Spontaneous and Estrogen-Produced Tumors in Nb Rats and Their Behavior After Transplantation

[CANCER RESEARCH 35, 766-780, March 19751

Spontaneous and Estrogen-produced Tumors in Nb Rats and Their Behavior after Transplantation

Robert L. Noble, Brenda Clayton Hochachka,' and Diane King2

Cancer Research Centre,' University of British Columbia, Vancouver, British Columbia, Canada V6 T I W5

SUMMARY INTRODUCTION

Tumors in rats of the Nb strain, arising either spontane This paper will introduce a large series ofdifferent types ously or after prolonged treatment with s.c. pellets of of tumors, arising either spontaneously or under the influ estrogen, were transplanted to establish whether hormone ence of estrogen in our colony of the Nb strain of hooded conditioning was required for their growth. Whereas all rats over the past 8 years. One objective was to establish spontaneous tumors arising in males and many of those in transplantable tumor lines in order to study hormone females were autonomous on transplant, most of those dependency and factors controlling the change to autono arising in estrogenized rats continued to require hormones mous growth. Hormonal carcinogenesis is a slow process, for growth after transplantation. The latter included carci extending over one-half of the life-span of the normal rat, nomas of the adrenal cortex, breast, pituitary, pituitary and even transplant experiments may involve observations ectopic tissue, ovary (thecomas), uterus (leiomyomas), over a year. Preliminary observations on a few types of cervix, vagina (fibrosarcoma), Leydig cells oftestis, thymus, tumors, however, have been presented in abstract form pancreas, salivary glands, orbital gland (fibroadenoma), (15â€”18). liposarcoma, and lymphoma. Many of the tissues of origin Our interest in rat endocrine tumors first arose in Dr. J. of the tumors have not been considered to be under the B. Collip's laboratory at McGill University in 1940, when it influence of estrogens. A type of hormone-responsive tumor was shown that mammary carcinomas followed estrogen that was inhibited by estrogen and that grew only in normal application and that such tumors regressed if the estrogen rats is described. All estrogens tested, including estriol, were pellet was removed (20, 22). These studies were confirmed interchangeable in action. and extended by Cutts some years later, using the same The incidence of the more common tumors of the strain of rats (3, 5). Our present initial objective was to adrenal, breast, and pituitary was very low in normal rats, restudy such tumors and to establish transplantable hor but higher in females. All tumors were more common after mone-dependent tumor models. In many cases tumors of estrogenization in both sexes, particularly in older animals. other organs were encountered, so that the study now The secretion of steroids and pituitary hormones by many includes more than 196 tumors of 18 organs including tumors led to obvious biological effects. Pituitary secretion hormone-induced tumors or tumors of spontaneous origin. led to severe lesions frequently associated with diseases in Endocrine tumors of the rat lend themselves not only to humans, but the signs of such diseases in the rat apparently studies on induction by hormones but also to the effects of were hormone dependent and disappeared if the tumor was hormones on the growth and secretion by the tumors. Many removed. The overall results raised the possibility that contributions have been made in this field but have been estrogens were not carcinogenic per se but stimulated the limited mainly to breast and pituitary tumors. Older growth of previously altered cells and that, following their publications on mammary tumors of rats (2 1) and the more transplantation, this hormone requirement was retained. general subject of tumors and hormones (I, 14) have been Irrespective of the mechanism of carcinogenesis, hormone reviewed. Although the work, particularly that of Furth, dependent tumor growth was not irreversible but was Huggins, and Dunning, has led to a number of transplanta controlled in an unexpectedly wide spectrum of organs by ble tumor lines that are widely used, it is difficult to define exogenous estrogen. Host factors may play a major role in these in an acceptable classification of â€œhormonedepend controlling the growth of many tumors and the ultimate ency.â€• A few of these tumors, although not induced by course of the disease. hormones, have been widely distributed and accepted for studies as models from which generalizations have become accepted. As will be shown, many of the new tumors to be described do not fit into these established patterns. More recentexperimentshaveindicatedthattestosterone 1 Collaborated in the Research Program, 1969 and 1970.

2 Senior Assistant, 1962 to 1966 and 1969. may also induce tumors if applied over prolonged periods,

3 A cancer unit supported by the National Cancer Institute of Canada. particularly to females or to rats of either sex following Received May 21, 1974: accepted December 9, 1974. pretreatment with estrogen. It appears that different hor

766 CANCER RESEARCH VOL. 35

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 1975 American Association for Cancer Research. Hormone-dependent Rat Tumors mones acting together or sequentially may cause tumors tumor takes). The exact absorption of estrone from similar differing in type and location from those induced by pellets has been measured in this laboratory using radioac estrogen alone, but such results will be reported separately. tive steroid (10). The results showed that the weight change From all observations, a striking similarity is apparent in of the pellet closely reflected the loss of hormone. All the rat tumors to be described, not only morphologically but tumors arising in estrogen-treated rats were considered to also in hormone responsiveness. be induced by the hormone, in contrast to spontaneous ones found in normal male or female animals. Experimental Procedures. Tumor transplants were made MATERIALS AND METHODS from finely scissored pieces of viable growing tumor areas (avoiding hemorrhage and necrosis), were placed in 0.9% Rats. A black-hooded rat colony has been maintained by NaCI solution and antibiotics, and were injected via a the author for 33 years (originating from Dr. J. B. Collip's trochar into the midline intrascapular area of the neck (2 to laboratory at McGill University) and has recently been 3 pieces of I to 3 mg) into groups of 2 or 3 hormone-condi assigned the abbreviation Nb strain by the Medical Re tioned and control rats. The most rapidly growing tumors search Council, Carshalton, England. The exact origin of were routinely selected for transplant. Rats receiving trans the strain is somewhat clouded by antiquity, but it probably plants were I .5 to 3 months of age. The use of rats older arosefrom Long-Evansrats obtained from Dr. Philip Smith than 6 months led to conflicting results. Intact animals were at Columbia University. Tumors will rarely grow in this used in most cases, representing unconditioned controls, but strain but will not grow in a hooded strain of McGill origin ovariectomized or ovariectomized-hysterectomized rats sold commercially in Montreal. Originally albino, yellow-, were also used. The latter eliminated a common complica and red-hooded offspring were discarded and black-hooded tion of cystic changes and infection in the ovaries and rats were used for breeding. Dr. Cutts had little difficulty in uterus, which frequently occurred after estrogen treatment starting inbred strains (accepting crossed skin grafts) at the and particularly after estrogen-androgen therapy. Ectopic University of Western Ontario. The present strain is of tissue transplants were grafted into the s.c. abdominal random litter-mate breeding over 12 to 14 generations in 12 tissue. Rats were inspected weekly or more frequently when years. Animals are always black-hooded, moderately sized, bearing tumors. Early mammary and some other superficial hardy rats that breed readily, are extremely docile, and are tumors could be located by palpation. Many gross tumors, ideal for experimental work. The colony is not pathogen however, were only discovered with a complete postmortem free. The breeding colony and stock animals are isolated in examination. Any rats that showed signs of illness were areas separate from all experimental rats. Exposure to killed and examined. Animals that died of natural causes known exogenous viruses or to infected mice seems virtually were also examined so that most gross tumors were impossible. An inbred colony of Fischer rats originally recorded. Hormone-dependent tumors have had to be obtained from Dr. A. Segaloff some 10 years ago is also maintained by serial transplantation because they appear to maintained in the laboratory. The rats are fed Purina chows be particularly vulnerable, seldom survive freezing tech and have tap water ad !ibitum. niques, and have not retained hormone-dependent charac Steroid Pellets. Pellets were made individually in a hand teristics after tissue culture. press and, for routine work, consisted of approximately 10 Experimental Interpretation. In assessing whether a mg (2 mm in diameter) of a 90% steroid: 10% cholesterol tumor would grow on transplant, an arbitrary time of 6 mixture. Pure steroid pellets may be made if large crystals months was selected for the presentation of results in this of the hormone are used, but the use of cholesterol as a paper (most growing tumors in estrogenized hosts allow a binding agent facilitates making the pellet. Pellets of the survival of only 2 to 3 months, but many slower-growing same size containing various percentages of hormones were sublines were established). Tumors were listed as not made by increasing the amount of cholesterol. A EP4 was growing if they could not be maintained by transplant for 3 hard and did not flake or crumble but retained its shape generations. Successfully transplanted tumors were classed even when reduced to about one-half of its original size after as dependent or autonomous only when they showed the a year in the s.c. tissue. A 10 or 20% pellet lost only 1 to 2 same growth patterns for at least 3 generations. Grafts of mg in the same period, as the cholesterol binder was very normal tissue or of tumors remained viable for indefinite insoluble. Similar pellets were prepared using other steroids. periods in this inbred strain of rats, and no evidence of tissue Estriol was somewhat more soluble, and a 90% pellet rejection or histological reaction was observed. disappeared in approximately 6 to 7 months (stilbestrol pellets dissolved in 3 months). Pellets could be palpated and removed surgically at any time. In most experiments a pellet RESULTS was inserted into the rat at the time of tumor implant (prior pelleting was of no advantage, and the removal of the pellet Endocrine Effects of Estrogen Pellets did not impart a subsequent hormonal status suitable for The continuous absorption of estrogen from pellets had a profound endocrinological action on both male and female 4 The abbreviations used are: EP, 90% estrone + 10% cholesterol pellet; STH, somatotrophic hormone: ACTH. adrenocorticotrophic hor rats, which is pertinent to this paper. The dosage of estrogen more. was estimated by weighing the pellets after prolonged

MARCH 1975 767

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 1975 American Association for Cancer Research. R. L. Nob!e et a!. sojourns in the animal. Although pellets from 6 to 13 mg to increase gradually with age throughout life, from 3.5 dissolve at slightly different rates, it was found that the mg/l00 g body weight at 6 weeks to 6.4 mg/l00 g at 58 weight of 94 pellets (from 7. 1 to I 3.2 mg) composed of 80 to weeks, whereas in males it did not change from 3.4 mg/l00 100% estrone after 370 days in the rat, was reduced to an g. This early observation on 230 rats has not been tabulated average of 46% of the original, indicating an average but, in confirmation, data from 23 normal males and 21 absorption of 0.42 mg of estrone per month. The effects females averaging 16 and 15 months of age, respectively, were reflected particularly in the body weight and testes size were taken from consecutive autopsies. The females aver in the male rat, and these parameters have been used for aged 250 g in weight and the pituitary averaged 20.5 mg (14 simple biological measurements. In view of the unexpected to 38 mg) = 0.8 mg/ 100 g, whereas the figures for the males activity shown by pellets of estriol, some biological data were 360 g to 12.5 mg (10 to 15 mg) = 0.35 mg/l00 g body comparing estrogens have been summarized in Chart I and weight. Table 1. In other experimentsrats wereexposedto pelletscontain The initial growth of the young rat was relatively normal ing lower levels of estrone ( I and 10%), estriol ( 10%), and after receiving a EP. The growth rate then diminished and, diethylstilbestrol (10%). Only the more soluble 10% stilbes finally, the weight plateaued at about 180 and 220 g for trol pellets had detectable estrogen effects. Male rats with females and males, respectively. When animals above these other pellets were found to be fertile and sired normal weights received pellets, the weight plateaued sooner; in litters, although in females a continuous estrus vaginal larger animals, weight loss was seen. smear was usually present. The effects on various organ weights of hormones As will be described, estrogen-dependent tumors have absorbed from pellets are shown in Table 1. The numbers been noted in a number of organs that are supposedly not indicate the same groups of rats corresponding to those under endocrine control, such as the salivary gland (sub whose weight curves were shown in Chart I. mandibular glands). This tissue was readily dissected and Rats receiving the higher doses of estrogen showed weighed in groups of 20 rats after the rats had received testicular atrophy and the lack of secondary androgen various hormones. The weight in relation to body weight secretion, reflected by atrophy of the ventral lobes of the was remarkably constant in rats of either sex (mg gland prostate (in 19 (2.7%) of690 rats that had been pelleted for a weight per g body weight; females, 1.32 Â±0.15; males, 1.23 minimum of 2 months, the expected atrophy of the testes Â± 0. 14) and was not significantly affected by estrogen or was not found, even though other signs of estrogenization androgen pellets. Rats bearing STH-secreting pituitary were present). Removal of estrogen pellets even after many tumors with or without a EP did not show a corresponding months [except when pelleting was before 2 weeks of age increase in weight of the salivary gland. (19, 23)] was followed by a rapid resumption of body growth and increased weight of the gonads associated with a return of pituitary weights towards normal. Tumor Incidence An increase in size of the anterior lobe of the pituitary was a constant finding in estrogen-treated rats. The pitui The incidenceof the most frequently encounteredtumors tary weight of normal female, but not male, rats was found of the pituitary, breast, and adrenal cortex over a 4-year period is shown in Chart 2. Tumors were not noted in rats before 8 months of age (except for pituitary hypertrophy in estrogenized animals). @0 With the exception of mammary fibroadenomas, spontane @00 ously arising tumors were seldom found in 299 males and JI were uncommon in 565 females, although more occurred in animals over 1 year of age. In 660 estrogenized rats, the

320 tumor incidence in all organs was sharply increased in both sexes but was greater in females and in animals treated for I 280 year or more. In certain experiments to induce breast p-240 tumors, it was noted that, ifestrogen treatment commenced I â€˜a in 2- to 3-week-old rats, carcinoma, usually of multiple @200 breasts, eventually developed in nearly all animals of either 0 Â£260 sex. On the other hand, if treatment was delayed until 4 months of age, few tumors developed. These findings may 120 be seen in Chart 3.

â€¢0 Adrenocortical carcinomas were induced readily in both sexes. In a review of 274 animals with such tumors, there 40 was no difference if rats were pelleted at 4 or 9 weeks of age, although, in a few rats receiving pellets at 6 months of age, _0_@4_@ I 12 16 20 24 28 32 WEEKS small or microscopic tumors appeared to be more frequent. Chart I . Effects of estrogens (treatment or removal) on body weight of Male rats appeared to have a slightly higher incidence of male Nb rats. I, average of untreated rats: bars, variation. 2 to 13, average pituitary tumors. Old estrogenized rats frequently had a of treated groups of rats. Treatments listed in Table I. number of tumors occurring in different organs, but no

768 CANCER RESEARCH VOL. 35

Table 1 Organ weights of male rats pelleted with various eszrogens'@ Rats killed after treatment for 52 or more weeks.

VentralGroupTreatmentNo. of (mg)IControls323446Â±300b309Â±4044Â±515Â±22EP13260Â±6010Â±441Â±5184Â±733E3P'@12310Â±10110Â±545Â±10190Â±654STP8265Â±17014Â±650Â±1262Â±645EP(x2)14350Â±8010Â±241Â±7198Â±486EP+E3P(x2)6320Â±11012Â±541Â±7140Â±317E3P(x2)6230Â±4511Â±244Â±2145Â±358EPat8wkfor53wk7300Â±4914Â±552Â±9156Â±ratsTestes (mg)prostate (mg)Adrenals (mg)Pituitary

329EPatl2wkfor53wk5375Â±3010Â±254Â±7158Â±5210EPat38wkforlOwk8658Â±21048Â±

1644Â±9IIEPat3wk:outatl6wk91092Â±40033Â±2546Â±1723Â±8for 1666Â±

days12EPat3wk:outatl8wk121660Â±87074Â±5554Â±1815Â±8for5wk13EPat3wk:outat24wkS3350Â±400193Â±13756Â±1716Â±4for17

32 wk

a Same groups of rats as shown in Chart I. b Mean Â± S.D.

C E3P, 90% estriol + 10% cholesterol pellet: STP. 90% diethylstilbestrol + 10% cholesterol pellet.

100 IL?5

In 0 I U) 80 0

z C.) a I- 70 â€¢12I In 4 4 U) @ 90 32 @60 LII 80 C) D @l70 >- 60 @50 â€¢25

50 I â€˜4 60 I 40 â€¢@iâ€¢36 30 w -J i;i30 â€¢27 > I ADRENAL BREAST ANT. PIT. I- @ CA FIBROAD CA (v60mg) 20 â€¢io U) I-. 12â€¢SIB Chart 2. Incidence of the commonest tumors in normal and estroge 4 @ nized Nb rats. CA, carcinoma: FIBRO AD, fibroadenoma; ANT. PIT., 10 â€¢to anterior pituitary. Â£5 U)@1 2 4 6 8 10 consistent pattern was seen. In a small series of 80 retired AGE OF RAT AT EP (Weeks) female breeders (12 to 14 months old) 2 pituitary and 2 Chart 3. Effect of age at the start of EP treatment on the development adrenal tumors were noted, a higher incidence than was of gross mammary tumors. Numbers, number of rats at each point. expected in normal females. Observations have also been made on a small number (approximately 100) of estroge nized Fischer rats. Pituitary adenomas occurred earlier and Tumor Induction in Ectopic Transplanted Tissues grew more rapidly than did similar tumors in Nb rats (50 and 99% incidence was found at 7.5 and 10.5 months, Anterior Pituitary. The anterior pituitary from normal respectively) but could not be maintained through 3 genera female rats was grafted to the s.c. tissues of other female tions by transplantation. No adrenal or mammary tumors animals. In 14 instances remnants of the graft could be were found, but the life expectancy was relatively short due identified, even after 26 months. In another group of to the development of pituitary tumors. animals that received an estrogen pellet at the time of the

MARCH 1975 769

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 1975 American Association for Cancer Research. R. L. Nob!e et a!. graft and an additional estrogen pellet at 5 months, growth assumed abnormal cell characteristics and showed local occurred in 5 of 12 grafts, so that after 11 months they areas ofcell extension into the basement membrane. Tumor weighed an average of 37.1 mg (18.6 to 70.4 mg). Similar cell masses, although apparently of a benign adenomatous experiments have been reported by others (25). Such grafts appearance, transplanted readily and then developed more resembled small chromophobe adenomas and transplanted aggressive cellular characteristics. Extension into surround successfully to become rapidly growing carcinomas. En ing tissue was uncommon, as was involvement of regional larged pituitaries obtained from rats pelleted from 2 to 4 lymph nodes. Early metastases were found with only a few months were also grafted in the same way, but these did not types of tumors and usually arose from blood-borne tumor show an increased incidence of tumor formation. cells. Successive transplanted lines of most dependent Adrenal. Adrenal cortical tissue obtained from glands tumors retained the same structure as the primary line, stimulated to 10 times normal size by an ACTH-secreting although the glandular secretory structure tended to be tumor and grafted s.c. to estrogen-conditioned rats has replaced by more solid cell carcinoma and stromal materials failed to grow in 4 tests. often tended to increase, particularly in cancer of the cervix and some breast tumors. The primary chromophobe struc Tumor Transplantation ture of pituitary adenomas, after the 1st transplant, was rapidly replaced by carcinoma with increasing anaplasia and cystic degeneration. Spontaneously arising tumors, In order to obtain transplantable tumor lines, most although similar to dependent types, were usually more spontaneous and induced tumors were tested to see if they anaplastic and showed rapid growth even in the initial would grow in young untreated or EP-conditioned rats. transplant. A mast cell infiltration of the stroma was a Mammary fibroadenomas have not been recorded because they grew irregularly when transplanted and seldom exhib striking feature of many growing tumors, as noted by ited a consistent response to hormone manipulation. A type Combs and Purnell (2), whereas eosinophil cell infiltration of pituitary tumor is described and referred to as a was seenonly in regressing tumors ( I 3). Tumor growth rates hormone-inhibited tumor. At present 4 of these tumors are extended over a wide range, but rapidly growing tumors being maintained by transplant, all of which apparently are exhibited many mitotic figures and tended to become hemorrhagic with necrotic centers. Slowly growing tumors unable to grow in estrogenized animals (they regress in the normal rat if estrogen is administered and exhibit the had few mitoses and showed no signs of a chronic cellular reaction. opposite types of responses when compared with hormone Adrenal Cortex. In 22 primary induced tumors (5 in dependent tumors). Transplanted tumors of the adrenal, males), the age of the rat when the tumor was discovered breast, and pituitary have been most extensively studied and averaged 13 months (10 to 22 months). The tumors arose are listed in Table 2. In the case of tumors of other organs, singly and varied from 60 mg to 24 g in weight; 8 tumors the total numbers occurring spontaneously or in estroge nized animals are noted, as is the hormonal status of their weighed less than 1 g. The left adrenal was involved twice as frequently as the right in 25 animals. The opposite gland transplants. The difference in the total number of tumors and successful transplants usually indicates tumors that did was always atrophic. Small subcapular adenomas were not not grow after transplantation, as nearly all tumors were uncommon in this strain of rats, but carcinomas appeared to transplanted. Tumors of the breast and pituitary could not arise from the zona reticularis near the medulla (Fig. I) and be transplanted successfully in approximately 30% of cases. could be induced with estriol (Fig. 2). Microscopically, the Similarly, tumors of smooth muscle of the uterus and cervix tumors resembled normal adrenal structure. Metastases and Wilms' tumor of the kidney rarely grew on transplant. were a striking feature in 15% of animals with primary From Table 2 it may be seen that transplanted tumors of tumors. In subsequent studies of 806 transplants of 14 14 organs have shown an estrogen effect on their growth. different tumors, an overall gross rate of 13% metastases These include such tumors as carcinomas of the salivary was noted (in some tumor sublines, however, the incidence gland and pancreas, a liposarcoma, and a lymphoma arising varied from 0 to over 50%). In 100 consecutive autopsies, in organs not usually considered under estrogen control. metastases were present in the liver (58%) lungs (54%), Some tumors of the pituitary and kidney were found to be adrenals (18%), ovaries (5%), thymus (1%), spleen (1%), and inhibited by estrogen upon transplant. A striking finding kidney (1%). Metastases were usually multiple and fre was that all spontaneous tumors arising in males and most quently massive. In the ovary, metastases grew only in of those arising in femaleswere autonomous on transplant. corpora lutea and the cortex of the adrenal. Most tumor On the other hand, transplants of most tumors arising in lines killed the host after about 2 to 4 months, but some estrogenized animals ofeither sex were estrogen dependent. slower-growing lines only required transplanting every 6 months. Neither spontaneous adrenal tumors of males nor autonomous transplants have been observed. Tumors: Description and Morphology Mammary Gland. Induction studies have resulted in over 500 mammary tumors, but only a small number has been Most primary hormone-dependent tumors and their transplanted. In 15 rats with primary tumors from which transplants were slowly growing, well-differentiated carci transplant lines were established, the average age at autopsy nomas of a low grade of cancer. Early epithelial hyperplasia was 13 months, and tumors varied from 0.25 to 31 g in resulted in accumulation of cellular areas that gradually weight. Tumors usually occurred singly in a breast, but

770 CANCER RESEARCH VOL. 35

Table 2 A new series of tumors and transplants that respond to estrogen

oftransplants and No.responseSpon ofprimary tumorsNo. hormone

taneousSpontaneousOrganTumortypeM

InducedAdrenal FInducedM F

cortex 24D Breast Carcinoma 1 5 2O@ 1A 5A 19D, IA Anteriorpituitary Carcinoma I 2 28a IA 2D 24D,2A,2Hl Anterior pituitary Carcinoma 5 3D (ectopic graft) Ovary Thecoma 6 4 2D, 4A 2D, 2A Uterus Leiomyoma 4 2 2D, 2A ID Cervix Leiomyoma 6 4 ID, IA Carcinoma I ID Vagina Fibrosarcoma I 2 ID. IA Carcinoma I Testis(Leydigcell) Carcinoma 2 1 IA ID Thymus Carcinoma II 2 IA IA 2D Lymphosarcoma I IA Liposarcoma I ID Orbit gland Fibroadenoma I ID Salivary gland Fibroadenoma I ID Carcinoma I 2 5@ IA 2A 3D, IA Pancreas Acinar carcinoma I 4 2D, 2A Lymphoid Reticulum cell 5 1 1 5A IA ID KidneyCarcinoma Carcinoma I 1HI 2Aa Wilms' nephroblastoma5 I 324a @a5D4

At least 20% of tumors in males. 4 D, dependent: A. autonomous: HI, hormone inhibited. multiple breasts were frequently involved. Small tumors of showed localized areas of proliferative growth. The ectopic 20 to 40 mg, usually in the nipple area, consisted mainly of pituitary grafts showed a similar picture. Transplants fibromatous stroma with overgrowth of areas of duct-like seldom retained the chromophobe structure (Fig. 5) but epithelial cells. As the tumor size increased, the glandular rapidly became carcinomas composed mostly of acidophilic element predominated and areas of epidermoid metaplasia anaplastic cells (Fig. 6). Rats with primary enlarged pitui frequently developed. Malignant lesions were not associated tary glands showed an absence of trophic hormonal effects, with excessive mammary gland stimulation of the host. in contrast to rats with the transplanted carcinomas, many Dependent tumors were of all types but predominantly of which secreted 5TH, ACTH or, less frequently, luteo papillary or solid adenocarcinomas with little stroma and trophic-like hormones. Metastases were infrequent. Exami little evidence of local invasiveness. Bone metaplasia was nation of the glands from which 2 estrogen-inhibited lines found occasionally. Estriol was an effective estrogen even in originated showed that they apparently were both well-dif the male (Fig. 3), (fibroadenomas usually appeared as ferentiated, slowly growing masses of chromophobe cells. single, hard, lobulated white oval tumors recognizable in the The interesting observation has beenmade repeatedlythat, gross). Spontaneoustumors aroseat an earlier agethan did if an EP rat bears a STH-secreting tumor, pituitary induced tumors and, in 5 females, were all found to be hypertrophy and adenoma formation occur rapidly so that, autonomous on transplant. Similarly, a tumor in a male of in a period of 3 months, the pituitary might weigh 100 mg or only 2.5 months of age grew rapidly upon transplant (Fig. more. A spontaneously arising tumor was found in a male 4). In some cases tumors appeared as rapidly growing rat of 17 months. It showed marked ACTH stimulation, undifferentiated spindle cell neoplasms, similar to those and this characteristic was retained in rapidly growing frequently found when transplanted dependent carcinomas autonomous transplants. became autonomous. Very few tumors metastasized, usu Ovary. Thecomas were found in untreated or estrogenized ally to liver or lungs and, exceptionally, to regional lymph rats, although in the latter case cystic inflammatory tissue nodes. tended to obscure tumor formation. Tumors were found in Pituitary. Adenomas of the anterior pituitary occurred in old rats averaging I 3 months of age, although 2 induced all estrogenized rats and usually attained a weight of 60 mg tumors were found in 4-month-old rats. Spontaneous or within 6 months. Within a year most glands that might induced tumors showed closely packed whorls oftumor cells weigh 300 mg appeared as diffuse masses of highly vascular and did not produce hormones (Fig. 7). Estrogen-dependent channels surrounded by chromophobe cells and only rarely transplants grew readily in normal female rats but not in

MARCH 1975 771

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 1975 American Association for Cancer Research. R. L. Noble et a!. ovariectomized or male rats. and all rats with swellings of this gland have been carefully Uterus. Tumors of the uterus occurred in normal or examined. Inflammatory lesions and abscesses were not estrogenized rats usually when over I year of age. They infrequent. Some tumors were found in relatively young caused elongated swellings in the muscle layers, reaching 5 g pelleted rats, as 3 animals were between 3 and 4.5 months in weight. Epidermoid metaplasia occurred in estrogenized old. These well-differentiated secretory carcinomas, arising rats and in some cases showed areas resembling carcinoma in estrogenized females, have been totally estrogen-depend in situ. Most tumors appeared to be leiomyomas (Fig. 8), ent on transplant (Fig. 19); 1carcinoma, arising in a treated although a transplantable autonomous tumor arising in an male, was autonomous. All 3 spontaneously arising tumors 8-month-old rat showed increased mitosis and sarcomatous were autonomous and showed marked anaplasia or papil change (Fig. 9). lary growth (Fig. 20). Cervix. Tumors were confined to the cervical area in 10 Pancreas. Carcinomas of the glandular pancreas were rats of an average age of 13 months and, in some cases, found in 4 female rats estrogenized from 9 to 12 months and protruded from the vagina. Most tumors in normal or in I retired breeder. In all cases the animals were killed estrogenized rats resembled the leiomyomas of the uterus because of increasing abdominal distension caused by and transplanted poorly. In I case, however, a typical hemorrhagic fatty ascitic fluid and hundreds of small carcinoma arose in a rat pelleted for only 2.5 months. This white metastatic nodules throughout the peritoneal cavity. showed only small amounts of stromal material initially, Transplants of 2 tumors showed well-differentiated glandu but later the epithelial element was contained in extensive lar structure and were under estrogen influence (Fig. 2 1). areas of fibrous substance. Transplants retained this mor The autonomous tumors were more scirrhous or papillary in phology and were estrogen dependent (Fig. 10). type (Fig. 22), and some tranplants metastasized to the Vagina. Extensive infiltrating tumors protruding from spleen and other organs. and confined to the vagina were found in 2 rats after 8 and Lymphoma. Spontaneously arising lymphomas, usually in 10 months of estrogenization. On section these appeared to male rats over I year of age, have presented as enlarged be fibrosarcomas resembling sarcoma botryoides (Fig. I I). livers, spleens, or lymph nodes and have been transplanted Transplants of 1 tumor grew slowly in unconditioned hosts in 6 cases. They grew progressively in unconditioned but more rapidly in estrogenized rats. A tumor in a animals. Most tumors appeared to be of reticulum cell type 15-month-old EP rat was found to be a carcinoma (Fig. 12), and spread readily to various organs. Giant cells may occur but this tumor was not transplanted. in the spleen (Fig. 24), but these may respresent bone Testis. Tumors have been found in a testis of a 13-month marrow metaplasia. A diffuse lymphoma was found in a old pelleted rat (Fig. 13) and in two 22-month-old retired male estrogenized for 10 months with enlargement of all breeders (Fig. 14). The estrogenized animal had a large lymphoid tissue but with a normal blood smear. Transplants scrotal hernia. All tumors were similar carcinomas of the of this unique tumor were readily stimulated in growth by Leydig interstitial cells. In the spontaneous tumors, the estrogen and usually spread throughout the body, even to seminiferous tubules of both testes were atrophic, suggest massive invasion of the pituitary and kidney (Fig. 23). ing a pituitary factor deficiency. Slowly growing estrogen Kidney. A carcinoma of renal tubule appearance was dependent sublines of the induced tumor have been carried found in an estrogenized female, but some months after the in transplant for 5 years and continue to cause intense EP had been removed. This tumor did not grow in androgenization and hernia of male or female hosts. In estrogenized rats through 5 transplant generations, similar contrast, early transplants of a spontaneous primary tumor to pituitary tumors, noted previously that were estrogen were autonomous and have not produced androgenization. inhibited. Nephroblastomas were not uncommonly found as Thymus. Diffuse or lobular hypertrophy of the thymus large tumors in 6- to 12-month-old normal or treated rats. has been observed frequently, but in 10 cases such lymphoid Transplantation has only been successful in 2 of the tumors could not be successfully transplanted. Epithelial autonomous Wilms' tumors and, in 1 animal, was associ tumors arose spontaneously in 2 old rats and were autono ated with marked changes characteristic of 5TH stimula mous on transplant (Fig. 16), as was a lymphosarcoma. On tion. the other hand, a similar induced tumor, after estrogeniza Other Tumors. Interest has focused on the preceding tion for 2 months, showed greater epidermoid metaplasia tumors of 14different organs and on their transplants, all of and was initially dependent on transplant (Fig. 15). A l0-g which had the common property of estrogen-influenced tumor found in the thymus of a 9-month-old female was growth. A number of tumors of other organs (usually found to be a liposarcoma (Fig. 17). Transplants of this arising spontaneously) have been autonomous on trans unusual tumor have been estrogen dependent for 6 genera plant. These included: brain, a rapidly growing undifferen tions. tiated astrocytoma (a slower-growing and a well-differen Orbital Gland. A tumor arising in the orbit and extending tiated blastoma arose in a pelleted rat); ear duct, sarcomas around the eye was discovered in an estrogenized female of and a sebaceous gland adenocarcinoma found in old 7 months. The exact origin is uncertain, but the sections animals; leukemia, transplantable monocytic leukemia showed a fibroadenomatous tumor, transplants of which occurring in 2 rats over a year in age; sarcoma, rapidly were hormone dependent (Fig. 18). growing spindle cell sarcomas found in 3 old rats. Salivary Gland. Eight carcinomas and I fibroadenoma, 3 A number of nonspecificlesionshasbeennoted. In earlier to 5 g in weight, of this gland have been found since 1970, studies 8 rats showed a severe arteritis, particularly in

772 CANCER RESEARCH VOL.35

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 1975 American Association for Cancer Research. Hormone-dependent Rat Tumors tortuous mesenteric vessels. Only 4 of the rats were induction times of I year or more has its limitations, but the estrogenized, contradicting Cutts' conclusion that similar described derived, hormone-dependent transplantable lines lesions were related to estrogen treatment (4). Pyometritis with more rapid growth may be of greater practical value. and ovarian cysts with infection were common in estrogen Many of the tumor models bear a striking resemblance, not treated animals, especially if combined with androgen. only in growth patterns but also in morphology, to counter Chronic lung infection, middle-ear disease, and subacute parts in humans. The unexpected finding that rat tumors of arthritis occur in this colony of rats, and overgrowth of such organs as the salivary gland, pancreas, lymphoid, and incisor teeth (despite a hard pellet diet) is frequent in old fatty tissue may be dependent on estrogens for growth raises animals. the question of whether similar responsive tumors might occur in man. Pellets of estrone allowing a slow and continuous release Pathological Changes Secondary to Hormone-secreting Tu of hormone have been used for routine studies, but pellets of mors. diethylstilbestrol and related synthetic estrogens as well as estriol have also induced tumors. Estriol pellets were not The postmortem findings on rats bearing many of the shown to have a competitive action against estrone ( 12) but types of tumors described may be unusual because of exerted an unexpectedly powerful biological endocrine â€œhormonesâ€•secretedby the tumors. Some pituitary trans action. It is possible that the tumorigenic action of estrogens plants produced tumor hormones or combinations that was enhanced by the pellet form of administration; this seemed to show selective organotrophic activity, suggesting would suggest that the use of estrogens in pellet form in the possibility that modified types of hormones predomi humans should be viewed with caution. On the other hand, nantly affected breast, thymus, liver, kidney, etc. Gonad the Nb strain of hooded rat may be particularly sensitive to atrophy in the male without associated mammary develop tumor formation by estrogen pellets, as rat strains have ment accompanied some autonomous tumors. The hosts been found by others to exhibit different responses to bearing adrenal tumors sometimes showed biological effects estrogen (3, 6, 11). associated with glucocorticoid or mineralocorticoid excess, The finding that transplants of tumors of 14 organs that particularly in early transplant generations. Transplants of developed under the influence of estrogens continued to I adrenal tumor produced intense androgenization of cas have hormone-influenced growth suggests that some com trated rats. Ectopic ACTH production was found with 2 mon mechanism may be involved, particularly since sponta transplanted breast tumors. There was no constant relation neous tumor transplants were autonomous. Tumors of ship between the hormone dependency or autonomy of different organs and even of the same organ in many cases tumors and hormone secretion by the tumors, and even showed individual characteristics. Carcinomas of the adre sublines of the same tumor might give different findings. nal cortex transplanted readily and grew at widely different Severe pathological lesions resembling those found in rates, while retaining hormone dependency, and have been human disease were associated with the secretion of differ used as a typical model. These tumors spontaneously ent tumors. Such changes included cystic mammary disease metastasized widely, in contrast to most of the other types (without malignant change), kidney and cardiac hypertro of tumors, and the distribution of metastases could be phy and nephrotic changes (with extreme albuminuria), duplicated by i.v. injection of tumor cells. Such models were obstructive jaundice, and large abdominal and scrotal unusual in that cells remained dormant in various organs hernias. until stimulated to grow by estrogen. Transplants of estro gen-induced mammary carcinomas were hormone depend ent, and such models could readily be maintained indefi DISCUSSION nitely, even though an abrupt spontaneous change might occasionally occur in a single tumor, leading to a rapidly This introductory paper not only describes the incidence growing autonomous scirrhous type of lesion. To produce a of spontaneous and estrogen-induced tumors in Nb rats but high incidence of mammary tumors, it was necessary to also includes a study of the ability of their transplants to commence estrogen treatment before the animal was 1 grow autonomously or in conditioned hosts. Although few month old. These results suggest that the induction of this tumors arose spontaneously in males, all were found to be disease in the rat is related to a cell change that occurs autonomous on transplant; in females with normal ovarian before puberty but becomes manifest in later life under function, 10 transplants of 26 spontaneous tumors were estrogen stimulation. It is possible that this model has some dependent. Mammary carcinoma appeared to be unusual, in characteristics that simulate changes in the developing that all 5 tumors found in normal females were autono vagina of the human embryo that may follow the ingestion mous. On the other hand, 90% of99 tumors that occurred in of estrogen by the mother during pregnancy but that do not estrogenized rats of either sex were hormone dependent become manifest as carcinoma until adolescence, when when transplanted. Transplants of autonomous tumors hormone levels are increasing (8). A carcinoma of the usually grew rapidly, in contrast to those that were hormone vagina, similar to such tumors reported in humans, was dependent. A number of pituitary tumors and a kidney noted in an untreated rat but was not transplanted. Two carcinoma represent a type of tumor whose growth was other vaginal tumors, however, found in estrogenized ani estrogen inhibited. The experimental use of tumors with mals, resembled sarcoma botryoides, a tumor occasionally

MARCH 1975 773

found in young girls. In 1 case transplants, although normal female rats, they would not grow in ovariectomized growing slowly without estrogen, were stimulated by treat or male animals. A Leydig cell carcinoma of the testis arose ment with the hormone. The usual tumors, however, found in a pelleted rat that, although estrogenized, gradually in the uterus and cervix were of the muscle. Slower-growing became androgenized from the secretion of the developing hormone-dependent tumors appeared to be leiomyomas, tumor ( I 8). Animals of either sex bearing transplants of this whereas autonomous tumors exhibited more mitotic figures, estrogen-dependent tumor have shown the same sex hor characteristic of sarcomatous change. In only I case was a mone-transformation effect. The development of large carcinoma of the cervix induced by estrogen, and its scrotal or femoral hernias was associated with the growth of transplants have continued to be hormone dependent. this tumor, similar to those reported in some strains of mice Tumors of the anterior pituitary can be induced readily with Leydig cell tumors (14). The same type of carcinoma by a number of agents and have been studied extensively by arising spontaneously in an old rat was autonomous on Furth et a!. (7). The gland appears to be particularly transplant and was not associated with androgen production sensitive to estrogens and continues to increase in weight [similar tumors in aged Fischer rats have been studied by through life only in female rats, suggesting that this change Jacobs and Huseby (9)]. The finding of other types of tu is related to normal ovarian function. Marked hypertrophy mors in a number of organs apparently associated with the leading to large chromophobe adenomas followed estrogen action of estrogen and androgen will be described in a sepa treatment of both sexes of rats. This was not associated with rate paper. biological evidence of increased secretion. The pituitary Tumors arising spontaneously in a number of other hypertrophy was accelerated by simultaneous treatment organs have been listed, and these were all autonomous on with pituitary STH (secreted by grafted STH-secreting transplant. Nephroblastoma or Wilms' tumor was the pituitary carcinomas). Adenoma formation was hormone common kidney lesion found in the Nb rats, although it is dependent, since the gland returned to normal size if apparently a rare tumor (24). estrogen treatment was discontinued (Table I). The initial The apparent involvement of estrogen in the induction of transplantation of pituitary adenomas, however, or those a large number of tumors of various organs and the induced in ectopic grafts of normal gland, was followed in dependence of their transplants on the same hormone for most cases by the development of aggressively growing optimal growth would appear to answer the polemic carcinomas (25), many of which secreted hormones. Some question of whether or not estrogens cause cancer. How of these became autonomous on sucessivetransplants, ever, any conclusion requires qualification, for, as has been although some retained hormone-dependent properties. A emphasized on many occasions, estrogens are not locally single spontaneous ACTH-secreting tumor in a male rat carcinogenic in the classic test of skin painting. Tissue was autonomous on transplant. Many of the transplanted damage of a premalignant nature is not caused by estrogens, pituitary tumors produced biological changes in the host, and tumors do not arise adjacent to the hormone pellet. In indicative of the secretion of ACTH or 5TH. Unlike the the experiments described, tumors did occur in target tissues common finding, reported by others, of prolactin effects in of the hormone such as the breast and pituitary, but other other strains of rats with similar tumors, the secretion of target organs such as the uterus, cervix, and vagina showed this hormone by tumors of Nb rats was seldom encountered. a low incidence of carcinoma. On the other hand, tumors Hormone secretion by transplanted tumors was not related under estrogen control were encountered in many organs to tumor growth dependency on estrogen and appeared to not thought to be influenced by the hormone. Such organs change haphazardly in successive generations. The continu as the salivary glands, pancreas, lymphatic system, cervix, ous secretion of pituitary hormones in some cases led to and breast, however, have frequently been suggested as sites severe pathological lesions, similar to ones found in some of viral carcinogenesis. human diseases. In the rat the signs of disease were In the results described, estrogens have been considered, apparently hormone dependent and disappeared if the for descriptive purposes. to act directly on tumor cell tumor was removed. induction or growth. Proof of this is lacking and, indeed, Tumors developed in estrogenized rats in a number of considerable evidence exists that the effect of estrogen may organs not usually believed to be under estrogen control. be indirect or may involve pituitary hormone stimulation. These included the salivary gland, glandular pancreas, Irrespective of its mode of action, however, the experiments lymphoid tissue, and epithelial and lipoid tissue of the would suggest possible mechanisms of tumorigenesis. thymus. In all cases some of the transplanted tumor lines Tumor induction might follow the effects of estrogens on were either estrogen dependent or stimulated by estrogen, dormant but potentially malignant cells that had survived a suggesting an implication of the hormone in the primary previous exposure to carcinogenic factors of viral, chemical, tumor induction (although in the case ofthe normal salivary or physical origin. Under the influence ofthe hormone, such glands, no evidence of a trophic action of estrogen could be cells might be stimulated to become tumors that are demonstrated). Spontaneously arising tumors in the same dependent on the same hormone requirement for sucessful organs were found in many cases; these tumors were usually growth of other transplants. In such a case estrogens might autonomous on transplant. Thecomas were the only tumors be considered as promoters of previously altered cells rather of the ovary found in 10 animals. Estrogen-dependent than primary carcinogens. The unexpected wide range of transplant lines apparently required very small amounts of organs affected would support such a concept. On the other estrogen for growth because, although they grew readily in hand, estrogen, when administered in the form of pellets

774 CANCER RESEARCH VOL.35

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 1975 American Association for Cancer Research. Hormone-dependent Rat Tumors allowing a constant released level of hormone over pro the Rat. I. Induction and Behaviour of Tumors. Cancer Res., 24: longed periods, may only be stimulating, in a random 1116â€”1123,1964. manner, clones of cells that are slightly more responsive to 6. Dunning, W. F. Strain Differences in Response to Estrone and Induction of Mammary Gland, Adrenal. and Bladder Cancer in Rats. the hormone. Through gradual division over the long latent Cancer Res.,7:511 521,1947. periods, such cells might eventually result in tumors that 7. Furth, J., Clifton. K. H., Gadsden, E. L., and Buffett, R. F. Dependent would be expected to retain dependency for growth on the and Autonotnous Mammotrophic Pituitary Tumors in Rats: Their hormone after transplant. Somatotrophic Features. Cancer Res., 16: 608-616. 1956. Irrespective of the basic mechanism of carcinogenesis, 8. Herbst, A. L., Ulfelder, H., and Poskanzer, D. C. Adenocarcinoma of however, the ultimate development and growth of certain the Vagina. New EngI. J. Med., 284: 878-881. 1971. cells into the tumors under discussion was controlled by 9. Jacobs, B. B., and Huseby. R. A. Neoplasms Occurring in Aged estrogen levels that could be adjusted experimentally and so, Fischer Rats with Special Reference to Testicular, Uterine and presumably, could be under hormone control by the host. Thyroid Tumors. J. NatI. Cancer Inst., 39: 3O3@3O9, 1967. The findings at present reinforce the studies on pituitary 10. Jellinck, P. H., and Woo, J. Changes in Oestrogen Metabolism tumors in 1956 by Furth et a!. (7) and, similarly, lead to and Induced in Rat Liver Microsomes by Subcutaneous Pellets of Estrone and Testosterone. Endocrinology. 39: 99- 104, 1967. expand the somewhat heretic conclusion that the initiation I I. MacKenzie, W. F., and Garner, F. M. Comparison of Neoplasms in and control of growth of various types of cancer of many Six Species of Rats. J. NatI. Cancer Inst., 50: 1243- 1258, 1973. organs may be markedly influenced by various factors that 12. Lemon, H. M. Abnormal Estrogen Metabolism and Tissue Estrogen reside in the host. Since this overall project now involves Receptor Proteins in Breast Cancer. Cancer, 25: 423-435, 1970. some 42 different types of tumors arising in at least 24 13. Nichols, T. M. The Morphological Effects of Estrogen Removal on an organs of rats, either spontaneously or under some type of Estrogen-dependent Adrenocortical Carcinoma in Rats. Cancer Res., hormone stimulation, it would suggest that a new look with 31: 1042-1050,1971. expanded horizons might be propitious, not only at the role 14. Noble, R. L. Tumors and Hormones. In: G. Pincus (ed.), The of hormones as a cause of tumor formation, but also as Hormones. Vol. 5, pp. 559â€”625. New York: Academic Press, Inc., 1964. factors that may be active in the host in the control of 15. Noble, R. L. Induced Transplantable Estrogen-dependent Carcinoma cancer. of the Adrenal Cortex in Rats. Proc. Am. Assoc. Cancer Res., 8: 51, 1967. 16. Noble, R. L. Biology of Experimental Hormone-dependent Tumors. ACKNOWLEDGMENTS Especially Mammary Carcinoma. Abhandl. Deut. Akad. Wiss. Berlin Kl. Chem. Geol. Biol., I: 21-28, 1967. Participants in this research during the period 1964 to 71 included 17. Noble, R. L. Estrogen-induced, Estrogen-dependent Mammary Can Valerie Parker, Margaret Birrell. Carol Spady, and Kathy Foster. Rilla cer in the Rat. Proc. Am. Assoc. Cancer Res., 12: 103, 1971. Hendry was responsible for the extensive histological preparations and Bill 18. Noble, R. L. An Estrogen-dependent Testicular Carcinoma ofthe Rat Siep was responsible for the microphotographs. Professor W. L. Dunn, Causing Unusual Androgen Activity. Proc. Am. Assoc. Cancer Res., Pathology Department, offered valuable assistance in tumor pathology. 13:121,1972. Dr. G. L. Slemmer has kindly criticized and assisted with the preparation 19. Noble, R. L. Permanent but Reversible Aspermatogenesis in the Rat of this paper. as a Sole Aftermath of Brief Exposure to Estrogen. Excerpta Med. Sect. Ill, p. 55, 1972. 20. Noble, R. L., and Collip. J. B. Regression of Estrogen-induced Mammary Tumors in Female Rats following Removal of the Stimu REFERENCES lus. Can. Med. Assoc. J., 44: 1-5, 1941. 21. Noble, R. 1., and Cutts, J. 1-I. Mammary Tumors of the Rat: A I. Clifton, K. H. Problems in Experimental Tumorigenesis of the Review. Cancer Res., 19: 1125-I 139, 1959. Pituitary Gland, Gonads, Adrenal Cortices and Mammary Glands: A 22. Noble, R. L., McEuen, C. S., and Collip, J. B. Mammary Tumors Review. Cancer Res., 19: 2@22, 1959. Produced in Rats by the Action of Oestrone Tablets. Can. Med. 2. Combs, J. W., and Purnell, D. M. Functional Characteristics of Mast Assoc.J.,42:413-417,1940. Cells Associated with Rat Mammary Tumors Induced by 7:12 23. Takewaki, K. Some Aspects of Hormonal Mechanism Involved in Dimethylbenz(a)anthracene. J. NatI. Cancer Inst., 50: 1003- 101 1, Persistent Estrus in the Rat. Experientia, 18: 1-48, 1962. 1973. 24. Tomashefsky, P., Furth, J., Lattimer, J. K., Tannenbaum, M., and 3. Cutts, J. H. Estrogen-induced Breast Cancer in the Rat. Can. Cancer Priestley, J. The Furth-Columbia Rat Wilms' Tumor. J. Urol., /07: Conf., 6: 50-68, 1966. 348-354, 1972. 4. Cutts, J. H. Vascular Lesions Resembling Polyarteritis Nodosa in Rats 25. Welsch, C. W., Jenkins, T., Amenomori, Y., and Meites, J. Tumorous Undergoing Prolonged Stimulation with Oestrogen. Brit. J. Exptl. Development of in Situ and Grafted Anterior Pituitaries in Female Pathol.. 47:401-404, 1966. Rats Treated with Diethylstilbestrol. Experientia, 27: 1350-1852, 5. Cutts, J. H., and Noble, R. L. Estrone-induced Mammary Tumor in 1971.

MARCH 1975 775

Figs. I to 24. Microphotographs of 20 types of tumors of 17 organs. These include 14 tumors arising in estrogenized hosts under the influence of estrogens and 8 corresponding spontaneously arising autonomous tumors. H & E, x 45, x I 13, or x 181. Fig. I. Primary early adrenal carcinoma (24 mg) arising in zona reticularis adjacent to medulla in female EP for 12 months. Fig. 2. Primary adrenal carcinoma (4 g) in a female 90% estriol + 10% cholesterol pellet for 14 months. Fig. 3. Primary mammary adenocarcinoma (125 mg) in male 90% estriol + 10% cholesterol pellet for I I months. Fig. 4. Primary spontaneous papillary mammary carcinoma (3 g) in 3-month-old male. Autonomous on transplant. Fig. 5. Pituitary carcinoma (with 5TH effect). Second transplant (dependent) oftissue (70 mg), arising in female EP for 10 months from ectopic graft. Fig. 6. Pituitary carcinoma. Eleventh transplant (autonomous) (with 5TH effect and anaplasia) ofprimary tumor (66 mg). arising in female EP for 12 months. Fig. 7. Primary ovarian thecoma ( I.2 g) arising after EP for 9 months. Transplants under study. Fig. 8. Leiomyoma of uterus. Fourth transplant (dependent) of primary tumor (200 mg) arising after EP for 15 months. Fig.9. Leiomyosarcoma of uterus. Fourth transplant (autonomous) of primary tumor (29 g) arising spontaneously in 8-month-old rat. Fig. 10. Adenocarcinoma of cervix. Thirty-first transplant (dependent) of primary tumor (0.3 g) arising after EP for 2.5 months. Fig. II. Primary fibrosarcoma of vagina (8 g) arising after EP for 10 months. Autonomous on transplant but stimulated by estrogen. Fig. 12. Primary carcinoma of vagina (16 g) arising after EP for 15 months. Not transplanted. Fig. 13. Primary interstitial (Leydig) cell carcinoma of the testis (8 g) arising after EP for 13 months. Dependent on transplant. Fig. 14. Primary spontaneous interstitial (Leydig) cell carcinoma of the testis (3 g) in 22-month-old male. Transplants under study. Fig. IS. Carcinoma of thymus. Fourth transplant (dependent) of primary tumor (7 g) arising in female EP for 12 months. Fig. 16. Primary spontaneous carcinoma of thymus (5 g) in male of 16 months. Autonomous on transplant. Fig. 17. Liposarcoma of thymus. Fifth transplant (dependent) of primary tumor ( 10 g) arising in 9-month-old female. inset: frozen section stained oil red0. Fig. 18. Fibroadenoma of gland of orbit. Second transplant (dependent) of primary tumor (0.7 g) arising in female EP for 7 months. Fig. 19. Carcinoma of submaxillary gland. Seventh transplant (dependent) of primary tumor (3 g) arising in female EP for I I months. Fig. 20. Primary spontaneous carcinoma of submaxillary gland (3.2 g) in 5-month-old male. Autonomous on transplant. Fig. 2 1. Primary adenocarcinoma of pancreas in female EP for 9 months. Dependent on transplant. Fig. 22. Primary adenocarcinoma of pancreas in female rat EP for 12 months. Autonomous on transplant. Fig. 23. Lymphoma, reticulum cell. Second transplant (estrogen-stimulated) showing kidney invasion from primary tumor presenting as enlarged lymph nodes (7.7 g) and thymus (4.9 g) in male EP for 9 months. Fig. 24. Lymphoma. reticulum cell. First transplant (autonomous) showing spleen invasion with giant cells from primary tumor presenting as enlarged spleen (8 g) and liver (42 g) in male EP for 5 months.

776 CANCER RESEARCH VOL.35

- -.

@ ,-(,@_. 1(j@.

,@I ,@. â€¢4I

@ M\R(II

R. L. Nob!e el a!.

6@ i: @@ - F ,,,- .- f. . .@â€”.. 4 / ,, -_â€¢@_@_1_',â€¢ :. . . @a â€˜ . .,,__4. -@ â€¢_â€˜â€˜â€¢, -â€” .: @ :, â€¢,, ,@; r@, â€˜@, â€˜I,,.., ...J_, ..@, - â€” @.. _._d. @ ..._v-, .frÃ¸:.c2,;,,..bv@.#: â€˜- @. . @, . .. ;...... â€˜@,â€˜ .6 - @ . a- ,,i'@@'-â€¢- @. @ . a @@ â€˜: â€¢ @ .. - . [email protected] ,@ â€˜â€¢.â€˜.â€˜@,t@â€˜F,..@â€˜ . â€˜ ,,. , , ,@t.. .@ @%.-. . @ @â€œ@â€˜t'. . -, I

@ .@-.,, .# @: @1Y 44 ,J,P @ -a ,@ 1â€•@ C. @ . â€” .1_-.-.@._.. , @. C .I

@ .... @@._:.â€œ.@â€¢:.;6:.':i, -,â€˜

@@@@ I' 6 , @@@@ :@: :@@ ;â€˜ ;t@6&.lbf@ .@â€˜L1@':@ I J@Q@

@ %

L@ a@ @ a V @1 .4 b p @- â€˜ . @ â€œ â€œSi

ffr

O_'_p â€¢ [email protected]@

@ @-@Sh -

@ @L . ,. [email protected] @ p. ,, @- t 1. â€˜\.@ ,. ;i@;'ir. .@@ @I's,m@ I@%@

@@@ FI.I -,-. .1@â€¢

@@ 1' @#â€˜ â€œ ..\ @@@ :â€˜ w .â€˜â€˜ @@ â€¢1 â€œ , I' : .â€˜c A . . . C â€˜ 1' â€˜-â€œI.:@.:t.@;:@.:.

â€œ 6 .

@ .; C , .4

@ â€˜ : , :@ -

778 CANCER RESEARCH VOL.35

@vl.\RCII V)7@s 779

R. L. Noble et a!.

@ .â€˜:- @1 @ :@ ...... â€˜.; [email protected]...... 5 5 ..â€˜@-Jr@@@ @-@r : -@ @@ C ,, .., . . % â€˜ . .: : â€˜a

- @â€˜ @@@@@ .: @â€¢v4- .@ â€˜ â€¢ . S â€¢ @@@@@@@@@@@ .. . . .b@4' â€˜P.,. â€¢. C@ . .: :@ @ â€¢@ . -. 11@@ â€˜@;.@$ @. -. , @@@@@ @..C. #â€˜@ ,:â€˜ -â€˜ @. @.. @@@@ *@ â€˜I, â€œ--a : @:

@ _.@,.; â€¢S..4@ S @ . . .. - .5. . .. .5 .@ . . .. @-a. @@ .â€¢â€˜@.â€˜J)I @â€¢ ., @@@@@@ .. @â€˜. , . S ,@ â€¢ . : 7.@ @@@ â€¢-,-: â€˜ @. .. â€˜ :

@@@ . . , . . :@ . : -. @.. :â€¢

@ :@ @.. @ . .@ .1'-' â€˜ z..J:

@@@ â€˜ . ,, ,@- : :::a@

!â€˜Ã˜s 5

@ â€˜. - --â€˜.5

\. @ . @ .1 p.'. -S. ;i@ â€œ .@k -â€˜4:@.4

a-a @ . , 51

.@â€˜

@@ .t@@}Tdu!S 4. a' @ 1. ., .@ â€¢.@

SS @@ @55/ I,

@ . ,- . .@ . ( @ p

@@ . @55 S @ .1 - , .@_._S;'. @ 4r&@7@[email protected]'[email protected] @4. @s':.I,w@ @ .- :@â€˜)_.â€œâ€”

780 CANCER RESEARCH VOL.35

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 1975 American Association for Cancer Research. Spontaneous and Estrogen-produced Tumors in Nb Rats and Their Behavior after Transplantation

Robert L. Noble, Brenda Clayton Hochachka and Diane King

Cancer Res 1975;35:766-780.

Updated version Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/35/3/766

E-mail alerts Sign up to receive free email-alerts related to this article or journal.

Reprints and To order reprints of this article or to subscribe to the journal, contact the AACR Publications Subscriptions Department at [email protected].

Permissions To request permission to re-use all or part of this article, use this link http://cancerres.aacrjournals.org/content/35/3/766. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC) Rightslink site.