical study protocols are difficult to standardize if engraftment rates vary.[12] There are financial challenges that slow the adaption of PDTX models as the standard for drug development. The cost to develop PDTX models is not covered by insurance and can potentially cost a patient $25,500 just for doctors to have access to the technology to potentially guide the patient's treatment.[29] Furthermore, the cost just for researchers rise steeply when considering the resources one must utilize to house mice, maintain histopatholigcal cores for frequent testing,[12] and pay for the expertise necessary to perform ex vivo passaging of tumors in mice with high tumor burdens.[2] Therefore, many institutions cannot afford to run PDTX labs because of limited funds.
Commercial Companies and Contract Research Organizations
Commercial companies and other contract research organizations are beginning to emerge on the market. As more companies and organizations are formed, the competition may help to lower the costs and provide the expertise needed to develop and maintain PDTX models.
Canada
- Living Tumor Laboratory Vancouver, British Columbia, Canada
China
- Pharmaron Beijing, China
- WuXi AppTec Shanghai, China
France
- Oncodesign Dijon Cedex, France
- Urolead, Strasbourg, France
- XenTech Paris, France
Germany
- Experimental Pharmacology and Oncology Berlin-Buch, Germany
- Oncotest Freiburg, Germany
India
- Deshpande Laboratories Bhopal, Madhya Pradesh, India
United States of America
- Aveo Oncology Cambridge, Massachusetts, USA
- Champions Oncology Hackensack, New Jersey, USA
- Charles River Laboratories Wilmington, Massachusetts, USA
- Crownbio Santa Clara, California, USA
- GenScript Piscataway, New Jersey, USA
- The Jackson Laboratory Bar Harbor, Maine, USA
- Molecular Response San Diego, California, USA
- Taconic Hudson, New York, USA
References
- ^ Morton CL, Houghton PJ (2007). "Establishment of human tumor xenografts in immunodeficient mice". Nature Protocols. 2 (2): 247–50. doi:10.1038/nprot.2007.25. PMID 17406581.
- ^ a b c Siolas D, Hannon GJ (2013). "Patient-derived tumor xenografts: transforming clinical samples into mouse models". Cancer Research. 73 (17): 5315–9. doi:10.1158/0008-5472.CAN-13-1069. PMC 3766500. PMID 23733750.
{{cite journal}}: Unknown parameter|month=ignored (help) - ^ Williams SA, Anderson WC, Santaguida MT, Dylla SJ (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.
{{cite journal}}: Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Zvibel I, Smets F, Soriano H (2002). "Anoikis: roadblock to cell transplantation?". Cell Transplantation. 11 (7): 621–30. doi:10.3727/000000002783985404. PMID 12518889.
{{cite journal}}: CS1 maint: multiple names: authors list (link) - ^ a b Jin K, Teng L, Shen Y, He K, Xu Z, Li G (2010). "Patient-derived human tumour tissue xenografts in immunodeficient mice: a systematic review". Clinical & Translational Oncology. 12 (7): 473–80. doi:10.1007/s12094-010-0540-6. PMID 20615824.
{{cite journal}}: Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Cutz JC, Guan J, Bayani J; et al. (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.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Morton CL, Houghton PJ (2007). "Establishment of human tumor xenografts in immunodeficient mice". Nature Protocols. 2 (2): 247–50. doi:10.1038/nprot.2007.25. PMID 17406581.
- ^ a b c d Tentler JJ, Tan AC, Weekes CD; et al. (2012). "Patient-derived tumour xenografts as models for oncology drug development". Nature Reviews. Clinical Oncology. 9 (6): 338–50. doi:10.1038/nrclinonc.2012.61. PMC 3928688. PMID 22508028.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Daniel VC, Marchionni L, Hierman JS; et al. (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.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ a b Williams SA, Anderson WC, Santaguida MT, Dylla SJ (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.
{{cite journal}}: Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ a b Hutchinson L, Kirk R (2011). "High drug attrition rates--where are we going wrong?". Nature Reviews. Clinical Oncology. 8 (4): 189–90. doi:10.1038/nrclinonc.2011.34. PMID 21448176.
{{cite journal}}: Unknown parameter|month=ignored (help) - ^ a b c d Malaney P, Nicosia SV, Davé V (2014). "One mouse, one patient paradigm: New avatars of personalized cancer therapy". Cancer Letters. 344 (1): 1–12. doi:10.1016/j.canlet.2013.10.010. PMID 24157811.
{{cite journal}}: Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ 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.
{{cite journal}}: Explicit use of et al. in:|author=(help)CS1 maint: multiple names: authors list (link) CS1 maint: unflagged free DOI (link) - ^ a b Richmond A, Su Y (2008). "Mouse xenograft models vs GEM models for human cancer therapeutics". Disease Models & Mechanisms. 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. 2 (4 Suppl 1): S134–9. PMID 14508091.
- ^ Talmadge JE, Singh RK, Fidler IJ, Raz A (2007). "Murine models to evaluate novel and conventional therapeutic strategies for cancer". The American Journal of Pathology. 170 (3): 793–804. doi:10.2353/ajpath.2007.060929. PMC 1864878. PMID 17322365.
{{cite journal}}: Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Meyer LH, Debatin KM (2011). "Diversity of human leukemia xenograft mouse models: implications for disease biology". Cancer Research. 71 (23): 7141–4. doi:10.1158/0008-5472.CAN-11-1732. PMID 22088964.
{{cite journal}}: Unknown parameter|month=ignored (help) - ^ Romanelli A, Clark A, Assayag F; et al. (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.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ 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.
{{cite journal}}: Explicit use of et al. in:|author=(help)CS1 maint: multiple names: authors list (link) - ^ Guenot D, Guérin E, Aguillon-Romain S; et al. (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.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ 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.
{{cite journal}}: Explicit use of et al. in:|author=(help)CS1 maint: multiple names: authors list (link) CS1 maint: unflagged free DOI (link) - ^ Krumbach R, Schüler J, Hofmann M, Giesemann T, Fiebig HH, Beckers T (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.
{{cite journal}}: Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Bertotti A, Migliardi G, Galimi F; et al. (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.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Hidalgo M, Bruckheimer E, Rajeshkumar NV; et al. (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. PMID 21673092.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Laheru D, Shah P, Rajeshkumar NV; et al. (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.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ a b "Pediatric Preclinical Testing Program (PPTP)." Pediatric Preclinical Testing Program. Nationwide Children's Hospital, 2009. http://pptp.nchresearch.org/index.html
- ^ "Pediatric Preclinical Testing Program (PPTP)". CTEP. August 4, 2014. Retrieved August 31, 2014.
- ^ Houghton PJ, Morton CL, Tucker C; et al. (2007). "The pediatric preclinical testing program: description of models and early testing results". Pediatric Blood & Cancer. 49 (7): 928–40. doi:10.1002/pbc.21078. PMID 17066459.
{{cite journal}}: Explicit use of et al. in:|author=(help); Unknown parameter|month=ignored (help)CS1 maint: multiple names: authors list (link) - ^ Pollack, Andrew (September 25, 2012). "Seeking cures, patients enlist ice stand-ins". The New York Times.
{{cite news}}: CS1 maint: date and year (link)