SPECIAL ARTICLE

The Origin and Pathogenesis of Epithelial Ovarian : A Proposed Unifying Theory Robert J. Kurman, MD and Ie-Ming Shih, MD, PhD

assume that the is the source of these ovarian Abstract: is the most lethal gynecologic neoplasms. Finally, preliminary data suggest that mucinous and . Efforts at early detection and new therapeutic transitional (Brenner) tumors arise from transitional-type approaches to reduce mortality have been largely unsuccessful, epithelial nests at the tubal-mesothelial junction by a process because the origin and pathogenesis of epithelial ovarian cancer of . Appreciation of these new concepts will allow are poorly understood. Despite numerous studies that have for a more rationale approach to screening, treatment, and carefully scrutinized the for precursor lesions, none have prevention that potentially can have a significant impact on been found. This has led to the proposal that ovarian cancer reducing the mortality of this devastating disease. develops de novo. Studies have shown that epithelial ovarian cancer is not a single disease but is composed of a diverse group Key Words: ovarian carcinogenesis, type I and type II tumors, of tumors that can be classified based on distinctive morphologic p53 mutation, KRAS, BRAF, PTEN, PIK3CA mutations and molecular genetic features. One group of tumors, designated (Am J Surg Pathol 2010;34:433–443) type I, is composed of low-grade serous, low-grade endo- metrioid, clear , mucinous and transitional (Brenner) carci- nomas. These tumors generally behave in an indolent fashion, are confined to the at presentation and, as a group, are relatively genetically stable. They lack mutations of TP53, but ‘‘Nothing will come from nothing’’ each histologic type exhibits a distinctive molecular genetic profile. Moreover, the exhibit a shared lineage with King Lear, Act I the corresponding benign cystic neoplasm, often through an The origin and pathogenesis of epithelial ovarian intermediate (borderline tumor) step, supporting the morpho- cancer has perplexed investigators for decades. Despite logic continuum of tumor progression. In contrast, another numerous studies that have carefully scrutinized the group of tumors, designated type II, is highly aggressive, evolves ovaries for precursor lesions, none have been found. This rapidly and almost always presents in advanced stage. Type II has led to the proposal that ovarian cancer develops tumors include conventional high-grade serous , de novo.2 ‘‘Nothing will come from nothing,’’ but each undifferentiated carcinoma, and malignant mixed mesodermal year in the United States, approximately 21,550 women tumors (). They displayTP53 mutations in over develop ovarian cancer ‘‘de novo,’’ and 14,600 women die 80% of cases and rarely harbor the mutations that are found from this disease.18 Ovarian cancer is, in fact, the most in the type I tumors. Recent studies have also provided cogent lethal gynecologic malignancy. It is clear that de novo evidence that what have been traditionally thought to be reflects our ignorance about the early events of ovarian primary ovarian tumors actually originate in other pelvic organs carcinogenesis rather than our insight into its perplexing and involve the ovary secondarily. Thus, it has been proposed origin. The time-honored concepts that have forged our that serous tumors arise from the implantation of views of ovarian carcinogenesis can be summarized as (benign or malignant) from the . Endometrioid follows: (1) although it is recognized that there are and clear cell tumors have been associated with profound differences among the various histologic types, that is regarded as the precursor of these tumors. As it is the vast majority of ovarian carcinomas are high-grade generally accepted that endometriosis develops from endome- serous carcinomas and therefore ovarian cancer is trial by retrograde menstruation, it is reasonable to regarded as a single disease; (2) ovarian cancer originates from the ovarian surface epithelium () that invaginates into the underlying resulting in From the Departments of , Gynecology and Obstetrics and inclusion that eventually undergo malignant trans- Oncology, The Johns Hopkins University School of Medicine, Baltimore, Maryland. formation; (3) ovarian cancer spreads from the ovary to Supported by the grants: DOD-OCRP 080469, RO1CA116184, the pelvis, abdomen, and distant sites. On the basis of these RO1CA129080, RO1CA103937. views of ovarian carcinogenesis, efforts at improving the Correspondence: Robert J. Kurman, MD, Departments of Pathology, survival have focused on early detection of ovarian can- Gynecology and Obstetrics, and Oncology, The Johns Hopkins University School of Medicine, Baltimore, Maryland (e-mail: cer, when it is still confined to the ovary, and on the devel- [email protected]). opment of new chemotherapeutic drugs and routes of Copyright r 2010 by Lippincott Williams & Wilkins delivery irrespective of the histologic type. Unfortunately,

Am J Surg Pathol  Volume 34, Number 3, March 2010 www.ajsp.com | 433 Kurman and Shih Am J Surg Pathol  Volume 34, Number 3, March 2010 these efforts have not been successful as evidenced by the mirrored by marked differences in their molecular genetic fact that the overall survival for women with ovarian features.7 As a group, type I tumors are genetically more cancer has not changed over the last 50 years. The reasons stable than type II tumors and display specific mutations for this are that the concepts of , on which in the different histologic cell types.21 Thus, KRAS, these approaches are based, are flawed. BRAF, and ERBB2 mutations occur in approximately Recent morphologic and molecular genetic studies two thirds of low-grade serous carcinomas, whereas TP53 have illuminated our understanding of ovarian carcino- mutations are rare in these tumors. Low-grade endome- genesis in ways that have been quite unexpected and have trioid carcinomas have aberrations in the Wnt signaling challenged the conventional wisdom regarding their pathway involving somatic mutations of CTNNB1 origin and development. Indeed, they have resulted in a (encoding b-), PTEN and PIK3CA.7 Mucinous paradigm shift that has important implications for research carcinomas have KRAS mutations in more than 50% of and for radically changing our approaches to early detec- specimens.1,28 Clear cell carcinoma is unique in that it has tion, prevention, and treatment. a high percentage of PIK3CA activating mutations when purified tumor samples and cell lines are analyzed.22 There is little available molecular genetic data on THE MORPHOLOGIC AND MOLECULAR transitional cell (Brenner) tumors. High-grade serous HETEROGENEITY OF EPITHELIAL carcinoma, the prototypic type II tumor is characterized OVARIAN CANCER by very frequent TP53 mutations (>80% of cases) and One of the major problems in elucidating the patho- CCNE1 (endcoding cyclin E1) amplification but rarely genesis of ovarian cancer is that it is a heterogeneous mutations that characterize most type I tumors, such as disease composed of different types of tumors with widely KRAS, BRAF, ERBB2, PTEN, CTNNB1, and PIK3- differing clinicopathologic features and behavior. On the CA.7 Although only a small number of malignant mixed basis of a series of morphologic and molecular genetic mesodermal tumors have been analyzed molecularly, the studies, we have proposed a dualistic model that categorizes few that have been analyzed, display a similar molecular various types of ovarian cancer into two groups desig- genetic profile. In summary, type I tumors as a group, are nated type I and type II.43 Type I tumors are clinically genetically more stable than type II tumors and display a indolent and usually present at a low stage. They exhibit a distinctive pattern of mutations that occur in specific cell shared lineage between benign cystic neoplasms and the types (low-grade serous, low-grade endometrioid, clear corresponding carcinomas, often through an intermediate cell and mucinous). In contrast, the type II tumors (high- (borderline tumor) step, supporting the morphologic grade serous, high-grade endometrioid, malignant mixed continuum of tumor progression in these neoplasms. This mesodermal tumors, and undifferentiated carcinomas) stepwise sequence of events parallels the -carcinoma show greater morphologic and molecular homogeneity, sequence that occurs in colorectal carcinoma. Type I tu- are genetically unstable, and have a very high frequency mors include low-grade serous, low-grade endometrioid, of TP53 mutations. These findings suggest that different clear cell and mucinous carcinomas. In contrast to the types of ovarian carcinomas develop along different clear-cut and distinctive morphologic differences among molecular pathways. type I tumors, the morphologic differences among the type II tumors are more subtle and, as a result, there is considerable overlap in the diagnosis of these tumors by THE CELL OF ORIGIN OF MOST EPITHELIAL different pathologists. Type II tumors exhibit papillary, OVARIAN CANCER IS NOT OVARIAN glandular, and solid patterns and are diagnosed as high- The cell of origin of ovarian cancer and the grade serous, high-grade endometrioid. and undifferen- mechanisms by which cancer develops have been long tiated carcinomas depending on the dominant pattern. debated. The traditional view of ovarian carcinogenesis Generally, most pathologists classify them as high-grade has been that the various tumors are all derived from the serous carcinomas even though they bear little resem- ovarian surface epithelium (mesothelium), and that blance to tubal-type epithelium (the basis for typing a subsequent metaplastic changes lead to the development tumor as serous); arguably many of those lacking distinc- of the different cell types [serous, endometrioid, clear cell, tive serous or endometrioid features could be classified as mucinous, and transitional cell (Brenner)], which mor- ‘‘high-grade ’’. In addition to these neo- phologically resemble the epithelia of the fallopian tube, plasms, malignant mixed mesodermal tumors (carcino- endometrium, , or endocervix and ) are included in the type II category, because , respectively. The normal ovary, how- they have epithelial components identical to the pure type ever, has no constituents that resemble these tumors. II carcinomas. Type II tumors are highly aggressive and Moreover, the , endometrium, and fallopian tubes almost always present in advanced stage. As they account are derived from the mu¨llerian ducts, whereas the ovaries for approximately 75% of all epithelial ovarian carcino- develop from mesodermal epithelium on the urogenital mas and have relatively similar morphologic features and ridge separate from the mu¨llerian ducts. Therefore, an a uniformly poor outcome, ovarian cancer has been alternate theory proposes that tumors with a mu¨llerian erroneously regarded as a single disease. The morphologic phenotype (serous, endometrioid, and clear cell) are derived differences between type I and type II tumors are from mu¨llerian-type tissue and not from mesothelium.11

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This mu¨llerian-type tissue (columnar epithelium, often and therefore do not qualify as mu¨llerian-type tumors. A ciliated) lines cysts located in paratubal and paraovarian similar problem exists for transitional cell (Brenner) locations that have been referred to collectively as the tumors resembling urothelium that is not mu¨llerian in ‘‘secondary mu¨llerian system’’.23 According to this origin. theory, ovarian tumors develop from these cysts. As the The most compelling evidence suggests that the vast tumor enlarges, it compresses and eventually obliterates majority of what seems to be primary ovarian , ovarian tissue resulting in an adenxal tumor that seems to namely serous, endometrioid, and clear cell carcinomas, have arisen in the ovary. More recently, another theory are derived from the fallopian tube and endometrium and has been advanced, which argues that the majority of not directly from the ovary. Sporadic reports of tubal ovarian carcinomas that are high-grade serous carcino- carcinoma and ‘‘dysplasia’’ had been reported in the mas, arise from high-grade intraepithelial serous carcino- past,15 but in 2001, a group of Dutch investigators mas in the fallopian tube, which then spread to the ovary. described these lesions that closely resemble high-grade These conflicting views led us to undertake a review of the ovarian serous carcinoma in women with a genetic literature in an effort to determine which of the theories is predisposition to ovarian cancer.33 This was a surprising best able to explain the various aspects of ovarian finding, as numerous studies over the past two decades that carcinogenesis. carefully examined the ovaries of women with a genetic Evaluating these theories is problematic, because it predisposition to ovarian cancer never reported similar is difficult to construct experimental systems to test their lesions. In addition, other studies of normal appearing validity. Accordingly, our evaluation is based on critical ovaries contralateral to sporadic (nonhereditary) unilat- analysis of these studies in the light of observations we eral ovarian carcinomas had never identified a convincing have made in the course of pathologic examination of precursor lesion. These latter studies reported a number ovarian tumors. The discussion that follows is an attempt of morphologic changes in grossly normal appear- to distill the most plausible components from the various ing ovaries, such as an increased number of surface theories of cellular origin and integrate them with the papillae and cortical inclusion cysts, including some dis- clinicopathologic and molecular genetic data from the playing minor degrees of atypia. The data, however, have dualistic model to construct a unifying theory of ovarian been conflicting, some studies reporting a significant carcinogenesis. difference of these changes in cases versus controls, and The theory of origin from ovarian surface epithe- other studies reporting no difference. In any event, none lium (mesothelium) has a number of limitations. Histo- of these changes even remotely resemble high-grade serous logically, the single layer of generally attenuated mesothelium carcinoma. It was precisely because of a lack of convinc- overlying the ovaries bears no resemblance to serous, ing precursor lesions that the de novo hypothesis was endometrioid, mucinous, clear cell, or transitional (Bren- invoked. ner) carcinomas. As noted above to account for this apparent In hindsight, because it was assumed that precursors contradiction, it was proposed that the mesothelium over- of ovarian carcinoma would logically be in the ovaries, lying the ovary invaginates into the underlying stroma to the fallopian tubes were not carefully examined.10,42 form so-called ‘‘cortical inclusion cysts’’. These cysts Subsequent studies, in which fallopian tubes were more under the influence of local factors, possibly steroid hor- carefully examined, confirmed that in situ and small, early mones, undergo a metaplastic change, which results in the invasive tubal carcinomas occurred in women with a mesothelium being converted to mu¨llerian-type epithe- genetic predisposition for the development of ovarian lium. These inclusion cysts, with their newly acquired cancer.4,5,8,12,27,29,41 This led to fallopian tube carcinoma mu¨llerian phenotype, can then undergo malignant trans- being included as part of the cancer spectrum associated formation resulting in carcinomas corresponding to the with inherited BRCA mutations. It was subsequently different cell types (serous, endometrioid and clear cell proposed that a proportion of ovarian carcinomas might carcinomas).6 Although cortical inclusion cysts lined by develop as a result of implantation of malignant cells ciliated (mu¨llerian-type epithelium) are frequently obser- from the tubal carcinoma to the ovary.34,35 The next ved in the ovarian , well-documented examples important step linking what had been termed ‘‘tubal of what can be interpreted as a transition from these cysts intraepithelial carcinoma’’ and, subsequently, ‘‘serous tubal to carcinoma have not been reported. Moreover, cortical intraepithelial carcinoma’’ (STIC) with ovarian carcinoma inclusion cysts lined by intestinal-type epithelium to was the observation that over 70% of sporadic (non- account for the development of mucinous carcinomas hereditary) ovarian and peritoneal high-grade serous are distinctly rare. The same can be said for the absence of carcinomas showed mucosal tubal involvement including transitional-type epithelium lining cortical inclusion cysts STICs.19 This observation gave substantial support to the to account for the development of Brenner tumors. proposal that STICs, which almost always are detected in The limitations of the secondary mu¨llerian system the fimbria, may be the source of ovarian high-grade theory are that precursor lesions resembling serous, endomet- serous carcinoma in both women with hereditary muta- rioid, and clear cell carcinomas have rarely, if ever, been tions in BRCA and women who did not have a known reported in paratubal and paraovarian cysts. Moreover, genetic predisposition for ovarian cancer. Although it can the vast majority of mucinous tumors display intestinal be argued that mucosal tubal involvement could represent rather than endocervical-type mucinous differentiation secondary spread from an ovarian carcinoma present in r 2010 Lippincott Williams & Wilkins www.ajsp.com | 435 Kurman and Shih Am J Surg Pathol  Volume 34, Number 3, March 2010 the same specimen, the presence of focal noncontiguous mutations.13,17,41 Like STICs, p53 signatures express g- intraepithelial lesions (STICs) would be an unusual mani- H2AX that localizes to areas of DNA damage in nuclei.24 festation of metastasis. Furthermore, the identification of When associated with STICs and ovarian carcinoma, the STICs in prophylactic specimens from women with a p53 signature has had the identical TP53 mutation as the hereditary predisposition to ovarian cancer, in which STIC and the carcinoma in some cases but not in others. complete microscopic evaluation of the fallopian tubes Based on these findings, a sequence of pathogenetic and ovaries failed to identify invasive carcinoma in these events has been proposed beginning with genotoxic organs, lends additional support to the concept that the DNA damage, followed by TP53 mutation and progres- serous neoplastic process may well begin in the fallopian sive loss of cell cycle control, which then eventuate in the tube rather than in the ovary. Further support for this development of carcinoma.24 There are a number of argument is the finding that nearly all STICs overexpress questions that must be resolved, however, before this p53, similar to high-grade serous carcinoma (Fig. 1). hypothesis can be completely accepted. First, as noted in Laser capture microdissection studies of these lesions have some instances, TP53 mutations, when present in the p53 shown that they harbor mutated TP53.19 In addition, signature, are not always identical with the mutations in STICs associated with a concomitant ovarian carcinoma the STICs and carcinomas in the same specimen. Second, share not only morphologic features but also identical women at high risk have the same frequency of p53 TP53 mutations indicating a clonal relationship between signatures as women who are not at high risk. Third, the them. Adnexal malignant mixed mesodermal tumors high prevalence of p53 signatures (a third of all women) (another type II tumor) have also been associated with compared with the low prevalence of high-grade serous STICs supporting the existence of a common precursor ovarian carcinoma suggests that either a small minority of lesion for type II tumors.14 Further evidence implicating p53 signatures progress or that they are not related to the fallopian tube rather than ovarian surface epithelium carcinoma. It is conceivable that p53 signatures reflect as the site of origin of serous neoplasms comes from a an appropriate and physiologic upregulation of p53 in gene profiling study showing that the gene expression response to DNA damage based on the observation that profile of high-grade serous carcinoma is more closely TP53 mutations are absent in nearly half of p53 signa- related to the fallopian tube than to the ovarian surface tures. Although the proposal that the p53 signature is a epithelium.25 In addition, high-grade serous carcinomas precursor lesion is intriguing, its role in the genesis of express PAX8, a mu¨llerian marker, but not calretinin, a ovarian high-grade serous carcinoma is far from clear at mesothelial marker (Shih, Unpublished data). this time. As fallopian tubes are more carefully examined A recent finding has been the identification of and these lesions studied, the nature of p53 signatures and benign tubal epithelium, specifically secretory as opposed their relationship to STICs will become better defined. to ciliated cells, that express p53 and in which laser Generally, before a carcinoma acquires the ability capture microdissection studies have reported TP53 to metastasize, it must first invade and gain access to mutations in 57% of cases.24 These lesions termed ‘‘p53 vessels or lymphatics. We have observed that the signatures’’ are found in association with STICs and in fimbria contain a rich angiolymphatic vasculature. More- normal appearing fallopian tubes of women without over, they are in almost direct contact with the basement STICs or carcinoma; they have been observed in approxi- membrane of the tubal epithelium, and therefore a tubal mately one third of women with and without BRCA carcinoma may not need to attain a very large size to invade this highly accessible angiolymphatic network. In addition, invasion in the case of a STIC may not be a necessary prerequisite for dissemination. Tubal intra- AB epithelial carcinomas are morphologically and immuno- histochemically similar to endometrial intraepithelial carcinomas that are regarded as precursors or early forms of uterine serous carcinoma. These lesions have also been termed ‘‘uterine surface serous carcinomas’’. They have been shown to disseminate throughout the peritoneal cavity presumably by the passage of malignant cells through the fallopian tube without requisite myometrial invasion.45 The cells that comprise both endometrial and tubal intraepithelial carcinomas are highly anaplastic and identical morphologically to high-grade serous carci- noma. The lesions form papillary tufts and the constitu- ent cells are loosely cohesive. Presumably, these cells can shed and implant on the surface of the ovary and the FIGURE 1. Serous tubal intraepithelial carcinoma (STIC). A, in the absence of invasive growth in the High magnification. Hematoxylin and eosin stain. B, Immuno- fallopian tube. Evidence supporting this possibility are histochemical stain for p53. Arrows point to STIC and an the reports of positive pelvic washings in women whose asterisk defines the boundary of the lesion. only lesion was a STIC.4

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As earlier noted, in studies of ovarian and primary and not by the usual (type II) pathway that begins with a peritoneal high-grade serous carcinomas in which the TP53 mutation. Third, clear-cut mucosal tubal involve- entire fallopian tubes were carefully sectioned, mucosal ment could have been obscured by overgrowth of the involvement of the tube, including STICs, was identified pelvic carcinoma. Fourth, the fimbria of the fallopian in approximately 70% of cases.19 The question arises as tube normally is in intimate contact with the ovarian to the source of the remaining ovarian carcinomas that surface at the time of ovulation. It is conceivable that lack evidence of tubal involvement. There are a number when the ovarian surface epithelium is disrupted at the of possible explanations. First, despite thorough section- time of ovulation, normal tubal epithelial cells from the ing, a small STIC could have been missed (unpublished fimbria may be dislodged and implant in the ovary to data). Second, occasionally high-grade serous carcinomas form an inclusion (Fig. 2) from which a high-grade are intimately associated with serous borderline tumors serous carcinoma could develop (see below). Evidence to and low-grade serous carcinomas. In these cases, the support this notion is the observation that fallopian tube high-grade tumors have had KRAS mutations identical epithelial cells are easily obtained for culture by flushing to those in the serous borderline tumors and lacked TP53 the fallopian tube (Shih, Unpublished data).34 This mutations.9 This finding suggests that some high-grade mechanism could also explain the development of endo- serous carcinomas arise from low-grade serous tumors salpingiosis, a lesion composed of and papillary

FIGURE 2. Transfer of normal tubal epithelium to the ovary. A, Anatomical relationship of fallopian tube with the ovary at the time of ovulation. The fimbria envelops the ovary. B, Ovulation. The ovarian surface ruptures with expulsion and transfer of the oocyte to the fimbria. The fimbria is in intimate contact with the ovary at the site of rupture. C, Tubal epithelial cells from the fimbria are dislodged and implant on the denuded surface of the ovary resulting in the formation of an inclusion cyst. r 2010 Lippincott Williams & Wilkins www.ajsp.com | 437 Kurman and Shih Am J Surg Pathol  Volume 34, Number 3, March 2010 structures lined by tubal-type epithelium that is found on carcinoma as an intermediate step. According to this peritoneal surfaces in the pelvis, omentum, and beneath view, both low-grade and high-grade serous carcinomas the capsule of pelvic and para-aortic lymph nodes. Endo- are ultimately of tubal (mu¨llerian) origin, and, in a , salpingiosis is frequently found in association with low- the ovary is involved secondarily (Fig. 3). grade serous tumors and has been viewed as a possible It has been well established both by morphologic precursor of these tumors. Finally, the possibility that some and, more recently, by molecular genetic studies that low- high-grade serous carcinomas arise in cortical inclusion grade endometrioid and clear cell carcinomas develop cysts as a metaplastic process from the ovarian surface from endometriotic cysts (endometriomas)that are fre- epithelium rather than from implantation of normal quently associated with implants of endometriosis else- fallopian tube epithelium cannot be entirely dismissed. where in the pelvis.44 Although the precise origin of Direct implantation of tubal epithelium into the endometriosis has not been completely established, ovary to form an inclusion cyst, which in turn is the site specifically, whether it develops in situ in the peritoneum of origin of ovarian serous carcinoma, although not through a process of metaplasia or from retrograde yet shown, is an attractive alternative theory to that of menstrual flow, the preponderance of data favor the latter metaplasia from the surface epithelium (mesothelium). mechanism.3 Admittedly, the former theory is more Implantation of fallopian tube epithelium from the difficult to prove experimentally. Thus, if retrograde fimbria at the time of ovulation when the surface epithe- menstruation accounts for most cases of endometriosis, it lium is disrupted can explain the derivation of low-grade is logical to assume that endometrioid and clear cell tumors and high-grade serous carcinomas. In the case of a low- develop from endometrial tissue (mu¨llerian derived) that grade serous carcinoma, the process develops slowly from implanted on the ovary and therefore the ovary is invol- a serous and then a serous borderline tumor ved secondarily26 (Fig. 4). Of further interest has been the after a KRAS or BRAF mutation, whereas in the case of observation that the eutopic endometrium in women with a high-grade serous carcinoma, the process evolves rapidly, endometriosis exhibits intrinsic molecular abnormalities, presumably from a cortical inclusion cyst after a TP53 including activation of oncogenic pathways. Presumably, mutation with the development of an intraepithelial these changes permit the endometrial tissue to implant,

A B

cystadenoma

inclusion cyst SBT occasionally LG STIC TP53 cells HGHG

FIGURE 3. Proposed development of low-grade (LG) and high-grade (HG) serous carcinoma. A, One mechanism involves normal tubal epithelium that is shed from the fimbria, which implants on the ovary to form an inclusion cyst. Depending on whether there is a mutation of KRAS/BRAF/ERRB2 or TP53, a LG or HG serous carcinoma develops, respectively. LG serous carcinoma often develops from a serous borderline tumor, which, in turn, arises from a serous cystadenoma. Another mechanism involves exfoliation of malignant cells from a serous tubal intraepithelial carcinoma (STIC) that implants on the ovarian surface resulting in the development of a HG serous carcinoma. B, A schematic representation of direct dissemination or shedding of STIC cells onto the ovarian surface on which the carcinoma cells ultimately establish a tumor mass that is presumably arising from the ovary. Of note, there may be stages of tumor progression that precede the formation of a STIC.

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reported that Brenner tumors are small (median size 0.5 cm, range 0.02 to 20 cm), whereas mucinous cystade- nomas are large (median size 9 cm, range 1 to 30 cm). The investigators speculated that as a small Brenner tumor grows, the mucinous component becomes dominant resulting in the development of a that, as it enlarges, compresses and eventually obliterates the adjacent ovary giving the appearance that it arose in CCC the ovary. The findings in this study are intriguing but must be regarded as preliminary. Additional morphologic and molecular genetic studies are necessary to determine whether this concept is valid. EM/CC In summary, none of the existing theories adequately borderline tumor reconciles all aspects of ovarian carcinogenesis. All of endometriosis them have something to offer in explaining the develop- ment of ovarian carcinomas, but none are all inclusive. It does seem that the vast majority of what have been thought to be primary epithelial ovarian and primary peritoneal carcinomas are, in fact, secondary. Thus, the FIGURE 4. Proposed development of LG endometrioid and most persuasive data support the view that serous tumors clear cell carcinoma. Endometrial tissue, by a process of retrograde menstruation, implants on the ovarian surface to develop from the fimbriated portion of the fallopian tube, form an endometriotic cyst from which a LG endometrioid endometrioid, and clear cell tumors from endometrial or clear cell carcinoma can develop. CCC indicates clear cell tissue passing through the fallopian tube resulting in carcinoma of the ovary; EMC, LG endometrioid carcinoma of endometriosis and mucinous, and Brenner tumors from the ovary. transitional-type epithelium located at the tubal-mesothe- lial junction where the fimbria makes contact with the peritoneum. The concept that the majority of epithelial ovarian carcinomas originates outside the ovary and survive, and invade on ovarian and peritoneal surfaces.3 involves it secondarily has emerged only recently, because This hypothesis, by which endometrioid and clear cell in the past, the default diagnosis of carcinomas involving carcinoma develop from endometrial tissue implanted the pelvis and abdomen was that they were ovarian. A on the ovary, is supported by epidemiologic evidence showing carcinoma was classified as tubal in origin only when the that a protective effect for tubal ligation was seen only for bulk of the tumor involved the fallopian tube rather than endometrioid and clear cell carcinoma of the ovary.37 the ovary, and there was evidence of an intraepithelial (in Finally, the derivation of mucinous tumors of situ) tubal carcinoma.39 A diagnosis of primary peritoneal gastrointestinal type and transitional cell (Brenner) tumors carcinoma is even more restrictive. Even with extensive may also not involve the ovaries directly. The origin of tumor involving the peritoneum, omentum, and other these tumors is puzzling, as unlike serous, endometrioid, abdominal organs, a carcinoma is classified as primary and clear cell tumors, they do not display a mu¨llerian ovarian, if there is as little as 5 mm of tumor involving the phenotype. Although it has been argued that these muci- ovaries. Thus, there has been an inherent bias in nous tumors bear some relationship with the endocervix, classifying pelvic tumors as being ovarian in origin. the mucinous epithelium that characterizes these neo- Although the data suggesting that epithelial ovarian plasms more closely resembles gastrointestinal mucosa. It carcinoma arises in extraovarian sites and involves the seems most unlikely that they develop from cortical ovaries secondarily are compelling, serous neoplasms (low- inclusion cysts. as mucinous metaplasia involving cortical and high-grade) involve the ovaries and other pelvic and inclusion cysts is a very rare finding. On the other hand, abdominal organs, such as the omentum and mesentery, the association of Brenner tumors and mucinous tumors much more extensively than the fallopian tubes. Similarly, has been recognized for many years. In a provocative although endometrioid carcinomas develop from endome- study of mucinous and Brenner tumors, it triosis that frequently involves multiple sites in the pelvis, was reported that after extensive sectioning, mucinous these neoplasms are almost always confined to the ovaries. cystadenomas contained foci of Brenner tumor in 18% of It is likely that the propensity for growth in the ovary is cases.40 Interestingly, mucinous tumors were frequently mulifactorial, but the precise reasons for this are unknown. associated with Walthard cell nests that are composed of benign transitional-type epithelium, frequently found in paraovarian and paratubal locations. This raises the IMPLICATIONS FOR RESEARCH, SCREENING, possibility that mucinous tumors and Brenner tumors PREVENTION, AND TREATMENT have the same histogenesis arising from these microscopic The implications of this new paradigm of ovarian transitional cell nests at the tubal-mesothelial junction in carcinogenesis for investigators, clinicians, and women keeping with their nonmu¨llerian appearance. The study are significant. For researchers, the implication of tubal r 2010 Lippincott Williams & Wilkins www.ajsp.com | 439 Kurman and Shih Am J Surg Pathol  Volume 34, Number 3, March 2010 origin of ovarian serous carcinoma challenges many of the malignant mixed mesodermal tumors ()] earlier reports showing ‘‘overexpressed’’ ovarian cancer- originate outside the ovary illustrates the underlying flaws associated genes in which their expression levels in carci- in screening approaches designed to detect these tumors noma are almost always compared with their ‘‘normal’’ while confined to the ovary. Moreover, type II tumors counterpart, ovarian surface epithelium. As the gene represent approximately 75% of all ovarian carcinomas expression profiles in ovarian surface epithelium that is of and are responsible for 90% of ovarian cancer deaths.16 It mesothelial origin, are distinct from fallopian tube epithe- is the type II tumors that should be targeted for screening, lium which is of mu¨llerian origin, experiments in which but unfortunately these tumors are rarely confined to the ovarian surface epithelium (mesothelium) has been used ovary, even at their inception. In a study of nearly 400 as a control may not be valid. Whether the overexpressed patients who were carefully staged from the Washington genes that have been reported are indeed upregulated, Center Hospital in Washington DC, which is largely a when they are compared with the more likely source of primary care hospital, less than 1.25% of high-grade ovarian serous carcinoma, that is, fallopian tube epithe- serous carcinomas were confined to the ovary (Seidman et lium, needs to be revisited. In fact, a recent molecular al, unpublished data). Similarly, the British Columbia genetic study showed that the different histologic types of Tumor Registry reported that only 0.5% of high-grade ovarian cancer do indeed display distinct expression serous carcinomas were limited to the ovary at diag- profiles that are concordant with the normal tissues they nosis.38 The futility of detecting early-stage ovarian resemble and show little similarity to ovarian surface cancer was recently underscored in a large multiinstitu- epithelium (mesothelium). Thus, the genes expressed tional prospective study [, , Colorectal, and in serous carcinoma were similar to those expressed in Ovarian Cancer Screening Trial] in which, despite intensive normal fallopian tube, whereas the expression profiles of annual screening of nearly 35,000 women with CA 125 endometrioid and clear cell carcinomas resembled endome- and transvaginal ultrasound, 70% of the women presented trial epithelium. Interestingly, the expression profile of with advanced stage disease. This was no different from mucinous tumors resembled normal colonic epithelium.25 unscreened populations.31 For the type II tumors, the We have also observed (unpublished data) that PAX8, a goal in screening should be the detection of low volume, marker of mu¨llerian-type epithelium, is expressed in ovarian not low stage disease. This can only be accomplished by serous carcinoma but not in ovarian surface epithelium developing a panel of sensitive and specific biomarkers (mesothelium), whereas calretinin, a mesothelial marker, that are expressed early in ovarian carcinogenesis. reacts with ovarian surface epithelium and As with early detection, the treatment of type I but not with tubal epithelium or ovarian serous carci- and type II tumors must be individualized. Type I tumors noma (Fig. 5). In the future, the analysis of overexpressed are generally low-grade, slow growing and localized to genes in ovarian cancer should take into account the the ovary at diagnosis spreading late in their evolution. histologic type of the tumors being studied and the data Accordingly, when confined to the ovary, salpingo- compared with the appropriate normal tissue. oophorectomy may suffice. On the other hand, when they From a clinical perspective, the implications of this have spread beyond the ovary, chemotherapeutic agents new paradigm are even more far reaching. For the last that are effective against the more rapidly proliferating type two decades, numerous studies, including large clinical II tumors are not as effective for type I tumors, because the trials, have been conducted in an effort to develop screening latter are slow growing. Therefore, new approaches for tests for ovarian cancer. The goal of these studies is to advanced- stage type I tumors are needed. Deregulation of detect tumors, when they are still confined to the ovaries, protein kinase activity as a result of somatic mutation in thereby increasing the likelihood of cure and reducing the these genes occurs in many type I tumors. Mutations in mortality of the disease. The modalities that are currently these genes constitutively activate the signaling pathways being used to screen women are pelvic examination, they control, and tumor cells with mutations become transvaginal ultrasound, and measurement of serum CA dependent on those mutations for progression. Therefore, 125. An awareness of the dualistic model, which high- these genes could provide potential targets for therapeutic lights the heterogeneity of ovarian carcinoma, clearly intervention. For example, in many type I carcinomas, indicates that one screening test will not be effective in there is constitutive activation of the MAPK signaling detecting all the different types of ovarian carcinomas. pathway because of mutations in ERBB2, KRAS or Type I tumors (low-grade serous, low-grade endome- BRAF, the upstream regulators of MAPK. It is therefore trioid, clear cell, and mucinous) are slow growing and conceivable that BRAF inhibitors and other MAPK kinase attain a large size while still confined to the ovary. They inhibitors could prolong disease-free interval and improve are easily detected by pelvic examination and/or transva- overall survival in patients with these types of advanced ginal ultrasound. They constitute, however, only 25% of stage type I tumors, when combined with conventional ovarian cancers and account for approximately 10% of therapeutic modalities. ovarian cancer deaths.16 Therefore, it can be argued that The approach to the treatment of type II tumors the development of a biomarker screening test is not should be completely different from that of the type I urgently needed for type I tumors. More importantly, tumors. Treatment for type II tumors should be initiated the recognition that the majority of type II tumors on the basis of detection of sensitive and specific [high-grade serous and undifferentiated carcinomas, and biomarkers before the appearance of morphologically

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H&E PAX8 calretinin

ovarian surface epithelium

fallopian tube epithelium

“ovarian” serous carcnoma

FIGURE 5. Comparison of the immunohistochemical staining pattern for ovarian surface epithelium (mesothelium), normal fallopian tube epithelium, and HG serous carcinoma. PAX8 is a marker of mu¨llerian-type epithelium, such as fallopian tube epithelium, and calretinin is a marker of mesothelium. recognizable disease, when therapy will likely be more using oral contraceptives for 5 or more years.36 Parity has effective. A precedent exists for this approach, as women also been shown to be protective conferring approximately with hereditary BRCA mutations are treated on the basis a 50% decrease in risk compared with nulliparity.32 of that information only. Another important treatment Accordingly, the entire approach to prophylaxis, not issue that needs to be considered is whether patients found only for women at high risk of developing ovarian cancer to have a STIC require adjuvant chemotherapy. The but also for the general female population, needs to be finding of positive pelvic washings in patients with only a reevaluated in the light of the evolving new paradigm of STIC indicates that these microscopic lesions can shed ovarian carcinogenesis as discussed here. The traditional malignant cells.4 At present there is no consensus as to approach for reducing risk for women with a family whether or not these women should be treated. This will history of ovarian carcinoma or who are found to have have to be determined by a randomized clinical trial. BRCA1/2 mutations has been hysterectomy and bilateral Finally, the mounting evidence that ovarian cancer salpingo-oophorectomy. The ovarian tumors that develop does not develop in the ovary, and the lack of success of are almost always high-grade serous carcinomas and ovarian cancer screening provides a strong rationale for there has been no convincing evidence that these women directing efforts at primary prevention. It has been well are at a higher risk of developing uterine serous carci- established in epidemiologic studies that the use of oral nomas. If it can be unequivocally shown that the serous contraceptives reduces the risk of ovarian cancer sub- carcinomas in these women develop almost exclusively in stantially. The risk is reduced by about 50% for women the fimbria, then salpingectomy alone would be sufficient r 2010 Lippincott Williams & Wilkins www.ajsp.com | 441 Kurman and Shih Am J Surg Pathol  Volume 34, Number 3, March 2010 to reduce the risk of ovarian cancer. This approach would Recent studies on the origin of ovarian cancer have have to be evaluated in a randomized clinical trial compar- directed attention to a putative precursor lesion in the ing it to the standard treatment of bilateral salpingo- fallopian tube that morphologically and molecularly oophorectomy. For women who are not considered to be resembles high-grade ovarian serous carcinoma, and that at high risk but who undergo a hysterectomy for benign has been designated ‘‘serous intraepithelial tubal carcino- uterine disease, many gynecologists have argued that bi- ma (STIC)’’. Thus, rather than developing de novo from lateral oophorectomy should be carried out to reduce the the ovary, as earlier proposed, the majority of type II risk of developing ovarian cancer. In a recent prospective tumors seem to arise from a STIC in the fimbriated end study of nearly 30,000 women in the Nurses’ Health Study, of the fallopian tube that spreads to the ovary. Another it was shown that compared with ovarian conservation, possible mechanism for the development of ‘‘ovarian’’ bilateral oophorectomy at the time of hysterectomy was carcinoma is dislodgement of normal tubal epithelium associated with an increased risk of all-cause mortality, from the fimbria, which implants on the site of rupture fatal and nonfatal coronary disease, and .30 where ovulation occurred resulting in the formation of Accordingly, for women undergoing a hysterectomy for an inclusion cyst that may then undergo malignant benign uterine disease, removal of only the fallopian tubes transformation. Thus, serous tumors may develop from with sparing of the ovaries would improve quality of life inclusion cysts, as has been thought, but by a process of and overall survival while still reducing the risk of ovarian implantation of tubal (mu¨llerian-type) tissue rather than carcinoma. Such an approach has important public health by a process of metaplasia from ovarian surface epithelium implications, as approximately 300,000 women in the (mesothelial). Endometrioid and clear cell carcinomas United States undergo elective oophorectomy each year. may also originate from nonovarian, mu¨llerian-type tissue, as it is widely accepted that these tumors develop from endometriosis that is thought to develop as a result of retrograde menstruation. The origin of mucinous and CONCLUSIONS transitional cell (Brenner) tumors is still not well established, A new paradigm for the pathogenesis of ovarian although recent data suggest a possible origin from cancer based on a dualistic model and the recognition that transitional epithelial nests located in paraovarian loca- the majority of ‘‘ovarian’’ carcinomas originate outside the tions. Thus, there is mounting evidence that type I and ovary assist in organizing this complex group of neoplasms type II ovarian tumors develop independently along diffe- and facilitates the development of new and novel approa- rent molecular pathways, and that both types develop ches to prevention, screening, and treatment. One group outside the ovary and involve it secondarily. This explains of tumors (type I) is generally indolent, presents in stage I why current screening strategies designed to detect ovarian (tumor confined to the ovary) and develops from well- cancer, when it is confined to the ovary, are ineffective in established precursors, so-called borderline tumors. These accomplishing this goal. tumors are characterized by specific mutations, including Given the obstacles in early detection (screening) KRAS, BRAF, ERBB2, CTNNB1, PTEN and PIK3CA, and the significant but relatively limited success in treatment, but rarely TP53. They are relatively genetically stable. attention should be directed to primary prevention. This The other group (type II) is composed of tumors that are takes on particular relevance with the recognition that the aggressive, present in advanced stage, and develop from majority of ovarian carcinomas are derived from cells in intraepithelial carcinomas in the fallopian tube. They the fallopian tube or from passage of the endometrial have a very high frequency of TP53 mutations but rarely tissue through the fallopian tubes and the important role harbor the mutations detected in type I tumors. They are of ovulation in ovarian carcinogenesis. Salpingectomy genetically highly unstable. alone may be sufficient to accomplish this, as removal of This proposed model is intended to serve as a the fallopian tubes would reduce the risk of ovarian framework for studying ovarian cancer. It is not complete cancer while preserving ovarian function. Ovarian conser- and does not resolve all issues. For example, clear cell vation seems to be particularly important for a woman’s carcinoma is classified as a type I tumor based on having health, as it has been shown that oophorectomy is a characteristic PIK3CA mutation, relative genetic associated with increased overall mortality and a higher stability, frequent presentation in stage I, and association frequency of nonfatal coronary heart disease. Other with endometriosis, a well-established precursor lesion. approaches should also be explored, for example the use But unlike other type I tumors, clear cell carcinoma is of oral contraceptives that presumably by preventing high-grade at presentation. The inability to reconcile all ovulation reduces the risk of ovarian cancer by as much of the many issues relating to ovarian pathogenesis does as 50%. In any case, new diagnostic, prevention and not invalidate or negate the utility of the paradigm. therapeutic approaches must be developed on the basis of Thomas Kuhn, who introduced the concept of paradigms our evolving understanding of ovarian carcinogenesis. as a way of explaining how science progresses, pointed out: ‘‘To be accepted as a paradigm, a theory must seem better than its competitors, but it need not, and in fact ACKNOWLEDGMENTS never does, explain all the facts with which it can be The authors thank Drs Kathleen Cho, Lora Hedrick confronted.’’20 Ellenson, Brigitte M. Ronnett, Jeffrey, D. Seidman, and

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