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Current concepts in the diagnosis of human soft tissue sarcomas
Perspectives in Pathology C U R R E N T C O N C E P T S IN T H E D I A G N O S I S OF H U M A N S O F T TISSUE SARCOMAS Robert Stern, M.D.*
Abstract Studies of tile stroma of human soft tissue sarcomas may provide a new classification scheme for these tumors. Immunoflttorescence microscopy using antisera directed against purified stromal components, particttlarly against type specific collagens, may become an important adjunct for the surgical pathologist in the diagnosis o f these tumors.
THE CURRENT CLASSIFICATION OF SOFT TISSUE TUMORS H u m a n sarcomas, particularly the soft tissue tumors, have long presented problems in diagnosis. Soft tissue neoplasms constitute a heterogeneous group of proliferative lesions purportedly arising from mesenchymal cells. These tumors vary significantly in histological structure and location o f origin, and in fact considerable controversy often surrounds the question o f their cell o f origin. A particular problem is that the biological activity of these diverse neoplasm varies, from benign localized growths to invasive, highly malignant entities9 These are also important tumors. It is estimated that one-fourth of all the solid tumors in children arise in the soft tissues.' Our ability to separate the different biological entities has been predicated on the basis o f a variety o f m o r p h o l o g i c a l a p p e a r a n c e s . G e n e r a l l y the classification schemes currently used are those o f Stout and Lattes ~ and Enzinger. s These schemes are remarkable in their diversity, the frequency with which eponyms are used, and their lack o f consistency. This may underly the particular inability to predict biological activity simply on the basis of an observed lnstologmal pattern 9 All too frequently, once a term has been applied to a lesion, on the basis of a particular cell pattern, the surgical pathologist feels that his obligations have ended. Determination of what cell has been transformed and what process has resulted in this histo9
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logical pattern should also be in the purview of the pathologist. Facility in applying a label to a surgical specimen often shortcircuits the ability to pose these important questions. Significant advances in both understanding proliferative lesions and predicting biological activities were occasioned with extensions of our means o f examination 9 Electron microscopic observation made more particular and more critical our ability to differentiate specific tumors. More recently the utilization o f a variety o f immunocytochemical techniques to s e p a r a t e d i f f e r e n t types o f l y m p h o m a s a n d leukemias has aided us in predicting biological activity and in selecting appropriate therapy for these particular tumors. It stands to reason that a similar biochemical approach might help in separating the several types of soft tissue growths. There should be no more reluctance on the part of the pathologist to accept and utilize these approaches than ther~ was in accepting tile extensions o f microscopy. The following discussion details the potential benefits to be gained from use of this system9 DIFFICULTY W I T H CURRENT CLASSIFICATION SCHEMES It has been our experience that cases o f many soft tissue tumors referred to our hospital have a litany o f diagnoses attached to them as the slides are passed t h r o u g h th~ h a n d s o f several surgical
Accepted for publication April 20, 1981. This study was supported by grant 1 RO1 CA 2517-01A1 PTHB awarded by the National Cancer Institute, DHHS. * Associate Professor, Departmentof Pathology,Universityof California,San Francisco,Schoolof Medicine.Attending Pathologist, Universityof California Hospitals, Universityof California, San Francisco,San Francisco,California. HUMAN PATHOLOGY--VOLUME 12, NUMBER9 September 1981
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HUMAN PATHOLOGY--VOLUME 12, NUMBER 9 September1981 p a t h o l o g i s t s . ~I'erms s u c h as s y n o v i a l s a r c o m a , fibroxanthosarcoma, epithelioid sarcoma, malignant fibrous histiocytoma, and alveolar soft part sarcoma o r e p o n y m s such as Ewing's s a r c o m a r e f e r to a constellation o f histological features, some highly variable. T h e s e terms do not r e f e r to the biology o f the t u m o r in any sense. One b r i e f study p r o v i d e d evidence o f the problems intrinsic to this area o f surgical pathology. T h e clinical records from all 11 patients with extraskeletal Ewing's sarcoma at tile National Cancer Institute between 1968 and 1976 were reviewed. T h e diagnoses p r o f e r r e d by the r e f e r r i n g pathologist f r o m outside hospitals are given in Table 1. T h e p r o b l e m in recognizing this soft tissue t u m o r is readily a p p a r e n t f r o m the disparate diagnoses r e n d e r e d . C u r r e n t classification schemes separate t u m o r s whose cell o f origin is known from those whose origin is not known. However, a m o n g the t u m o r s whose n o r m a l c o u n t e r p a r t s are known, an opinion regarding the assumed tissue o f origin may be based on a m i n i m u m o f evidence. Evidence that a synovial sarc o m a actually arises f r o m n o r m a l synovium is tdnuous at best. This lesion often arises in muscle far r e m o v e d f r o m any synovial space. Also confusing are those t u m o r s in which the n o r m a l tissue o f origin is known with s o m e c e r t a i n t y , b u t t h e t u m o r arises in an a n o m a l o u s location. Extraskeletal bone t u m o r s such as the soft tissue o s t e o g e n i c s a r c o m a s , c h o n d r o sarcomas, and Ewing's sarcomas and the o c c u r r e n c e o f r h a b d o m y o s a r c o m a s in the e x t r a h e p a t i c biliary tree are classic e x a m p l e s ? - 9 9T h e o t h e r major g r o u p o f soft tissue sarcomas are those whose histogenesis or normal nont r a n s f o r m e d c o u n t e r p a r t s are not known. Identification o f those tumors is d e p e n d e n t entirely on pattern r e c o g n i t i o n . Despite this general schema, 15 to 20 per cent o f the tumors defy diagnosis and remain unclassified. T h e gross a p p e a r a n c e o f these lesions is usually not helpful in diagnosis. It is often difficult to distinguish soft tissue masses that are benign f r o m those that are malignant. T h e f r e q u e n c y with which sarcomas are mistaken for benign processes is the result chiefly o f the deceptively harmless a p p e a r a n c e during the initial phase o f the disease. However, it is obvious that correct diagnosis and vigorous t h e r a p y at an early stage are imperative for cure. For the surgical p a t h o l o g i s t the c o n v e r s e may be even m o r e o f a p r o b l e m . N o d u l a r fasciitis can be easily mistaken for a sarcoma. Many malignant t u m o r s can a p p e a r e~capsulated grossly. Since sarcomas are not associated with a basement m e m b r a n e t h r o u g h which they must p e n e t r a t e to be c o n s i d e r e d invasive, they e x p a n d until the pressure o f adjacent tissue confines them. T h i s pressure may p r o d u c e a pseudocapsule, with a misleading gross a p p e a r a n c e o f being well encapsulated, but these lesions can be seen mlcroscopicall) to be infiltrative. Such t u m o r s r e c n r after enucleat i o n ? ~ T h i s occurs f r e q u e n t l y with fibrosarcomas, liposarcomas, leiomyosarcomas, and synovial sarcomas.
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T A B L E 1. D I A G N O S E S PROFFERED BY REFERRING PATHOLOGISTS Undifferentiated small cell sarcoma versus carcinoma Alveolar rhabdomyosarcoma Sarcoma, type unspecified or undifferentiated Malignant histiocytoma Reticulum cell sarcoma Synovial sarcoma Glomus tumor Embryonal rhabdomyosarcoma Undifferentiated malignant tumor Soft tissue tumor resembling Ewing's sarcoma Ewing's sarcoma
Soft tissue sarcomas can arise in a n u m b e r o f l o c a t i o n s - - m e d i a s t i n u m , r e t r o p e r i t o n e u m , the soft tissues o f tile visceral organs, and within muscle and connective tissues o f the extremities. However, the site o f origin is not part o f the classification scheme, and it is not usually helpful in making a diagnosis in an e f f o r t to u n d e r s t a n d their behavior. An additional disadvantage o f the c u r r e n t classification scheme is that the d e g r e e o f differentiation o f any single lesion is usually not relevant to the diagnosis. Some t u m o r s are so u n d i f f e r e n t i a t e d that they begin to resemble each o t h e r as their histological patterns a p p r o a c h that o f primitive mesenchyme. T h e c o n t r i b u t i o n o f the host r e s p o n s e to the malignant cells has also not been adequately evaluated in these lesions. J u s t as a scirrhous response o r desmoplastic reaction occurs in epithelial malignant tumors, there may also be a host response to malignant sarcomas. I n d e e d m u c h o f the dense fibrous stroma o f some sarcomas may be o f host origin. T h e b r o a d bands o f connective tissue that traverse the l y m p h n o d e in tile n o d u l a r s c l e r o s i n g f o r m o f Hodgkin's disease is o f particular interest in this regard. This may be the best known example o f a scirr h o u s r e a c t i o n by t h e host to m a l i g n a n t m e s e n chymal cells. O t h e r examples occur, but to date they have not been identified a m o n g o t h e r h u m a n sarcomas. O t h e r complications in the d i a g n o s i s p f soft tissue sarcomas include necrosis and ulceration. T h e s e att e n d a n t complications interfere m o r e frequently in the accurate diagnosis o f sarcomas than o f epithelial neoplasms. Ulceration may be the presenting sign o f a soft tissue t u m o r , p a r t i c u l a r l y o f an e x t r e m i t y . Epithelioid s a r c o m a is a relatively new entity not c o n t a i n e d in t h e a f o r e m e n t i o n e d diagnostic schemes. II This t u m o r can begin as an ulcerated lesion and may be i g n o r e d . A t may be confused with a g r a n u l o m a t o u s process, o r it may look like an ulcerated squamous cell carcinoma, a mistake m a d e all too frequently. L Occasionally an anaplastic u n d i f f e r e n t i a t e d small cell t u m o r presents a particularly difficult p r o b l e m for the surgical patholog!st. One example is the undifferentiated small cell osteogenic sarcoma, whose pattern may resemble that o f Ewing's sarcoma, t2 This d i s t i n c t i o n is o f clinical i m p o r t a n c e b e c a u s e the
HUMAN SOFT TISSUE SARCOMAS---STERN treatments o f these two lesions d i f f e r so widely. In the initial biopsy study a decision must be m a d e by the surgical pathologist based o n an exceedingly small a m o t m t o f tissue. U n f o r t u n a t e l y such e r r o r s b e c o m e obvious only when the ample tissue o f the a m p u t a t i o n specimen becomes available. Angiograms, l y m p h a n g i o g r a m s , c o m p u t e d tom o g r a p h i c scans, sonograms, and u l t r a s o u n d are also not helpfld in making an accurate diagnosis o f soft tissue tumors. T h e y help the s u r g e o n to develop an effective t r e a t m e n t plan, but such data are not relevant for c u r r e n t diagnostic schemes. It has become a p p a r e n t that megavolt irradiation for Hodgkin's disease and o t h e r t u m o r s may be associated with the d e v e l o p m e n t o f a second t u m o r , usually a soft tissue sarcoma, with an onset 20 to 30 years later. Such cases are only now c o m i n g to the attention o f surgical pathologists. 13 T h e accurate diagnosis o f h u m a n sarcomas will become increasingly i m p o r t a n t in the future. Every a t t e m p t must be m a d e t o b e g i n to u n d e r s t a n d these lesions. What is required is a conceptual f r a m e w o r k for the examination o f t h e s e lesions, an all e n c o m p a s s i n g classification scheme, which recognizes the biology o f these tumors. Only then will an u n d e r s t a n d i n g o f prognosis and d e v e l o p m e n t o f an a p p r o p r i a t e t r e a t m e n t regimen follow logically.
R E L A T I O N OF T U M O R S T O T H E NORMAL MESENCHYME
A n o b s e r v a t i o n b e c o m i n g i n c r e a s i n g l y well d o c u m e n t e d is that h u m a n t u m o r s reflect with great a c c u r a c y a n d fastidiousness c e r t a i n stages in the pathway o f differentiation o f n o r m a l cells. This has been shown by surface markers with particular clarity for h u m a n l y m p h o m a s and leukemias? 4 It is as if a b a r r i e r to f u r t h e r cell differentiation had been imposed at a particular stage in that process. D e f n i n g h u m a n sarcomas m o r e clearly will lead to o u r ability to resolve d i f f e r e n t t u m o r s for diagnostic purposes, and will also give us insight into the stages o f differentiation o f normal mesenchyme. T h e s e bland app e a r i n g lethal tumors, with their characteristic banal histologic m o n o t o n y , may provide i m p o r t a n t clues to the rich diversity o f the n o r m a l mesenchyme. For e x a m p l e , a working hypothesis for the cell o f origin o f the malignant fibrous histiocytomas would be an und i f f e r e n t i a t e d mesenchymal cell, p r e c u r s o r to both f b r o b l a s t s and histiocytes. It is this cell that may have u n d e r g o n e the malignant t r a n s f o r m a t i o n event, a cell that has h e r e t o f o r e escaped detection in histological sections o f normal tissues. A p p r o x i m a t e l y 25 p e r c e n t o f f i b r o s a r c o m a s o c c u r in scars. I t would be o f great i m p o r t a n c e to identify that cell at a particular stage o f w o u n d healing that is susceptibl e tq malignant transformation. T h e p l u r i p o t e n t i a l m e s e n c h y m a l cells, which must a b o u n d in the body, for the most part have a cryptic or transient existence. Only when they
u n d e r g o malignant transformation do they b e c o m e evident. T h e delicate periosteal layer o f bone's transf o r m a t i o n to a periosteal osteosarcoma is one example. We know so little a b o u t some o f these n o r m a l tissues that we often do not recognize the t u m o r s that originate from them. T h u s soft tissue t u m o r s are replete with biological and embryological data, although we are only now beginning to be receptive to the information these t u m o r s can provide. This is also an e x a m p l e o f how clinical material can provide basic biological information. We should not u n d e r e s t i m a t e the h u m a n material with which we work each day as a source o f knowledge. T h e diversity o f h u m a n t u m o r s a p p e a r s to be g r e a t e r than that o f any o t h e r animal. T u m o r s p a r a p h r a s e and are caricatures o f the stages o f differentiation o f n o r m a l tissue, and may shed great insight into that process. T h u s studies o f h u m a n soft tissue tumors not only may help us to differentiate difficult lesions one f r o m the other, but may help us to identify the n o r m a l cell o f origin o f these tumors, the biology, and the embryological histogenesis o f normal cells. It may be possible to apply this concept to a new diagnostic scheme for h u m a n s a r c o m a s - - t h a t each sarcoma represents with great accuracy a stage in the pathway o f differentiation o f a normal mesenchymal cell. B I O L O G Y OF T H E T U M O R M A T R I X
T h e surgical pathologist's diagnosis sets the stage for the t r e a t m e n t and care o f all cancer patients. An entirely new conceptual f r a m e w o r k is r e q u i r e d if we are to deal accurately and effectively with soft tissue sarcomas. In the past d e c a d e there has been rapid progress in molecular biology and in o u r u n d e r s t a n d ing o f tile chemistry o f structural proteins such as elastin, myosin, keratin, a n d in particular the collagen molecule. This i n f o r m a t i o n , t o g e t h e r with the recent p r o g r e s s in m o n o c l o n a l a n t i b o d y p r o d u c t i o n a n d i m m u n o f l u o r e s c e n c e t e c h n i q u e s , can resolve the controversy over the structural c o m p o n e n t s o f t u m o r matrices. Identification o f tile structural elements in tile matrix o f h u m a n sarcomas will provid~ a diagnostic scheme based on d e f i n e d biochemical criteria. Such a classification scheme will use both m o r p h o l o g y and biochemical markers, in m u c h the same m a n n e r used with great success in resolving B cell, T cell, and null cell l y m p h o m a s and their attendant subsets. A n u m b e r o f e x p e r i m e n t a l transplantable animal sarcomas exist whose stromal collagens have been well defined. T M Studies o f these tumors may act as prototypes for the systematic i~xamination o f h u m a n sarcomas. Some o f these sarcomas in e x p e r i m e n t a l animals histologically do not resemble their nontransf o r m e d counterparts. However, analysis o f the collagenous c o m p o n e n t s o f their stroma confirms their p r e s u m e d cell o f origin. O n e example is a mouse t u m o r o f known e n d o d e r m a l sinus origin that elaborates a matrix o f p u r e basement m e m b r a n e . 17 It is intriguing to postulate that groups o f h u m a n sar-
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HUMAN PATIIOLOGY--VOLUME 12, NUMBER 9 September1981 c o m a s exist with lfistological features r e s e m b l i n g no k n o w n n o r m a l tissue, yet whose n o n m a l i g n a n t count e r p a r t will b e c o m e evident a f t e r biochemical analysis o f their matrix proteins. Evidence has b e e n d o c u m e n t e d that distinctions o f stromal collagens do exist in h u m a n sarcomas and can be used for diagnostic p u r p o s e s ? s T h e profile o f collagen types in a case o f m a l i g n a n t fibrous histoc y t o m a has been e x a m i n e d r e c e n t l y ? 9 T h e evidence suggests that t h e r e is a r e s e m b l a n c e b e t w e e n tlfis t u m o r a n d osteosarcomas. It is i m p o r t a n t to point o u t that t u m o r m a r k e r s a n d such a classification s c h e m e will not replace the skills o f the surgical pathologist but will b r i n g the tools o f the biochemist to the surgical pathology bench. BIOCHEMISTRY OF COLLAGEN THE TUMOR MATRIX
AND
C o l l a g e n is the m a j o r c o n n e c t i v e tissue p r o i e i n - - t b e c h i e f c o m p o n e n t o f tile t u m o r m a t r i x as well as the most a b u n d a n t p r o t e i n o f the body. It is now recognized to be a family o f closely related proreins, each a d i f f e r e n t gene p r o d u c t . * Five types o f collagen are now recognized (Table 2). A m i n o acid s e q u e n c e s t u d i e s a n d c y a n o g e n b r o m i d e a n d e n z y m e p e p t i d e m a p p i n g analyses o f n o r m a l tissne reveal even g r e a t e r h e t e r o g e n e i t y t h a n that disclosed by conventional protein c h r o m a t o g r a p h y . M i n o r collagen types h a v e b e e n d e t e c t e d in n o r m a l tissues whose identity a n d relationship to the m a j o r collagen types have not yet b e e n established, z4 T h e same a p p r o a c h can now be a p p l i e d to the matrix o f h u m a n and e x p e r i m e n t a l sarcomas. Unique collagens m a y exist in the matrices o f t u m o r s that are also d i f f e r e n t f r o m those described h e r e t o f o r e . T h e re-
* For recent reviews see references 20 to 22. TABLE 2.
Type I
II III
IV V
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TYPES OF COLLAGEN
Location This is the most abundant type, making up the major protein component of skin, bone, and tendon. However it is ubiquitous in the body. A variant is found in fetal tissue and in some experimental tumors.23 This is cartilage-specific collagen, found also in the nucleu~ po]posus and in certain eye structures. This collagen is a form of reticulin though the relationship between the two is unclear. Type III is present wherever type I is found, but in decreased amounts in hard tissue such as bone and dentin, and in increased amounts proportionately in flexible distensible tissues such as the gastrointestinal tract and blood vessels. It is also found in rapidly proliferating tissue, in fetal tissues, in bone marrow, and in early wound healing and in early cirrhosis.. Type IV collagen"is basement membrane specific. This is a widespread molecule suspected to be a cell surface collagen or a collagen associated with the plasma membrane of cells.
cent pro.gress in t e c h n i q u e makes it possible to exa m i n e m t c r o g r a m quantities for structural identification. 24 Evidence o f the c o m p a r a b l e h e t e r o g e n e i t y o f e l a s tin exists as well as for the tissue specific adhesion proteins c o n t a i n e d within tissue matrix such as fibronectin, c h o n d r o n e c t i n , a n d laminin. 2~-29 T h e r e is also a b a t t e r y o f m a t r i x g l y c o p r o t e i n s ? ~ This p a n o p l y o f extracellular structural m a c r o m o l e c u l e s p o r t e n d s well for the eventual identification o f t u m o r specific matrix p r o t e i n s a n d t u m o r specific p r o t e i n products. T h e h y b r i d o m a t e c h n i q u e makes it possible to g e n e r a t e with relative ease large a m o u n t s o f m o n o clonal antibodies directed against each o f these struct u r a l m o i e t i e s a n d v i r t u a l l y all c o n n e c t i v e tissue macromolecules? z Monoclonal antibodies directed against type I, II, a n d IV collagen have b e e n reported, a3-3~ and it is within reason to a s s u m e that a battery o f such antibodies m i g h t soon be available for use in surgical p a t h o l o g y f o r i m m u n o f l u o r e s c e n c e staining o f frozen sections o f t u m o r s . S o m e o f the variants o f soft tissue s a r c o m a s are sufficiently rare that e v e n histopathologists in large medical centers are p r e s e n t e d with serious p r o b l e m s . T h e r e are no centers large e n o u g h for o n e individual to see e n o u g h e x a m p l e s o f the variants to be familiar with the a p p e a r a n c e o f all o f t h e m . An a r r a y o f antibodies for use in e x a m i n i n g specific stromal prot e i n s will a u g m e n t t h e a b i l i t y o f t h e s u r g i c a l pathologist to m a k e such diagnoses. Not only t u m o r specific structural m a t r i x proteins but u n i q u e patterns o f organization o f conventional matrix proteins m u s t be c o n s i d e r e d in this context as diagnostic aids. It is i n c u m b e n t o n surgical pathologists to c o o p e r a t e with biochemists at s o m e stage o f this dialogue in o r d e r to d e m o n s t r a t e the u s e f u l n e s s o f these reagents. T h e r e is a m a n d a t e a d d r e s s e d to all clinicians to translate recent a d v a n c e s in biology m o r e rapidly a n d m o r e effectively into the i m p r o v e m e n t o f medical care. A liaison b e t w e e n biochemists who study structural proteins a n d surgical pathologists m a y e x p e d i t e this process. Surgical p a t h o l o g y is at arf i m p o r t a n t j u n c t u r e ~ t h e o p p o r t u n i t y to b r i n g the r e c e n t advances in biology to b e a r on tile i m m e d i a t e care a n d t r e a t m e n t o f certain c a n c e r patients.
SUMMARY
- T h e surgical pathologist is often called u p o n to m a k e a r a p i d decision aboiat the m a l i g n a n t potential o f a soft tissue t u m o r . T h e s e t u m o r s have s p e c i f c stromas that s u p p o r t g r o w t h and e n d o w the t u m o r with u n i q u e quali~ies, as well as being manifestations o f the p h e n o t y p i c e x p r e s s i o n o f t u m o r cells. Collagen, the m a j o r p r o t e i n o f the s t r o m a o f t u m o r s , is now r e c o g n i z e d as a family o f proteins, each a d i f f e r e n t gene product. Immunofluorescence microscopy studies u s i n g a n t i b o d i e s d i r e c t e d a g a i n s t p u r i f i e d s t r o m a l c o m p o n e n t s , particularly the d i f f e r e n t type
H U M A N S O F T T I S S U E SARCOMAS---STERN s p e c i f i c c o l l a g e n s , m a y s o o n b e a v a i l a b l e in l a r g e q u a n t i t i e s . T h e s e a r e p o t e n t i a l l y u s e f u l tools f o r t h e surgical pathologist not only in m a k i n g that i m p o r t a n t d e c i s i o n , b u t also f o r d e v e l o p i n g a classification s c h e m e f o r h u m a n s a r c o m a s . T h i s s c h e m e will i n e v i t a b l y h a v e a m o r e s o u n d biological basis t h a n t h o s e currently used.
ACKNOWLEDGMENT I am indebted to Diane Jarest for her skillfld help in preparing tiffs manuscript.
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16. Breitkreutz, D., Diaz de Leon, L., Paglia, L., Gay, S., Swarm, R. L., and Stern, R.: Histological and biochemical studies of a transplantable rat chondrosarcoma. Cancer Res., 39:5093-5100, 1979. 17. Orkin, R. W., Gehron, P., McGoodwin, E. M., Martin, G. R., Valentine, T., and Swarm, R. L.: A murine tumor producing a matrix of basement membrane. J. Exp. Med., 145:204-220, 1977. 18. Stern, R., Wilczek, J., Thorpe, W. P., Rosenberg, S. A., and Cannon, G.: Procollagens as markers for the cell of origin of human bone tumors. Cancer Res., 40:325-328, 1980. 19. Hall, J , Tseng, S. C. G., Hendrix, M.J.C., Linsenmayer, T., Timple, R., and Stern, R.: The collagen profiles of human sarcomas. Malignant fibrous histiocytoma. Submitted for publication. 20. Fessler, J. H., and Fessler, L. I.: Biosynthesis of procollagen. Ann. Rev. Biochem., 47:129-162, 1978. 21. Prockop, D.J., Kivirikko, K. I., Tuderman, L., and Guzman, N. A.: Biosynthesis of collagen and its disorders. New Engl. J. Med., 301:13-85, 1979. 22. Bornstein, P., and Sage, H.: Structurally distinct collagen types. Ann. Rev. Biochem., 49:957-1003, 1980. 23. Mayne, R., Vails, M. S., and Miller, E. J.: Analysis of changes in collagen biosynthesis that occur when chick chondrocytes are grown in 5-bromo-2'-deoxyuridine. Proc. Natl. Acad. Sci. USA, 72:4511-4515, 1975. 24. Sage, H., Pritzl, P., and Bornstein, P.: A new mapping technique for collagen chains. Coll. Rel. Res., 1:3-15, 1981. 25. Quintarelli, G., Starcher, B. C., Vocaturo, A., Di Gianfilippo, F., Gotte, L., and Mecham, R. P.: Fibrogenesis and biosynthesis of elastin in cartilage. Connect. Tissue Res., 7:1-19, 1979. 26. Sandberg, L.B., Gray, W.R., and Franzblau, C. (Editors): Elastin and Elastic Tissue. New York, Plenum Press, 1977. 27. Ruoslahti, E., Engvall, E., and Hayman, E.G.: Fibronectin: current concepts of its structure and functions. Collagen Rel. Res., 1:95-128, 1981. 28. Hewitt, A. T., Kleinman, H. K., Pennypacker, J. P., and Martin, G. R.: Identification of an adhesion factor for chondrocytes. Proc. Natl. Acad. Sci. USA, 77:385-388, 1980. 29. Timple, R., Rohde, H., Robey, P. G., Rennard, S. I., Foidart, J.-M., and Martin, G. R.: Laminin, a glycoprotein from basement membranes. J. Biol. Chem., 254:9933=-9937, 1979. 30. Anderson,J. C.: Glycoproteinsoftheconnectivetissuematrix. Int. Rev. Connect. Tissue Res., 7:251-322, 1976. 31. Kornfeld, R., and Kornfeld, S.: Comparative aspects ofglycoprotein structure. Ann. Rev. Biochem., 45:217-237, 1976. 32. Milstein, C., and Lennox, E.: The use of monoclonal antibody techniques in the study of developing cell surfaces. Current Top. Develop. Biol., 14(Part II):1-32, 1980. 33. Linsenmayer, T. F., Hendrix, M.J.C., and Little, C. D.: Production and characterization of a monoclonal antibody to chick type I collagen. Proc. Natl. Acad. Sci. USA, 76:3703-3707, 1979. 34. Linsenmayer, T. F., and Hendrix, M.J.C.: Monoclonal antibodies to connective tissue macromolecules: type II collagen. Biochem. Biophys. Res. Comm., 92:440-446, 1980. 35. Foellmer, H., Madri, J. A., and Furthmayr, H.: Monoclonal antibodies to type IV collagen from human placenta and bovine lung. Fed. Proc., 40:794, 1981. Department of Pathology University of California Hospitals University.9f California, San Francisco San Francisco, Calif6rnia 94143
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Abstract Studies of tile stroma of human soft tissue sarcomas may provide a new classification scheme for these tumors. Immunoflttorescence microscopy using antisera directed against purified stromal components, particttlarly against type specific collagens, may become an important adjunct for the surgical pathologist in the diagnosis o f these tumors.
THE CURRENT CLASSIFICATION OF SOFT TISSUE TUMORS H u m a n sarcomas, particularly the soft tissue tumors, have long presented problems in diagnosis. Soft tissue neoplasms constitute a heterogeneous group of proliferative lesions purportedly arising from mesenchymal cells. These tumors vary significantly in histological structure and location o f origin, and in fact considerable controversy often surrounds the question o f their cell o f origin. A particular problem is that the biological activity of these diverse neoplasm varies, from benign localized growths to invasive, highly malignant entities9 These are also important tumors. It is estimated that one-fourth of all the solid tumors in children arise in the soft tissues.' Our ability to separate the different biological entities has been predicated on the basis o f a variety o f m o r p h o l o g i c a l a p p e a r a n c e s . G e n e r a l l y the classification schemes currently used are those o f Stout and Lattes ~ and Enzinger. s These schemes are remarkable in their diversity, the frequency with which eponyms are used, and their lack o f consistency. This may underly the particular inability to predict biological activity simply on the basis of an observed lnstologmal pattern 9 All too frequently, once a term has been applied to a lesion, on the basis of a particular cell pattern, the surgical pathologist feels that his obligations have ended. Determination of what cell has been transformed and what process has resulted in this histo9
i
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logical pattern should also be in the purview of the pathologist. Facility in applying a label to a surgical specimen often shortcircuits the ability to pose these important questions. Significant advances in both understanding proliferative lesions and predicting biological activities were occasioned with extensions of our means o f examination 9 Electron microscopic observation made more particular and more critical our ability to differentiate specific tumors. More recently the utilization o f a variety o f immunocytochemical techniques to s e p a r a t e d i f f e r e n t types o f l y m p h o m a s a n d leukemias has aided us in predicting biological activity and in selecting appropriate therapy for these particular tumors. It stands to reason that a similar biochemical approach might help in separating the several types of soft tissue growths. There should be no more reluctance on the part of the pathologist to accept and utilize these approaches than ther~ was in accepting tile extensions o f microscopy. The following discussion details the potential benefits to be gained from use of this system9 DIFFICULTY W I T H CURRENT CLASSIFICATION SCHEMES It has been our experience that cases o f many soft tissue tumors referred to our hospital have a litany o f diagnoses attached to them as the slides are passed t h r o u g h th~ h a n d s o f several surgical
Accepted for publication April 20, 1981. This study was supported by grant 1 RO1 CA 2517-01A1 PTHB awarded by the National Cancer Institute, DHHS. * Associate Professor, Departmentof Pathology,Universityof California,San Francisco,Schoolof Medicine.Attending Pathologist, Universityof California Hospitals, Universityof California, San Francisco,San Francisco,California. HUMAN PATHOLOGY--VOLUME 12, NUMBER9 September 1981
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HUMAN PATHOLOGY--VOLUME 12, NUMBER 9 September1981 p a t h o l o g i s t s . ~I'erms s u c h as s y n o v i a l s a r c o m a , fibroxanthosarcoma, epithelioid sarcoma, malignant fibrous histiocytoma, and alveolar soft part sarcoma o r e p o n y m s such as Ewing's s a r c o m a r e f e r to a constellation o f histological features, some highly variable. T h e s e terms do not r e f e r to the biology o f the t u m o r in any sense. One b r i e f study p r o v i d e d evidence o f the problems intrinsic to this area o f surgical pathology. T h e clinical records from all 11 patients with extraskeletal Ewing's sarcoma at tile National Cancer Institute between 1968 and 1976 were reviewed. T h e diagnoses p r o f e r r e d by the r e f e r r i n g pathologist f r o m outside hospitals are given in Table 1. T h e p r o b l e m in recognizing this soft tissue t u m o r is readily a p p a r e n t f r o m the disparate diagnoses r e n d e r e d . C u r r e n t classification schemes separate t u m o r s whose cell o f origin is known from those whose origin is not known. However, a m o n g the t u m o r s whose n o r m a l c o u n t e r p a r t s are known, an opinion regarding the assumed tissue o f origin may be based on a m i n i m u m o f evidence. Evidence that a synovial sarc o m a actually arises f r o m n o r m a l synovium is tdnuous at best. This lesion often arises in muscle far r e m o v e d f r o m any synovial space. Also confusing are those t u m o r s in which the n o r m a l tissue o f origin is known with s o m e c e r t a i n t y , b u t t h e t u m o r arises in an a n o m a l o u s location. Extraskeletal bone t u m o r s such as the soft tissue o s t e o g e n i c s a r c o m a s , c h o n d r o sarcomas, and Ewing's sarcomas and the o c c u r r e n c e o f r h a b d o m y o s a r c o m a s in the e x t r a h e p a t i c biliary tree are classic e x a m p l e s ? - 9 9T h e o t h e r major g r o u p o f soft tissue sarcomas are those whose histogenesis or normal nont r a n s f o r m e d c o u n t e r p a r t s are not known. Identification o f those tumors is d e p e n d e n t entirely on pattern r e c o g n i t i o n . Despite this general schema, 15 to 20 per cent o f the tumors defy diagnosis and remain unclassified. T h e gross a p p e a r a n c e o f these lesions is usually not helpful in diagnosis. It is often difficult to distinguish soft tissue masses that are benign f r o m those that are malignant. T h e f r e q u e n c y with which sarcomas are mistaken for benign processes is the result chiefly o f the deceptively harmless a p p e a r a n c e during the initial phase o f the disease. However, it is obvious that correct diagnosis and vigorous t h e r a p y at an early stage are imperative for cure. For the surgical p a t h o l o g i s t the c o n v e r s e may be even m o r e o f a p r o b l e m . N o d u l a r fasciitis can be easily mistaken for a sarcoma. Many malignant t u m o r s can a p p e a r e~capsulated grossly. Since sarcomas are not associated with a basement m e m b r a n e t h r o u g h which they must p e n e t r a t e to be c o n s i d e r e d invasive, they e x p a n d until the pressure o f adjacent tissue confines them. T h i s pressure may p r o d u c e a pseudocapsule, with a misleading gross a p p e a r a n c e o f being well encapsulated, but these lesions can be seen mlcroscopicall) to be infiltrative. Such t u m o r s r e c n r after enucleat i o n ? ~ T h i s occurs f r e q u e n t l y with fibrosarcomas, liposarcomas, leiomyosarcomas, and synovial sarcomas.
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T A B L E 1. D I A G N O S E S PROFFERED BY REFERRING PATHOLOGISTS Undifferentiated small cell sarcoma versus carcinoma Alveolar rhabdomyosarcoma Sarcoma, type unspecified or undifferentiated Malignant histiocytoma Reticulum cell sarcoma Synovial sarcoma Glomus tumor Embryonal rhabdomyosarcoma Undifferentiated malignant tumor Soft tissue tumor resembling Ewing's sarcoma Ewing's sarcoma
Soft tissue sarcomas can arise in a n u m b e r o f l o c a t i o n s - - m e d i a s t i n u m , r e t r o p e r i t o n e u m , the soft tissues o f tile visceral organs, and within muscle and connective tissues o f the extremities. However, the site o f origin is not part o f the classification scheme, and it is not usually helpful in making a diagnosis in an e f f o r t to u n d e r s t a n d their behavior. An additional disadvantage o f the c u r r e n t classification scheme is that the d e g r e e o f differentiation o f any single lesion is usually not relevant to the diagnosis. Some t u m o r s are so u n d i f f e r e n t i a t e d that they begin to resemble each o t h e r as their histological patterns a p p r o a c h that o f primitive mesenchyme. T h e c o n t r i b u t i o n o f the host r e s p o n s e to the malignant cells has also not been adequately evaluated in these lesions. J u s t as a scirrhous response o r desmoplastic reaction occurs in epithelial malignant tumors, there may also be a host response to malignant sarcomas. I n d e e d m u c h o f the dense fibrous stroma o f some sarcomas may be o f host origin. T h e b r o a d bands o f connective tissue that traverse the l y m p h n o d e in tile n o d u l a r s c l e r o s i n g f o r m o f Hodgkin's disease is o f particular interest in this regard. This may be the best known example o f a scirr h o u s r e a c t i o n by t h e host to m a l i g n a n t m e s e n chymal cells. O t h e r examples occur, but to date they have not been identified a m o n g o t h e r h u m a n sarcomas. O t h e r complications in the d i a g n o s i s p f soft tissue sarcomas include necrosis and ulceration. T h e s e att e n d a n t complications interfere m o r e frequently in the accurate diagnosis o f sarcomas than o f epithelial neoplasms. Ulceration may be the presenting sign o f a soft tissue t u m o r , p a r t i c u l a r l y o f an e x t r e m i t y . Epithelioid s a r c o m a is a relatively new entity not c o n t a i n e d in t h e a f o r e m e n t i o n e d diagnostic schemes. II This t u m o r can begin as an ulcerated lesion and may be i g n o r e d . A t may be confused with a g r a n u l o m a t o u s process, o r it may look like an ulcerated squamous cell carcinoma, a mistake m a d e all too frequently. L Occasionally an anaplastic u n d i f f e r e n t i a t e d small cell t u m o r presents a particularly difficult p r o b l e m for the surgical patholog!st. One example is the undifferentiated small cell osteogenic sarcoma, whose pattern may resemble that o f Ewing's sarcoma, t2 This d i s t i n c t i o n is o f clinical i m p o r t a n c e b e c a u s e the
HUMAN SOFT TISSUE SARCOMAS---STERN treatments o f these two lesions d i f f e r so widely. In the initial biopsy study a decision must be m a d e by the surgical pathologist based o n an exceedingly small a m o t m t o f tissue. U n f o r t u n a t e l y such e r r o r s b e c o m e obvious only when the ample tissue o f the a m p u t a t i o n specimen becomes available. Angiograms, l y m p h a n g i o g r a m s , c o m p u t e d tom o g r a p h i c scans, sonograms, and u l t r a s o u n d are also not helpfld in making an accurate diagnosis o f soft tissue tumors. T h e y help the s u r g e o n to develop an effective t r e a t m e n t plan, but such data are not relevant for c u r r e n t diagnostic schemes. It has become a p p a r e n t that megavolt irradiation for Hodgkin's disease and o t h e r t u m o r s may be associated with the d e v e l o p m e n t o f a second t u m o r , usually a soft tissue sarcoma, with an onset 20 to 30 years later. Such cases are only now c o m i n g to the attention o f surgical pathologists. 13 T h e accurate diagnosis o f h u m a n sarcomas will become increasingly i m p o r t a n t in the future. Every a t t e m p t must be m a d e t o b e g i n to u n d e r s t a n d these lesions. What is required is a conceptual f r a m e w o r k for the examination o f t h e s e lesions, an all e n c o m p a s s i n g classification scheme, which recognizes the biology o f these tumors. Only then will an u n d e r s t a n d i n g o f prognosis and d e v e l o p m e n t o f an a p p r o p r i a t e t r e a t m e n t regimen follow logically.
R E L A T I O N OF T U M O R S T O T H E NORMAL MESENCHYME
A n o b s e r v a t i o n b e c o m i n g i n c r e a s i n g l y well d o c u m e n t e d is that h u m a n t u m o r s reflect with great a c c u r a c y a n d fastidiousness c e r t a i n stages in the pathway o f differentiation o f n o r m a l cells. This has been shown by surface markers with particular clarity for h u m a n l y m p h o m a s and leukemias? 4 It is as if a b a r r i e r to f u r t h e r cell differentiation had been imposed at a particular stage in that process. D e f n i n g h u m a n sarcomas m o r e clearly will lead to o u r ability to resolve d i f f e r e n t t u m o r s for diagnostic purposes, and will also give us insight into the stages o f differentiation o f normal mesenchyme. T h e s e bland app e a r i n g lethal tumors, with their characteristic banal histologic m o n o t o n y , may provide i m p o r t a n t clues to the rich diversity o f the n o r m a l mesenchyme. For e x a m p l e , a working hypothesis for the cell o f origin o f the malignant fibrous histiocytomas would be an und i f f e r e n t i a t e d mesenchymal cell, p r e c u r s o r to both f b r o b l a s t s and histiocytes. It is this cell that may have u n d e r g o n e the malignant t r a n s f o r m a t i o n event, a cell that has h e r e t o f o r e escaped detection in histological sections o f normal tissues. A p p r o x i m a t e l y 25 p e r c e n t o f f i b r o s a r c o m a s o c c u r in scars. I t would be o f great i m p o r t a n c e to identify that cell at a particular stage o f w o u n d healing that is susceptibl e tq malignant transformation. T h e p l u r i p o t e n t i a l m e s e n c h y m a l cells, which must a b o u n d in the body, for the most part have a cryptic or transient existence. Only when they
u n d e r g o malignant transformation do they b e c o m e evident. T h e delicate periosteal layer o f bone's transf o r m a t i o n to a periosteal osteosarcoma is one example. We know so little a b o u t some o f these n o r m a l tissues that we often do not recognize the t u m o r s that originate from them. T h u s soft tissue t u m o r s are replete with biological and embryological data, although we are only now beginning to be receptive to the information these t u m o r s can provide. This is also an e x a m p l e o f how clinical material can provide basic biological information. We should not u n d e r e s t i m a t e the h u m a n material with which we work each day as a source o f knowledge. T h e diversity o f h u m a n t u m o r s a p p e a r s to be g r e a t e r than that o f any o t h e r animal. T u m o r s p a r a p h r a s e and are caricatures o f the stages o f differentiation o f n o r m a l tissue, and may shed great insight into that process. T h u s studies o f h u m a n soft tissue tumors not only may help us to differentiate difficult lesions one f r o m the other, but may help us to identify the n o r m a l cell o f origin o f these tumors, the biology, and the embryological histogenesis o f normal cells. It may be possible to apply this concept to a new diagnostic scheme for h u m a n s a r c o m a s - - t h a t each sarcoma represents with great accuracy a stage in the pathway o f differentiation o f a normal mesenchymal cell. B I O L O G Y OF T H E T U M O R M A T R I X
T h e surgical pathologist's diagnosis sets the stage for the t r e a t m e n t and care o f all cancer patients. An entirely new conceptual f r a m e w o r k is r e q u i r e d if we are to deal accurately and effectively with soft tissue sarcomas. In the past d e c a d e there has been rapid progress in molecular biology and in o u r u n d e r s t a n d ing o f tile chemistry o f structural proteins such as elastin, myosin, keratin, a n d in particular the collagen molecule. This i n f o r m a t i o n , t o g e t h e r with the recent p r o g r e s s in m o n o c l o n a l a n t i b o d y p r o d u c t i o n a n d i m m u n o f l u o r e s c e n c e t e c h n i q u e s , can resolve the controversy over the structural c o m p o n e n t s o f t u m o r matrices. Identification o f tile structural elements in tile matrix o f h u m a n sarcomas will provid~ a diagnostic scheme based on d e f i n e d biochemical criteria. Such a classification scheme will use both m o r p h o l o g y and biochemical markers, in m u c h the same m a n n e r used with great success in resolving B cell, T cell, and null cell l y m p h o m a s and their attendant subsets. A n u m b e r o f e x p e r i m e n t a l transplantable animal sarcomas exist whose stromal collagens have been well defined. T M Studies o f these tumors may act as prototypes for the systematic i~xamination o f h u m a n sarcomas. Some o f these sarcomas in e x p e r i m e n t a l animals histologically do not resemble their nontransf o r m e d counterparts. However, analysis o f the collagenous c o m p o n e n t s o f their stroma confirms their p r e s u m e d cell o f origin. O n e example is a mouse t u m o r o f known e n d o d e r m a l sinus origin that elaborates a matrix o f p u r e basement m e m b r a n e . 17 It is intriguing to postulate that groups o f h u m a n sar-
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HUMAN PATIIOLOGY--VOLUME 12, NUMBER 9 September1981 c o m a s exist with lfistological features r e s e m b l i n g no k n o w n n o r m a l tissue, yet whose n o n m a l i g n a n t count e r p a r t will b e c o m e evident a f t e r biochemical analysis o f their matrix proteins. Evidence has b e e n d o c u m e n t e d that distinctions o f stromal collagens do exist in h u m a n sarcomas and can be used for diagnostic p u r p o s e s ? s T h e profile o f collagen types in a case o f m a l i g n a n t fibrous histoc y t o m a has been e x a m i n e d r e c e n t l y ? 9 T h e evidence suggests that t h e r e is a r e s e m b l a n c e b e t w e e n tlfis t u m o r a n d osteosarcomas. It is i m p o r t a n t to point o u t that t u m o r m a r k e r s a n d such a classification s c h e m e will not replace the skills o f the surgical pathologist but will b r i n g the tools o f the biochemist to the surgical pathology bench. BIOCHEMISTRY OF COLLAGEN THE TUMOR MATRIX
AND
C o l l a g e n is the m a j o r c o n n e c t i v e tissue p r o i e i n - - t b e c h i e f c o m p o n e n t o f tile t u m o r m a t r i x as well as the most a b u n d a n t p r o t e i n o f the body. It is now recognized to be a family o f closely related proreins, each a d i f f e r e n t gene p r o d u c t . * Five types o f collagen are now recognized (Table 2). A m i n o acid s e q u e n c e s t u d i e s a n d c y a n o g e n b r o m i d e a n d e n z y m e p e p t i d e m a p p i n g analyses o f n o r m a l tissne reveal even g r e a t e r h e t e r o g e n e i t y t h a n that disclosed by conventional protein c h r o m a t o g r a p h y . M i n o r collagen types h a v e b e e n d e t e c t e d in n o r m a l tissues whose identity a n d relationship to the m a j o r collagen types have not yet b e e n established, z4 T h e same a p p r o a c h can now be a p p l i e d to the matrix o f h u m a n and e x p e r i m e n t a l sarcomas. Unique collagens m a y exist in the matrices o f t u m o r s that are also d i f f e r e n t f r o m those described h e r e t o f o r e . T h e re-
* For recent reviews see references 20 to 22. TABLE 2.
Type I
II III
IV V
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TYPES OF COLLAGEN
Location This is the most abundant type, making up the major protein component of skin, bone, and tendon. However it is ubiquitous in the body. A variant is found in fetal tissue and in some experimental tumors.23 This is cartilage-specific collagen, found also in the nucleu~ po]posus and in certain eye structures. This collagen is a form of reticulin though the relationship between the two is unclear. Type III is present wherever type I is found, but in decreased amounts in hard tissue such as bone and dentin, and in increased amounts proportionately in flexible distensible tissues such as the gastrointestinal tract and blood vessels. It is also found in rapidly proliferating tissue, in fetal tissues, in bone marrow, and in early wound healing and in early cirrhosis.. Type IV collagen"is basement membrane specific. This is a widespread molecule suspected to be a cell surface collagen or a collagen associated with the plasma membrane of cells.
cent pro.gress in t e c h n i q u e makes it possible to exa m i n e m t c r o g r a m quantities for structural identification. 24 Evidence o f the c o m p a r a b l e h e t e r o g e n e i t y o f e l a s tin exists as well as for the tissue specific adhesion proteins c o n t a i n e d within tissue matrix such as fibronectin, c h o n d r o n e c t i n , a n d laminin. 2~-29 T h e r e is also a b a t t e r y o f m a t r i x g l y c o p r o t e i n s ? ~ This p a n o p l y o f extracellular structural m a c r o m o l e c u l e s p o r t e n d s well for the eventual identification o f t u m o r specific matrix p r o t e i n s a n d t u m o r specific p r o t e i n products. T h e h y b r i d o m a t e c h n i q u e makes it possible to g e n e r a t e with relative ease large a m o u n t s o f m o n o clonal antibodies directed against each o f these struct u r a l m o i e t i e s a n d v i r t u a l l y all c o n n e c t i v e tissue macromolecules? z Monoclonal antibodies directed against type I, II, a n d IV collagen have b e e n reported, a3-3~ and it is within reason to a s s u m e that a battery o f such antibodies m i g h t soon be available for use in surgical p a t h o l o g y f o r i m m u n o f l u o r e s c e n c e staining o f frozen sections o f t u m o r s . S o m e o f the variants o f soft tissue s a r c o m a s are sufficiently rare that e v e n histopathologists in large medical centers are p r e s e n t e d with serious p r o b l e m s . T h e r e are no centers large e n o u g h for o n e individual to see e n o u g h e x a m p l e s o f the variants to be familiar with the a p p e a r a n c e o f all o f t h e m . An a r r a y o f antibodies for use in e x a m i n i n g specific stromal prot e i n s will a u g m e n t t h e a b i l i t y o f t h e s u r g i c a l pathologist to m a k e such diagnoses. Not only t u m o r specific structural m a t r i x proteins but u n i q u e patterns o f organization o f conventional matrix proteins m u s t be c o n s i d e r e d in this context as diagnostic aids. It is i n c u m b e n t o n surgical pathologists to c o o p e r a t e with biochemists at s o m e stage o f this dialogue in o r d e r to d e m o n s t r a t e the u s e f u l n e s s o f these reagents. T h e r e is a m a n d a t e a d d r e s s e d to all clinicians to translate recent a d v a n c e s in biology m o r e rapidly a n d m o r e effectively into the i m p r o v e m e n t o f medical care. A liaison b e t w e e n biochemists who study structural proteins a n d surgical pathologists m a y e x p e d i t e this process. Surgical p a t h o l o g y is at arf i m p o r t a n t j u n c t u r e ~ t h e o p p o r t u n i t y to b r i n g the r e c e n t advances in biology to b e a r on tile i m m e d i a t e care a n d t r e a t m e n t o f certain c a n c e r patients.
SUMMARY
- T h e surgical pathologist is often called u p o n to m a k e a r a p i d decision aboiat the m a l i g n a n t potential o f a soft tissue t u m o r . T h e s e t u m o r s have s p e c i f c stromas that s u p p o r t g r o w t h and e n d o w the t u m o r with u n i q u e quali~ies, as well as being manifestations o f the p h e n o t y p i c e x p r e s s i o n o f t u m o r cells. Collagen, the m a j o r p r o t e i n o f the s t r o m a o f t u m o r s , is now r e c o g n i z e d as a family o f proteins, each a d i f f e r e n t gene product. Immunofluorescence microscopy studies u s i n g a n t i b o d i e s d i r e c t e d a g a i n s t p u r i f i e d s t r o m a l c o m p o n e n t s , particularly the d i f f e r e n t type
H U M A N S O F T T I S S U E SARCOMAS---STERN s p e c i f i c c o l l a g e n s , m a y s o o n b e a v a i l a b l e in l a r g e q u a n t i t i e s . T h e s e a r e p o t e n t i a l l y u s e f u l tools f o r t h e surgical pathologist not only in m a k i n g that i m p o r t a n t d e c i s i o n , b u t also f o r d e v e l o p i n g a classification s c h e m e f o r h u m a n s a r c o m a s . T h i s s c h e m e will i n e v i t a b l y h a v e a m o r e s o u n d biological basis t h a n t h o s e currently used.
ACKNOWLEDGMENT I am indebted to Diane Jarest for her skillfld help in preparing tiffs manuscript.
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