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Mouse Models of Gynecologic Pathology

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   The   new england journal of    medicine   n engl j med 352;21   www.nejm.orgmay 26, 2005   2240  clinical implications of basic research   Mouse Models of Gynecologic Pathology  Jeff Boyd, Ph.D. Over the past two decades, the cataloguing of mu-tated oncogenes and tumor-suppressor genes in various types of human cancers has provided arich knowledge base for the creation of geneticmodels. Although powerful in terms of their abil-ity to elucidate gene function in a controlled envi-ronment in vivo, these models are prone to the per-ils of biologic reductionism. Often missed are theboilerplate goals of mimicking a particular humancancer sufficiently to render insight into the early natural history of the disease process and provid-ing a relevant preclinical tool for testing thera-pies. Ovarian carcinoma has proved exceptionally refractory to genetic modeling; only during thepast three years were the first two mouse modelsannounced. 1,2  Dinulescu et al. 3  have recently de-scribed a new model that is a tour de force in geneticmodeling of gynecologic abnormalities. They haveengineered both a model of the highly prevalent,nonmalignant condition of endometriosis and amodel of the endometrioid histologic subtype of invasive epithelial ovarian carcinoma, which some-times arises within endometriotic lesions.Mouse models of cancer are typically gener-ated by one of two strategies: the engineering of transgenic animals to express, through the germline, an activated oncogene, or the use of “knock-out” animals that carry a mutationally inactivatedtumor-suppressor gene. These relatively simplisticapproaches often lead to unpredictable or undesir-able outcomes, however. It is not unusual for eitherapproach, but especially the use of genetic knock-out animals, to result in embryonic lethality (whichis not always uninformative) or adult mice that havea cancer other than that seen in humans who carry or acquire a defect in the orthologous gene.The development of molecular techniques that allow the targeted manipulation of the mouse ge-nome in a tissue- or time-specific fashion 4  has pro- vided a welcome means to overcome these obsta-cles. For example, one such strategy was used togenerate invasive ovarian carcinomas in transgenicmice carrying the transforming oncogene fromsimian virus 40 DNA, which is under the transcrip-tional control of a promoter of a gene whose ex-pression is specific to the ovarian epithelial cell. 2 A powerful strategy for the conditional inacti- vation of tumor-suppressor genes typically involvesthe use of a recombination system composed of arecombinase enzyme and its target sequence. Al-though there are many variations of this concept,the basic approach is that the desired gene se-quence, flanked by stretches of DNA that mediaterecombination, is introduced into the germ lineand the resultant mice are crossed with mice bear-ing the Cre recombinase gene, generally under thecontrol of a tissue-specific or drug-inducible pro-moter. With the evolution of this technique, theCre   recombinase system can now be used to condi-tionally activate (gain of function) or inactivate (lossof function) gene expression in a time- or tissue-spe-cific manner and thus allow highly accurate mousemodeling of human disease. 4 In an effort to refine the existing models of mouseovarian carcinoma, Dinulescu and colleagues 3  set out to create a genetic model of endometrioid ovar-ian carcinoma, a compelling goal for two reasons.First, existing mouse models give rise to either un-differentiated ovarian cancer or the common seroushistologic variant, and models producing more dif-ferentiated or less common histologic variants (that is, endometrioid, clear-cell, and mucinous ovariancancer) would be of great value. Mutational activa-tion of the K- RAS  oncogene occurs in a small frac- Figure 1. (facing page). Making a Mouse Model of Gyne-cologic Disease. Dinulescu et al. 3  genetically engineered mice to carry la-tent alleles of active mutant K -ras (left side) or active mu-tant K -ras  and inactive Pten  (right side). Injection of an adenoviral Cre recombinase construct into the ovarian bursa led to tissue-specific expression of active mutant K -ras,  resulting in pelvic endometriosis, or tissue-specific expression of active mutant K -ras  and inactivation of Pten,  resulting in metastatic endometrioid ovarian carcinoma. Copyright © 2005 Massachusetts Medical Society. All rights reserved. Downloaded from www.nejm.org at HOSPITAL CLINICO STGO on May 31, 2005 .  n engl j med 352;21 www.nejm.orgmay 26, 2005 clinical implications of basic research 2241 Adenoviral Cre recombinaseLatent active mutant K- ras Latent active mutant K- ras Latent inactive Pten Active mutant K- ras Active mutant K- ras Inactive Pten OvaryCapsuleOviductPeritoneal spaceOvarian bursa Metastatic endometrioid ovarian carcinoma Metastatic endometrioid Pelvic endometriosis Adenoviral Cre recombinase Copyright © 2005 Massachusetts Medical Society. All rights reserved. Downloaded from www.nejm.org at HOSPITAL CLINICO STGO on May 31, 2005 .  clinical implications of basic research 2242 n engl j med 352;21 www.nejm.orgmay 26, 2005 tion of endometrioid ovarian carcinomas, and mu-tational inactivation of the PTEN   tumor-suppressorgene is found in a substantial fraction of endometri-oid ovarian cancers (but not in the other histolog-ic types), so the candidate genetic targets were inhand. Second, approximately 30 percent of en-dometrioid ovarian carcinomas (and a smaller frac-tion of clear-cell carcinomas) are associated withendometriosis, 5  but the molecular relationship of this common benign gynecologic entity to ovariancarcinoma remains obscure.In an elegant application of Cre recombinasetechnology, Dinulescu et al. attained these goals ina two-phase study (Fig. 1). 3  First, they generatedmice with a mutationally activated K -ras   gene, whoseexpression was induced by the presence of Cre re-combinase. After the delivery of a recombinant ad-enoviral vector expressing Cre recombinase to theovarian bursa, which induced the expression of ac-tivated K -ras,  benign endometrioid lesions involv-ing the ovarian epithelium developed in all the mice(Fig. 1). Remarkably, however, extensive peritonealendometriosis, often extending into the abdomen,also developed in approximately half the mice.These lesions met all the established histopatho-logical criteria for the human-disease counterpart,thus representing the first mouse model of sponta-neous human endometriosis. Additional data in-formed a long-standing debate over the histologicsrcin of endometriosis. The coelomic-metaplasiahypothesis, as its name suggests, involves the gen-esis of endometrioid lesions within the peritonealcavity. The data of Dinulescu et al. provide support for an alternative hypothesis, which states that the lesions are initiated by endometrium refluxedthrough the fallopian tubes into the peritoneal cav-ity. This model should prove invaluable for increas-ing our understanding of the natural history of en-dometriosis, its molecular and cell biology, and itssusceptibility to various treatments. It is difficult tooverstate the importance of this unique resource tothe gynecology-research community, in the light of the high prevalence of this disease and its effect on women’s health.In the second phase of the study, the mice in which K- ras   could be activated were crossed withmice that had a Pten  gene flanked by stretches of DNA targeted by recombinase. Injection of the ad-enoviral Cre recombinase construct into the ovari-an bursal tissue resulted in mice that expressed mu-tant K- ras   but lacked Pten . In these mice, invasiveendometrioid carcinoma of the ovary developed within 7 to 12 weeks after the injection (Fig. 1).Gross and histopathological examination at autop-sy revealed frequent hemorrhagic ascites and lungmetastases, an invariant srcin of cancer in the ovar-ian surface epithelium, and all of the histologic hall-marks of human endometrioid ovarian carcinoma.Thus, this model rigorously recapitulates the cor-responding human disease. Important clinical im-plications are evident from additional data show-ing that in tumor implants, the proto-oncogenicpathway    phosphatidylinositol 3' kinase–Akt–mam-malian target of rapamycin (mTOR), which is sup-pressed by PTEN, and p70 S6 kinase (a downstreamtarget of mTOR) were activated, as was MAP kinase,a downstream effector of oncogenic K- ras,  suggest-ing that this model will be helpful in testing the ef-ficacy of specific inhibitors of these pathways (suchas rapamycin and the small molecule PD 184352) inthe treatment of this subtype of ovarian carcinoma.These sophisticated genetic models of gyneco-logic disease illustrate the enormous power andpromise of conditional manipulation of the mousegenome in advancing our understanding of thecause and course of human diseases, as well asproviding essential tools for experimental thera-peutics. From the Departments of Medicine and Surgery, MemorialSloan-Kettering Cancer Center, New York. 1. Orsulic S, Li Y, Soslow RA, Vitale-Cross LA, Gutkind JS, VarmusHE. Induction of ovarian cancer by defined multiple genetic changesin a mouse model system. Cancer Cell 2002;1:53-62. 2. Connolly DC, Bao R, Nikitin AY, et al. Female mice chimeric forexpression of the simian virus 40 TAg under control of the MISIIR promoter develop epithelial ovarian cancer. Cancer Res 2003;63:1389-97. 3. Dinulescu DM, Ince TA, Quade BJ, Shafer SA, Crowley D, JacksT. Role of K-ras   and Pten  in the development of mouse models of endometriosis and endometrioid ovarian cancer. Nat Med 2005;11:63-70. 4.  van der Weyden L, Adams DJ, Bradley A. Tools for targetedmanipulation of the mouse genome. Physiol Genomics 2002;11:133-64. 5. Seidman JD, Russell P, Kurman RJ. Surface epithelial tumors of the ovary. In: Kurman RJ, ed. Blaustein’s pathology of the femalegenital tract. 5th ed. New York: Springer-Verlag, 2002:791-904. Copyright © 2005 Massachusetts Medical Society. Copyright © 2005 Massachusetts Medical Society. All rights reserved. Downloaded from www.nejm.org at HOSPITAL CLINICO STGO on May 31, 2005 .
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