[CANCER RESEARCH 34, 3018—3025,November 1974]

SUMMARY

The effect of perphenazine, a phenothiazine derivative, onserumprolactin levels,on maturation of the normalmammae,and on growth of mammary adenocarcinomas was studiedusing two transplantable rat mammary tumor systems, the13762 mammary tumor (13762 MT) in the syngeneic Fischer344 strain and the R-35 mammary tumor (R-35 MT) inallogeneic Sprague-Dawley stock. Perphenazine induced adose-response increase in the serum prolactin levels of bothnormal and tumor-bearing animals and stimulated maturationof the normal mammae. The resultant responses of the twomammary tumors to perphenazine treatment were in exactcontrast. Growth of the 13762 MT was accelerated whereasgrowth of the R-35 MT was inhibited. There was atrophy ofthe mammae and a concomitant decrease in serum prolactinlevels in young, sexually maturing females with progressivelygrowing 13762 MT tumors. Normal mammae and serumprolactin levels were unaffected in similar females bearing theR-35 MT. Treatment of the hosts with estrogen prior to tumorimplantation, as well as treatment of both tumor and host,enhanced the effects of perphenazine on serum prolactinlevels, on the normal mammae, and on mammary tumorgrowth, i.e., stimulation of 13762 MT and inhibition of R-35MT.

INTRODUCFION

Phenothiazine derivatives have been reported to inducenormal mammary growth and initiate milk secretions inhumans and in a variety of animals (3, 10, 12, 15). Ben-David(I) has recently shown that the perphenazine-mducedmammary development in intact rats is associated withstimulation of pituitary prolactin secretion. Danon et a!. (6)demonstrated that both intact females and recently spayedrats respond to perphenazine treatment with lobuloalveolardifferentiation of the mammae. On the other hand, when therats had been ovariectomized 12 days previously, perphenazinewas only weakly mammotrophic. In the latter case primingwith sufficient amounts of estrogen (8 pg/day) completely

1 This investigation was supported by Contract PH43-65-6 from the

Endocrine Evaluation Branch, General Laboratories and Clinics,National Cancer Institute, NIH.

Received December 14, 1972; accepted August 2, 1974.

restored the mammotrophic response to perphenazine andproduced lactation. That estrogen apparently acts on thehypothalamic-pituitary system to stimulate the secretion ofprolactin was shown both in vivo by Kanematsu and Sawyer(8) and in vitro by Nicoll and Meites (1 1) and by Ben-David eta!. (2). The results obtained by Danon et a!. (6) suggest thatprolactin secretion cannot be induced when pituitary secretionof gonadotrophins is high and that inhibition of gonadotrophm secretion by estrogen stimulates or potentiates thesecretion of prolactin.

Hilt et a!. (7) studied the effect of fluphenazine HC1, a

phenothiazine derivative, on growth of the R3230ACmammary adenocarcinoma and mammary glands of the ratand reported that, while administration of this tranquilizerstimulated normal mammary glands, it caused a decrease in thegrowth of R3230AC .tumor. They concluded that creating ahormonal milieu that stimulates normal mammary glands doesnot necessarily stimulate growth of mammary tumors. Thewell-recognized variability in the hormone responsiveness ofmammary tumors, both in animals and in man, raised thequestion concerning the sensitivity of the 13762 and R-35MT's2 to prolactin. These tumors have been widely used forchemotherapeutic studies (13, 14) as well as for studiesdesigned to evaluate therapy combinations (5). The 13762MT, originally induced with dimethylbenzanthracene, is anextremely useful experimental mammary tumor model with apredictable (almost 100%) incidence of lung and organmetastases from s.c.-implanted grafts and was found to be verysensitive to the oncolytic effects of the steroidal alkylatingagents (13, 14). The R-35 MT, a subline of the Hugginsfibroadenoma A, on the other hand, has a relatively lowincidence of metastases and, although it is responsive, it lacksthe same degree of sensitivity to the steroidal alkylatingagents.

This study was designed, therefore, to determine not onlythe effects of enhanced prolactin secretion, induced byperphenazine, on growth of the 2 mammary adenocarcmomasper Se, but also the effect of mammary tumor growth onprolactin-normal mammary tissue interaction. Quantitation ofserum prolactin levels by means of radioimmunoassay haspermitted the comparison of prolactin levels in the variousexperimental groups. The experimental design incorporates,therefore, estrogen priming of host prior to tumor implanta

2Theabbreviationusedis:MT.mammarytumor.

3018 CANCER RESEARCH VOL.34

The Effect of Perphenazine-induced Serum Prolactin Responseon Estrogen-primed Mammary Tumor-Host Systems, 13762and R-35 Mammary 1

Arthur E. Bogden,D. JaneTaylor, EricV. H. Kuo,MarcusM. Mason,andAnastasiaSperopoulos

Department of Immunobiology, Mason Research Institute, Worcester, Massachesetts 01608 [A. E. B., E. V. H. K., M. M. M., A. S.J , and EndocrineEvaluation Branch, Division ofCancer Biology and Diagnosis, National Cancerlnstitute, NIH, Bethesda, Maryland 20014 [D. J. T.J

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TreatmentPerphenazine

dose levels(mg/kg/day)Groupa-Study A:estrogen priming of hostStudy B:estrogen priming of tumor andhostITreated

with 0.5 ml sesame oil,1 dose/day for 10 days. Givenimplant of tumor tissue on Day11. Treatment with 0.5 ml0.03 N HCI initiated on Day 11,1 dose/day, continued to survivaltime.Given

implant of tumor tissue on Day 1.Treatment with 0.5 ml sesame oilinitiated on Day 9, 1 dose/day for 10days. Treatment with 0.5 ml 0.03 N HC1initiated on Day 19, 1 dose/day, continuedto survivaltime.IITreated

with 0.5 ml sesame oil,1 dose/day for 10 days. Givenimplant of tumor tissue on Day11. Treatment with perphenazineinitiated on Day 1 1, 1 dose/day,to survival time.Given

implant of tumor tissue on Day 1.Treatment with 0.5 ml sesame oilinitiated on Day 9, 1 dose/day for 10days. Treatment with perphenazineinitiated on Day 19, 1 dose/day, tosurvivaltime.2III4IV8VTreated

with 8 @.&gestradiol-1713,once each day for 10 days. Givenimplant of tumor tissue on Day 11.Treatment with perphenazineinitiated on Day 11, 1 dose/day,to survival lime.Given

implant of tumor tissue on Day 1.Treatment with 8 @gestradiol-17 @3initiatedon Day 9, 1 dose/day for 10 days. Treatment with perphenazine initiated on Day19, 1 dose/day, to survivaltime.2VI4VII8VIIITreated

with 8 @gestradiol-1 7@, 1dose/day for 10 days. Treatmentwith perphenazine initiated onDay 11, 1 dose/day, to survivaltime. No implant.Treatment

with 8 @sgestradiol-17 @3initiatedon Day 9, 1 dose/day for 10 days. Treatment with perphenazine initiated on Day19, 1 dose/day to survival time. Noimplant.2IX4x8

Prolactin and Mammary Tumor-Host Interaction

routine 1- to 2-cu-mm grafts of the syngeneic 13762 MT.R-35 MT. Forty-five to 50-day-old SCH:ARS(SD) stock

females were divided into the various experimental groups asshown in the basic experimental design outlined in Table 1 andwere maintained on Wayne Lab Blox and tap water ad libitum.The designated groups were given s.c. implants on the rightside of a routine Stadie slice of the allogeneic R-35 MT.Although R-35 MT originated and is transplantable inSprague-Dawley-derived stock, grafts implanted s.c. are lethalto only 85% of the animals. Therefore, animals with no takesor with growth followed by regression were deleted from thisstudy.

Drug Preparation and Administration. Estrogen treatmentconsisted of 8pg estradiol-l7@3 in sesame oil injected s.c. intothe nape of the neck daily for 10 days. Perphenazine (NSC150866, generously donated by the Research Division, SheringCorp., Bloomfield, N. J.) at 3 dose levels, 2, 4, and 8 mg/kg,

Table 1

tion as well as estrogen priming of host and tumor. In view ofthe apparent competition of mammary tumors with normaluteri for endogenousestrogensdemonstratedby Bogdenet al.(4), it was felt that the administration of the exogenous

estrogen would provide the excess of estrogen necessary notonly to permit maximal sensitization of both normal andneoplastic tissues to prolactin effects, but also to potentiatethe response of the pituitary to perphenazine.

MATERIALS AND METHODS

13762 MT. Forty-five to 50-day-old Fischer 344/CRLfemales were divided into the various experimental groups asshown in the experimental design outlined in Table 1 and weremaintained on Wayne Lab Blox and tap water ad libitum. Thedesignated groups were given s.c. implants on the right side of

Experimental design: 13762 MT system

a Groups contain 6 to 10 rats at eachsacrifice time.

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dissolved in 0.03 N HC1 in a volume of 0.2 ml, was injected s.c.in the left side. No injections were given in the right side wherethe tumor was to be implanted. All animals were maintained2/cage to minimize stress of crowding in a room that hadcontrolled light and dark cycles of 12 hr.

Experimental Design.The overallstudy wasdivided into 4experimental segments, i.e. , a Study A and a Study B for eachtumor system. Although the experimental protocols andtreatment regimens were designed specifically for each tumorsystem , being based upon the lag time and growth ratecharacteristic of each tumor, they are, nonetheless,fundamentally identical in design so that valid correlations andcomparisons between the 2 mammary tumors can be made.The experimental design for the 13762 MT is summarized inTable 1. In Study A, the hosts were treated with estrogen priorto implantation of tumor grafts. Perphenazine treatment wasinitiated on Day 1 of tumor implantation and continued dailythrough survival time. In Study B, both tumor and host weretreated with estrogen. Therefore, tumor grafts were implantedon Day 1 and estrogen treatment was not initiated until thegrafts showed 1st evidence of palpable growth. Perphenazinetreatment was initiated after 8 or 10 days of estrogenadministration and when the tumors were in mid-log phase ofmeasurable growth.

Radioimmunoassay of Serum Prolactin. Animals weresacrificed in early morning, solid food having been removedfrom the cages at 5 p.m. on the previous day leaving onlywater ad libitum. On the day of sacrifice, animals wereremoved from their cages, 1 at a time, with minimal noise andagitation and quickly decapitated. Blood was collected in smallsterile funnels, allowed to run into 10-ml test tubes, and storedovernight at 5°for clot retraction before being processed forserum. Serum prolactin levels were determined on duplicateserum samples at 2 dilutions, using reagents provided in theNational Institute of Arthritis and Metabolic Diseases radioimmunoassay kit for rat prolactin (NIAMD-rat prolactinand antisera were generously provided by the HormoneDistribution Office, National Institute of Arthritis andMetabolic Diseases, NIH), according to the procedure reportedelsewhere (9).

Organ Weight End Points. Recording of the initial bodyweight and the final body weight after tumor excisionpermitted organ weight determinations based upon 100 g offInal body weight for the following organs: ovaries, uterus,liver, spleen, kidney, thymus, adrenals, and pituitary. Tumorweight was determined at sacrifice.

Histology of the Mamm@. At autopsy the left inguinalmammary pad was removed along with the overlying skin,pinned on flat cardboard, fixed in formalin, sectioned, stainedwith hematoxylin and eosin, and examined for mammary ductgrowth, growth of the lobuloalveolar system, and for evidenceof secretion.

Mammary tumor development was graded histologically ona numerical basis, i.e., 1+, 2+, etc., with the highest number(4+) representing the highest degree of mammarydevelopment. Examinations were made of coded slides to aidin objectivity, and the average for each experimental groupwas reported as a numerical index that is an indication ofrelative mammary development.

RESULTS

The effect of perphenazine on serum prolactin levels, onorgan weights, on the normal mammae, and on mammarytumor growth was studied in an experimental design thatpermitted evaluation of the effect on mammary tumor growthper se as well as on the interaction of mammary tumor growthand perphenazine on the development of the normal mammae.Since 2 different transplantable mammary tumor systems wereused, the results obtained with each tumor system arepresented separately. Significant differences and similaritiesare stressed in “Discussion.―

Serum Prolactin Response to Perphenazine Treatment. Thedose response of serum prolactin levels to perphenazine inFischer 344/CRL rats is illustrated in Chart 1. Thepotentiation of the pituitary for prolactin release by estrogentreatment is also indicated by the dose-response curve, whichshows a consistently higher prolactin response at all dose levelsin estrogen-treated animals. The serum prolactin response toperphenazine in normal rats pretreated with estrogen indicatedthat estrogen potentiation of the pituitary declined with timefollowing the termination of estrogen treatment. There was aconsistently lower response in serum prolactin levels toperphenazine at all 3 dose levels at 35 days after estrogentreatment than at 21 days (Chart 2).

Serum Prolactin and Normal Mammary Development inFischer 344 Females. The correlation of serum prolactin levels

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Chart 1. The dose response of serum prolactin levels to perphenazmein Fischer 344/CRL rats and potentiation of the response by estrogentreatment. Mean ±S.D.

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Prolactin and Mammary Tumor-Host Interaction

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the day of tumor implantation. The prolactin responses onDay 21 , when the tumors were relatively small, wereperphenazine-dose related, and the subsequent rates of tumorgrowth were definitely related to serum prolactin levels. At the8-mg/kg dose level of perphenazine, which induced the highestserum prolactin levels on Day 21 and, consequently, thelargest tumors by Day 35, serum prolactin levels were lower onDay 35 than on Day 21 . This is interpreted as possibleprolactin “binding―by the tumor occurring during maximalprolactin response with a consequent depletion of serumprolactin levels.

Effect of 13762 MT Growth on the Mammary Glands of the

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pretreated with estrogen illustrating the decline of the potentiatingeffect with time.

and development of the normal mammae inperphenazine-treated animals is illustrated in Chart 3. Acomparison of the mammary development index and serumprolactin levels of perphenazine-untreated, I 3762 MT-bearingwith those of the perphenazine-treated groups revealed thatperphenazine at the higher dose levels not only increasedserum prolactin levels but also counteracted the atrophiceffects on the normal mammary glands induced by mammarytumor growth.

Serum Prolactin Response and 13762 MT Growth. Thestimulatory effect of perphenazine treatment on the growth ofthe 13762 MT is shown in Chart 4. In Study A (Chart 4)perphenazine treatment stimulated mammary tumor growthonly at the highest dose level. When the host was givenestrogen prior to tumor implantation and initiation ofperphenazine treatment, tumor growth was enhanced at alldose levels. Stimulation of tumor growth in perphenazinetreated animals was also evident in Study B, although it wasnot as marked as in Study A. However, all dose levels ofperphenazine appeared to be equally stimulatory, primarilywhen both host and tumor were treated with estrogen.Perphenazine treatment in Study A was initiated in early loggrowth phase and lasted for 25 days, whereas in Study Bperphenazine treatment was initiated in mid-log phase and hada duration of only 18 days.

That the rate of tumor growth in the experimental groupsgiven perphenazine was related to the resultant prolactin levelsis shown in Chart 5. Perphenazine treatment was initiated on

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Chart 5. Relationship of the rate of 13762 MT growth and serumprolactin levels in perphenazine-treated animals.

Sexually Maturing Rat. In view of the effect of prolactin ongrowth of the 13762 MT and the indication of a possible“prolactin binding― by this tumor, the effect of tumor growthon the normal mammae of the sexually maturing rat wasstudied in the vehicle-treated control groups. Chart 6summarizes the atrophic effects of mammary tumor growthobtained in the vehicle-treated controls of Studies A and B andillustrates that the atrophy of the mammae was directlyrelated to progressive tumor growth and to a decrease in serumprolactin levels.

Serum Prolactin and Normal Mammary Development inSprague-Dawley Stock Females. Chart 7 illustrates a typicaldose response of serum prolactin levels induced in normalSprague-Dawley-derived stock females by perphenazine, andthe direct relationship of serum prolactin levels with development of the normal mammae. Potentiation of the pituitary forprolactin release by estrogen was also seen in this stock ofanimals, and it was evident throughout the study that higherdoses of perphenazine not only increased serum prolactinlevels but also stimulated development of the normal mammaein tumor-bearing animals.

Serum Prolactin Response and R-35 MT Growth. The effectof perphenazine on growth of the R-35 MT is shown in Chart8 and is in contrast to the results obtained in the 13762 MTsystem. Growth of the R-35 MT tended to be inhibited inperphenazine-treated animals. When the host was givenestrogen prior to tumor implantation and perphenazinetreatment, there was a perphenazine dose-related inhibition oftumor growth. That the tumor inhibitory effect was related toserum prolactin levels is illustrated in Chart 9. The slopes ofthe prolactin curves indicate that serum prolactin levelscontinued to increase with time despite progressive growth ofthe tumors. There was no tendency for the R-35 MTmammary tumor to lower prolactin levels as had been seen inthe 13762 MT system.

Effect of R-35 MT Growth on the Mammary Glands of theSexually Maturing Rat. The absence of an effect on serumprolactin levels by the R-35 MT, as indicated in the previousparagraph, is supported by the uninhibited development of thenormal mammae in rats with progressively growing mammarytumors (Chart 10). This is in sharp contrast to the atrophy ofthe mammae seen in rats bearing the 13762 MT (Chart 6).

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Weights. Thymolysis and adrenal hypertrophy resulting fromthe stress of tumor growth was evident in all tumor-bearinggroups and was related to tumor size rather than to treatment

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Study B, there was a perphenazine-induced, dose-effectincrease in ovarian weight on Sacrifice Day 37.

In the 13762 MT system , uterine atrophy was evident in alltumor-bearing groups, even in the estrogen-treated animals.The somewhat greater atrophy in animals treated only withperphenazine as compared to the vehicle controls can beattributed to the relatively larger tumors. Estrogen- andperphenazine-treated animals had the largest tumors bututerine atrophy on SacrificeDay 37 was no greater than theother experimental groups. Alleviation of some of the atrophiceffects seen in tumor-bearing animals, therefore, can beattributed primarily to the estrogen administration rather thanthe subsequent treatment with perphenazine. With the R-35MT system, on the other hand, there was no consistentatrophic effect on the uterus in animals with progressivelygrowing tumors. An estrogen effect on uterus weight wasevident in both tumor-bearing and non-tumor-bearing animals.

Prolactin and Mamma@y Tumor-Host Interaction

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with perphenazine and/or estrogen. When calculated on a100-g final body weight basis there was an increase in thekidney weight of tumor-bearing animals that paralleled tumorgrowth. Estradiol and perphenazine had no effect on kidneyweight in the non-tumor-bearing animal.

There was an increase in the ovarian weight of 13762MT-bearing animals that appears to be associated with tumorgrowth in perphenazine as well as perphenazine- and estrogentreated animals. There were no apparent effects of perphenazine or estrogen on the ovaries of non-tumor-bearing animals.In the R-35 MT system there was no significant or consistenteffect on ovarian weight resulting either from tumor growth orfrom treatmentwith estrogenand/or perphenazinein Study A.However, in the estrogen-treated, tumor-bearing animals of

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In the 13762 MT system there was a direct relationshipbetween increase in pituitary weight and tumor size in allexperimental groups except those treated with estrogen.Estrogen administration appeared to increase pituitary weightsin both tumor-bearing and non-tumor-bearing animals. Theresponse of the pituitary in Sprague-Dawley stock rats bearingthe R.35 MT was similar to the 13762 MT system.

DISCUSSION

The effectiveness of perphenazine to stimulate mammarygrowth in the young, sexually maturing female rat has beendemonstrated in this study, supporting the reports of similaractivity in humans and in a variety of other animals (3, 10, 12,15). Of particular significance was the dose response of serumprolactin levels to perphenazine treatment and the relationshipof 13762 MT growth to the serum prolactin response. There islittle question that increasing serum prolactin levels stimulatedboth normal mammae, in terms of functional maturation, andmalignant mammary tissue, in terms of proliferation. Thatnormal mammary tissue and malignant tissue can compete forthe available endogenous prolactin was indicated by theresponse of normal mammae and carcinomatous mammarytissue to prolactin, by the atrophy of normal mammary tissueand concomitant decrease in serum prolactin levels occurringin untreated animals bearing progressively growing 13762MT's, and by the finding that the highest dose of perphenazinewas able to abrogate this atrophic effect in tumor-bearinganimals.

In sharp contrast to these observations was the inhibition ofR-35 MT growth associated with increased serum prolactinlevels. Significantly, therefore, there was no detectableimpairment of maturation of the mammae or effect on serumprolactin levels in rats bearing progressively growing R-35MT's.

Inhibition of tumor growth by increased serum prolactinlevels was reported by Hilf et a!. (7) for the R3230ACmammary tumor. Hilf also reported that uterine weight wassignificantly reduced in R3230AC tumor-bearing animalsreceiving fluphenazine HC1. In the present study on both theR-35 and 13762 MT systems, no consistent atrophic effect onthe uterus was seen that could be attributed to perphenazinetreatment. Uterine atrophy in young, sexually maturingfemales, bearing progressively growing 13762 MT's has beenreported by our group (4) and also was evidenced in this studyin vehicle-treated controls bearing the 13762 MT but not theR-35 MT. It would seem important to determine whether theestrogen-binding capacity of mammary tumors is related to thesensitivity and type of response induced by prolactin.

Treatment of the host with estrogen prior to tumorimplantation, as well as treatment of both tumor and host,enhanced the effects of perphenazine treatment on both thenormal mammae and the mammary tumors. Since themammotrophic effects appear to be due, primarily, to theresultant increase in serum prolactin levels, the suggestion byDanon et a!. (6) that estrogen treatment potentiates thesecretion of prolactin by inhibiting gonadotrophin secretion issupported. Action of estrogen on the hypothalamic-pituitary

system to “stimulate―the secretion of prolactin was shown invivo by Kanematsu and Sawyer (8) and in vitro by Nicoll andMeites (1 1) and by Ben-David et a!. (2). The results of thepresent study provided evidence that the enhanced prolactinresponse resulting from estrogen treatment persists, althoughdecreasing, with time.

Thus, the primary objectives of this study to determine theeffect of enhanced prolactin secretion on mammary tumorgrowth per se and of mammary tumor growth on prolactinnormal mammary tissue interaction have been achieved. Forpossible use as contrasting test systems in chemotherapystudies as well as for evaluating combinations of therapy, it isimportant that, whereas growth of the 13762 MT, which haspredictable metastasizing characteristics, was shown to bestimulated by prolactin in these studies, growth of the R-35MT was found to be inhibited. Both of these tumor systemshave stable and predictable transplantation characteristics, asdoes the R3230AC mammary adenocarcinoma. It is clear thattransplantable mammary tumors show the variability ofresponse to prolactin that is reflected clinically and that arealistic preclinical evaluation of chemotherapeutic agents andtherapeutic modalities, designed for the treatment of breastcancer, must include a spectrum of well-defined mammarytumor models.

Of particular clinical importance was the finding thatmammary tumor growth could be stimulated by prolactin.Since perphenazine treatment increased serum prolactin levels,caution in the use of phenothiazine derivatives for tranquiization in pre- or postoperative breast cancer cases is suggested.

REFERENCES

1. Ben-David, M. Mechanism of Induction of Mammary Differentiation in Sprague-DawleyFemale Rats by Perphenazine. Endocrinology,83: 1217—1223,1968.

2. Ben-David, M., Dikstein, S., and Sulman, F. C. The Effect ofDifferent Steroids on Prolactin Secretion in Pituitary-Hypothalamus Organ Co-culture. Proc. Soc. ExptL Biol. Med., 117:511—514,1964.

3. Ben-David, M., Dikstein, S., and Sulman, F. G. Production ofLactation by Non-SedativePhenothiazineDerivatives.Proc. Soc.Exptl. Biol. Med., 118: 265—270,1965.

4. Bogden, A. E., Taylor, D. J., Esber, H. J. and Menmnger, F. F., Jr.Anti-uterotrophic Effect of Mammary Tumor Growth. Proc. Am.Assoc.CancerRes.,12:55,1971.

5. Bogden, A. E., Taylor, D. J., Esber, H. J., and Menninger,F. F., Jr.Effect of Surgery, Chemotherapy and Immunotherapy, Alone andin Combination, on Metastasesof the 13762 MammaryAdenocarcinoma. Proc. Am. Assoc. Cancer Res., 13: 76, 1972.

6. Danon, A., Weller,C. P., and Sulman, F. G. Role of EstrogensandGonadotrophins in Perphenazine-Induced Mammogenesis. J. Endocrinol.,48: 365—371, 1970.

7. Hill, R., Bell, C., Goldenberg, H., and Michel, I. Effect ofFluphenazine HC1 on R3230AC Mammary Carcinoma and Mammazy Glands of the Rat. Cancer Res., 31: 1111—1117, 1971.

8. Kanematsu, S., and Sawyer, C. H. Effects of Intrahypothalamicand Intrahypophyseal Estrogen Implants on Pituitary Prolactinand Lactation in the Rabbit. Endocrinology, 72: 243—252, 1963.

9. Kuo, E. Y. H., and Gala, R. R. Radioimmunoassay of Rat ProlactinComparing Prolactin Obtained from Anterior Pituitary OrganCulture with that Distributed by the National Institute of Arthritis

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1974;34:3018-3025. Cancer Res   Arthur E. Bogden, D. Jane Taylor, Eric Y. H. Kuo, et al.   R-35 Mammary Adenocarcinomason Estrogen-primed Mammary Tumor-Host Systems, 13762 and The Effect of Perphenazine-induced Serum Prolactin Response

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