ANATOMIC PATHOLOGY Original Article

Evidence for the Transformation of Seminoma to Yolk Sac Tumor, with Histogenetic Considerations JOSEPH T. CZAJA, M.D., AND THOMAS M. ULBRIGHT, M.D.

pected changes in reactivity at the foci of such transformation. Four additional cases were regarded as either seminomas with artifactual microcystic change or the close association of seminoma and yolk sac tumor but lacking evidence for transformation. These data support the theory that seminoma is not an "endpoint" neoplasm but may serve a precursor role in the progression to nonseminomatous germ cell tumors. (Key words: Seminoma; Yolk sac tumor; Germ cell tumor; Testicular neoplasm; Histogenesis) Am J Clin Pathol 1992;97:468-477

The pathogenesis of testicular germ cell tumors has long been an area of controversy. One aspect of this subject that has stimulated considerable debate is the role that seminoma plays in the formation of other neoplastic germ cell elements.1"3 Some models of histogenesis describe a separate pathway for the formation of seminoma compared to the nonseminomatous forms of germ cell tumor.4-5 In these schemata, seminomas are "end-stage" neoplasms that do not undergo further morphologic transformation (Fig. 1). A great deal of recent evidence, however, suggests that seminoma plays a more integral role in the development of nonseminomatous germ cell tumors.'" 3,6,7 This concept originally was suggested by Friedman, 8 who thought that seminoma acted as a precursor to all other invasive testicular germ cell tumors. Although it has long been observed that seminomas and nonseminomatous tumors frequently exist together in mixed germ cell neoplasms,9 often in close apposition, 10 "

histologic documentation for differentiation of seminoma to nonseminomatous tumors has been sparse. 12 ' 3 In this study, we present six cases of germ cell tumors that displayed histologic continuity between seminoma and yolk sac tumor. The morphologic findings from these cases, supplemented by immunohistochemical stains for cytokeratin, alpha-fetoprotein (AFP), and placental-like alkaline phosphatase (PLAP), support the conclusion that yolk sac tumor may evolve from seminoma. MATERIALS AND METHODS

Thirty germ cell tumors from testicular cancer patients at Indiana University Hospital were identified prospectively as possibly displaying differentiation of seminoma into yolk sac tumor elements. After reexamination, blocks from 10 of these cases were selected that were thought to demonstrate areas of potential seminoma-yolk sac tumor transformation. All tissue had been fixed in 10% neutral buffered formalin and embedded in paraffin. Sections were cut of at 4 jum, deparaffinized in xylene, and rehydrated From the Department of Pathology, Indiana University School through a graded series of alcohols. Sections to be stained Medicine. Indianapolis. Indiana. for cytokeratins were predigested for 30 minutes in a soReceived July 8, 1991; manuscript accepted for publication August lution of 0.1% trypsin (Difco, Detroit, MI) and 0.1% cal2, 1991. cium chloride in phosphate-buffered saline, pH 7.2. ImPresented in part at the meeting of the American Society of Clinical Pathologists, New Orleans, Louisiana, September 1991. munohistochemical staining was performed with the use Address reprint requests to Dr. Ulbright: Department of Pathology, of the streptavidin-HRP system (HistoMark, Kirkegaard Indiana University Hospital—N340, 926 West Michigan Street, Indiaand Perry Laboratories, Gaithersburg, MD). The antinapolis, Indiana 46202-5280. 468

Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016

Recent ultrastructural, cytogenetic, and ploidy analyses indicate that seminoma acts as a precursor from which other forms of testicular germ cell tumor may originate. Ten cases of primary or metastatic testicular germ cell tumors were investigated that showed possible transformation of seminoma to yolk sac tumor. Such transformation was identified in six cases in which foci of abrupt change from seminoma to various patterns of yolk sac tumor occurred, often at the periphery of otherwise pure lobules of seminoma. Immunostains for cytokeratins, placental-like alkaline phosphatase, and alpha-fetoprotein demonstrated the ex-

CZAJA AND ULBRIGHT Transformation of Seminoma to Yolk Sac Tumor

phatase staining was enhanced by dipping the sections in 0.125% osmium tetroxide.14 All slides were counterstained with hematoxylin, dehydrated, and mounted with coverslips with Permount (Fisher, Fair Lawn, NJ). Positivity was graded as 3+ (strong chromogen reaction visible at scanning magnification), 2+ (reaction visible at intermediate magnification), or 1 + (reaction visible with X40 objective only). The pattern of staining (diffuse, focal, or individual cells) also was noted. Clinical information, including serum AFP studies and follow-up data, was obtained by review of medical charts or contacting referring physicians.

[ Germ Cell j

I Seminoma I

Yolk Sac

469

Somatic

Trophoblastic

bodies included monoclonal anti-cytokeratins (CAM 5.2 [Becton-Dickinson, Mountainview, CA] and AE1/AE3 [Boehringer-Mannheim, Indianapolis, IN]), two anti-AFP antibodies (monoclonal, Zymed, South San Francisco, CA; and polyclonal, Dako, Santa Barbara, CA), and antiplacental alkaline phosphatase (Dako), a polyclonal antibody. Appropriate tissues served as positive controls. Negative controls consisted of staining without applying the primary antibody and the substitution of nonimmune serum for the primary antibody. Chromogenic visualization was accomplished by immersion of the sections in 3,3'-diaminobenzidine solution (0.25 mg/mL with 0.003% hydrogen peroxide in phosphate buffer) for 3 minutes. Placental-like alkaline phos-

RESULTS Of the 10 cases included in the study, seven were primary orchiectomy specimens and three were metastatic tumors resected after the administration of chemotherapy. The patients ranged in age from 18 to 53 years, with a mean age of 29 years. Nine of the patients were treated for mixed germ cell tumor, whereas one (case 10) was treated for pure seminoma. Preoperative AFP levels ranged from normal (four cases) to 5,600 ng/dL. The clinical information is summarized in Table 1. The presence of focal yolk sac tumor elements within lobules of seminoma was demonstrated morphologically and by immunostaining in 6 of the 10 cases. In two cases, components of both seminoma and yolk sac tumor were present in close apposition, but evidence for transformation was not clear. In the remaining two cases, the tumor stained as pure seminoma with foci of microcysts and had no evidence of epithelial differentiation (Fig. 2). These two cases therefore were regarded as pure seminomas with histologic features that mimicked yolk sac tumor.

TABLE 1. CLINICAL AND HISTOLOGIC FEATURES OF 10 CASES SUSPECTED OF SHOWING TRANSFORMATION OF SEMINOMA TO YOLK SAC TUMOR Preop. Case

Age (years)

1 2 3 4 5 6 7 8 9 10

23 22 18 34 28 47 53 28 23 18

APP

Specimen

Preop. Treatment

Tumor Components

(ng/dL)

Transformation Demonstrated?

Follow-up

L testis R testis L testis L testis RP node met R testis PC node L testis Liver met R testis

None None None None Chemotherapy None Chemotherapy None Chemotherapy None

SEM, YST SEM, YST, teratoma SEM, YST, teratoma EC, YST, SEM, STBGCs SEM, YST SEM, YST SEM, YST Teratoma, SEM, YST SEM (with microcystic artifact) SEM (with microcystic artifact)

93.7 848 5600 nl nl NA nl 24.5 424 nl

Yes Yes Yes Yes Yes Yes No No No No

*DOD—3 years NED—1.7 years NED—1.2 years fDOD—2 years NED—3 years Lost to follow-up Lost to follow-up Lost to follow-up NED—4 years NED—2.5 years

AFP = alphafetoprotein; DOD = died of disease; EC = embryonal carcinoma; L = left; met = metastatic tumor; NA = not available; NED = no evidence of disease; nl = normal; PC = pericolic; R = right; RP = retroperitoneal; SEM = seminoma; STBGCs = syncytiotrophoblastic giant cells; YST = yolk sac tumor.

* Had multiple recurrences after chemotherapy and died with liver metastases after autologous bone marrow transplant. t Died of progressive tumor, refractory to chemotherapy, with pulmonary metastases.

Vol. 97 • No. 4

Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016

FIG. 1. "Traditional" view of testicular germ cell tumor histogenesis. This concept suggests that seminoma is incapable of further differentiation. (Adapted from Srigley JR, Mackay B, Toth P, Ayala A. The ultrastructure and histogenesis of male germ cell neoplasia with emphasis on seminoma with early carcinomatous features. Ultrastruct Pathol 1988; 12: 67-86, with permission from the publisher.)

470

ANATOMIC PATHOLOGY Original Article other forms of yolk sac tumor differentiation occurred in these cases, as detailed in Table 2. One case (case 1) displayed a rare endodermal sinus-like structure (Fig. 4), in addition to a microcystic pattern. A third case (case 3) displayed small glands (which were AFP positive; see below) intimately intermingled with seminoma (Fig. 5), whereas two additional cases (cases 4 and 5) displayed a mixed microcystic/glandular pattern. In case 6, microcystic as well as solid yolk sac tumor developed in apparent transformation from seminoma. In addition to the foci of yolk sac tumor that occurred within lobules of seminoma, several of these cases contained separate yolk sac tumor areas showing the usual features of that neoplasm.1516 The patterns in those areas were essentially the same as those that occurred in the areas of transformation.

All cases displayed areas of "classic" seminoma composed of the characteristic polygonal tumor cells arranged in sheets and cords separated by branchingfibrousbands. A variable, interstitial lymphocytic infiltrate was present in all cases, and a granulomatous reaction was identified in several cases. Transformation to yolk sac tumor occurred as several patterns but was most frequently identified at the periphery of seminomatous lobules as distinct, microcystic foci (Fig. 3). These microcystic areas remained confined to the neoplastic lobules, rather than representing invasion into the seminoma from an adjacent yolk sac tumor component. In addition to a microcystic pattern,

FlGS. 3A and B. Case 5. (A) Microcystic yolk sac tumor arising at the periphery of a lobule of seminoma outlined byfibrouspseudocapsule (bottom). Hematoxylin and eosin (X38). (B) Higher-power view demonstrates classic features of seminoma at top, and microcystic yolk sac tumor at bottom. Hematoxylin and eosin (X78). FIG. 4. Case 1. Schiller-Duvall-like structure in a yolk sac tumor arising at the periphery of a seminomatous lobule. Note also the presence of syncytiotrophoblastic giant cells. Hematoxylin and eosin (X168).

A.J.C.P. • April 1992

Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016

FlG. 2. Case 10. Pure seminoma displaying artifactual microcystic areas. This finding may mimic yolk sac tumor differentiation. Hematoxylin and eosin (XI68).

Immunohistologically, the transformation was most readily demonstrated using the two cytokeratin stains. In all six cases, the epithelial yolk sac tumor elements were diffusely and strongly positive, whereas the seminomatous elements were predominantly negative, although scattered cytokeratin-positive seminoma cells were identified in some cases. The change from negative to strongly positive tumor cells was abrupt in four of the cases. Strong cytokeratin positivity, therefore, marked microcystic foci of yolk sac tumor differentiation at the periphery of lobules of seminoma or scattered glandular elements of yolk sac tumor dispersed among seminomatous cells (Figs. 6 and 7). In two cases of seminoma with transformation to microcystic/glandular yolk sac tumor, individual, cytokeratin-positive tumor cells were intermingled with the adjacent seminomatous elements (Fig. 7). It was not clear, however, if such cells represented transitional epithelial forms or simply cytokeratin-positive seminoma cells. A summary of the immunohistochemical studies is provided in Table 2. Staining for polyclonal AFP was strongly positive in the yolk sac tumor component in the areas of transformation in cases 1 and 6, whereas weaker, more focal positivity was shown in cases 2 and 3. The monoclonal antiAFP stained less strongly and displayed more focal positivity in these four cases. Cases 4 and 5 were negative for both the mono- and polyclonal AFP stains. The seminomatous component of all cases was negative for AFP. Placental-like alkaline phosphatase was diffusely and strongly positive for the cell membranes of the seminomatous components in five of the six cases. There also was less intense, more spotty positivity for the accom-

CZAJA AND ULBRIGHT Transformation of Seminoma to Yolk Sac Tumor

471

Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016

1< S* W OV

472

ANATOMIC PATHOLOGY Original Article TABLE 2. PATTERNS OF IMMUNOPEROXIDFASE STAINING IN SIX CASES THAT DEMONSTRATE TRANSFORMATION OF SEMINOMA TO YOLK SAC TUMOR Immunoperoxidase Staining* (Seminoma/Yolk Sac Tumor)

Case 1

2 3 4 5 6

YST Pattern at Transformation Microcystic-ES Microcystic-glandular Microcystic-glandular Glandular Microcystic Microcystic-solid

PLAP

CAM 5.2

AE 113

AFPpoly

AFPmono

2+D/l+D 1+D/l+D 1+D/l+F 3+D/2+F

2+R/3+D —/3+D 2+R/3+D 1+R/3+D 2+R/3+D 1+F/2+D

—/3+D —/3+D 1+R/3+D 1+R/3+D 2+R/3+D 1+F/2+D

—/2+D —/2+F —/3+F —/— —/— —/3+D

—/2+F —/2+F —/3+F —/— —/— —/2+D

—/— 3+D/l+F

* Strength of positivity graded 1+ to 3+; Pattern ofpositivity: D = diffuse; F = focal: R = rare individual cells.

panying yolk sac tumor components in these five cases. In the zones of transformation there was a definite decrease in the intensity of PLAP staining from seminomalike to yolk sac tumor-like components in four of these five cases (Fig. 8). In case 5, both the seminoma and yolk sac tumor components were negative for PLAP, perhaps representing antigen degradation during tissue processing.

tumors containing both histologic types. Generally, as seminoma transformed to yolk sac tumor, staining intensity for PLAP decreased, and the cells became strongly cytokeratin positive. This pattern of PLAP and cytokeratin reactivity in these two neoplastic elements is consistent with previous reports that examined reactivities in these germ cell tumor types.22"25 The transformation of seminoma to yolk sac tumor often was accompanied by the expected acquisition of AFP positivity. Although it may be argued that the documentation of a true transition must be accomplished by the demonstration of strong cytokeratin or AFP positivity in morphologic seminoma cells, our study suggests that biochemical and morphologic transformation are associated, and such evidence will rarely be attained. We did note the presence of cytokeratin positivity in seminomatous cells in these cases, but such a finding has been documented before in pure seminomas,23,25 and its significance in our cases is not clear.

DISCUSSION One widespread view of germ cell tumor histogenesis hypothesizes that primitive malignant germ cells initially differentiate into either seminoma or embryonal carcinoma. 1517 Teratoma, yolk sac tumor, and choriocarcinoma are thought to differentiate from embryonal carcinoma, whereas seminoma is incapable of further differentiation (Fig. 1). Another model of germ cell tumor histogenesis allows for the differentiation of seminomatous cells into embryonal carcinoma cells but does not include a pathway from seminoma to yolk sac tumor. 18 These hypotheses are easily understood and provide a rational explanation for many of the histologic patterns presented by germ cell tumors. In addition, experimental support for the precursor role of embryonal carcinoma has been provided by animal models and xenograft studies.19-21 However, such methodologies have not generally proved fruitful for the investigation of the possible precursor role of seminoma in the formation of germ cell tumors. Direct histologic observation remains a valuable technique in the study of the possible precursor role of seminoma. Our findings suggest that seminoma can play a precursor role for the subsequent development of nonseminomatous elements—a concept that is in keeping with more recently proposed models for the development of testicular germ cell neoplasia.12-6 In six cases of mixed primary or metastatic testicular germ cell tumors, we identified areas of transformation of seminoma to yolk sac tumor. A related observation was made by Friedman and Moore 12 in 1946, who commented on apparent transition between seminoma and embryonal carcinoma in

Why is it that seminoma is transformed to yolk sac tumor and not visa versa? The small size and peripheral location of the yolk sac tumor elements suggest that these cases represent a focal change within a preexisting seminomatous tumor. The peripheral location of such yolk sac tumor differentiation within seminomatous lobules may not be a chance occurrence because the periphery of advanced seminomas is the most actively proliferating area26 and thus would be most susceptible to the genetic changes that probably induce transformation. Other observations support a precursor role for seminoma. Srigley and colleagues' ultrastructurally demonstrated "cell surface specialization in keeping with early carcinomatous transformation" in four cases of histologically typical seminomas. They thought that these cases represented "hybrids" of seminoma and embryonal carcinoma. Studies by Raghavan and associates13 and Monaghan and co-workers27 described xenografts in which human seminomas transplanted into nude mice developed histologic, ultrastructural, and functional features of yolk sac tumor. One of these studies27 also described

A.J.C.P.-April 1992

Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016

AEI/3 and CAM 5.2 = anti-cytokeratins; AFP = alphafetoprotein; ES = endodermal sinus: PLAP = placental alkaline phosphatase; YST = yolk sac tumor.

CZAJA AND ULBRIGHT Transformation of Seminoma to Yolk Sac Tumor

473

FlG. 5. Case 3. Small glandular structure (bottom) arising in seminoma. Hematoxylin and eosin (X205). FIG. 6. Case 1. Strong cytokeratin positivity in microcystic yolk sac tumor arising at the periphery of a lobule of cytokeratin-negative seminoma, Anti-AE1/AE3 immunostain (X101).

areas displaying a histologic continuum in one of the tumors. Recent ploidy studies found that seminomas have a higher DNA index than nonseminomatous germ cell tumors (1.66 and 1.43, respectively),3 suggesting that nonseminomatous tumors may develop from seminomas as a consequence of chromosome loss—a concept consistent with current theories concerning the role of cancer suppressor genes in malignant neoplasia.28 Our observation that the seminoma-yolk sac tumor transformation was abrupt is consistent with this theory. In addition, HLA studies suggest that seminomas "exist as a stage after in situ carcinoma through which all germ cell tumors progress".6 Furthermore, intratubular germ cell neoplasia, the acknowledged common precursor to virtually all forms of testicular germ cell tumors, shows a striking morphologic, ultrastructural, and immunohistochemical resemblance to seminomatous cells,29'30 an observation that

supports a precursor role for seminoma. Differentiation to nonseminomatous elements presumably may then develop from either an invasive seminoma or from intratubular germ cell neoplasia while still confined within the tubules. 4,3 ' Other observations that support the ability of seminoma to differentiate to other germ cell tumor types include (1) its frequent combination with nonseminomatous elements within mixed germ cell tumors 9 "; (2) its frequent admixture with syncytiotrophoblasts,32"34 and the presence of trophoblastic hormones within seminomalike cells lacking distinct syncytiotrophoblastic differentiation by light microscopic examination4,32; and (3) the occurrence of elevated serum AFP levels in patients with metastatic germ cell tumors who had pure seminoma in well-sampled orchiectomy specimens.35"37 Although nonsampled yolk sac tumor elements in the testis cannot be excluded definitely in such instances, the differentiation

Vol. 97 • No. 4

Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016

TO,",

474

ANATOMIC PATHOLOGY Original Article

'.*• -•

', '» « . •-" '

''•,

» '

.

"

*

..'•'.

*•



'

'

.

i '

-

- .



'

*

'

,

"

_ '.'•."•»*'•*,"

***?»**



' "

' . '

•.

Evidence for the transformation of seminoma to yolk sac tumor, with histogenetic considerations.

Recent ultrastructural, cytogenetic, and ploidy analyses indicate that seminoma acts as a precursor from which other forms of testicular germ cell tum...
10MB Sizes 0 Downloads 0 Views