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Suppression Ofautochthonous Grafts Ofspontaneous Mammary Tumor by Induced Allogeneic Graft Rejection Mechanism

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Suppression Ofautochthonous Grafts Ofspontaneous Mammary Tumor by Induced Allogeneic Graft Rejection Mechanism [CANCER RESEARCH 33, 645-647, April 1973] Suppression of Autochthonous Grafts of Spontaneous Mammary Tumor by Induced Allogeneic Graft Rejection Mechanism Reiko Tokuzen and Waro Nakahara National Cancer Center Research Institute, Tsukiji 5-chome, Chuo-ku, Tokyo, Japan SUMMARY existing between the allogeneic and autochthonous grafts are involved in determining the different susceptibilities to the Autochthonous grafts of spontaneous mammary adenocar- host-mediated antitumor effect of plant polysaccharides. cinoma of mice give 100% takes with no spontaneous More recently, we examined local cellular reactions around regression. However, the growth of these autografts was Sarcoma 180 grafts during the process of regression under markedly inhibited with a high rate of complete regression polysaccharide treatment, and we found a massive outpouring when the grafts were mixed with ascitic Sarcoma 180 and of lymphoid cells 1 week after tumor implantation. No such implanted under i.p. treatment with lentinan. Lentinan is one cellular reaction was noted in untreated controls or in mice of the so-called antitumor-active plant polysaccharides; it treated with inactive polysaccharides. Similarly, autoch strongly inhibits Sarcoma 180 but has no effect on thonous grafts of spontaneous mammary tumors, not at all autochthonous grafts. Without the lentinan treatment, the inhibited by polysaccharides, called forth no local lymphoid mixed autografts and Sarcoma 180 or Sarcoma 180 alone cell infiltration. constantly produced large tumors. These results indicate that a The question arose as to what would happen to autoch local rejection mechanism against allogeneic tumor (Sarcoma thonous grafts if such local cell reaction were induced around 180), induced by injections of lentinan, failed to recognize them as in that which occurred around Sarcoma 180 under autografts as "self and destroyed them along with the polysaccharide treatment. In the experiments presented in this allogeneic transplants. paper, we attempted to answer this question by implanting The frequency of local tumor recurrence after surgical autochthonous grafts mixed with Sarcoma 180, followed by removal of the primary tumors and the development of new the usual polysaccharide treatments. The critical point in this tumors in other parts of the mammary gland suggest that the experiment was whether or not the induced allogeneic graft rejection of autografts might be a strictly local process and rejection mechanism could discriminate the autochthonous that it does not induce adequate systemic resistance against cells from the cells of allogeneic Sarcoma 180. Somewhat to the mammary tumor. our surprise, when autochthonous tumor grafts were mixed with Sarcoma 180 and implanted, they were suppressed to a great extent by subsequent polysaccharide treatment. INTRODUCTION Intensive studies over the past several years have disclosed MATERIALS AND METHODS that injections of certain plant polysaccharides induced the state of resistance to implantation of certain allogeneic Spontaneous mammary adenocarcinomas were taken from tumors, particularly Sarcoma 180 in mice. The antitumor- female Swiss albino mice, about 5 to 9 months old. We have active polysaccarides have been isolated as potent prepara never observed any spontaneous regression of these tumors, tions, many of which have definable chemical structures, such either primary growth or autochthonous graft. Tumors of as wheat straw (3), certain edible mushrooms (1), and lichens about the size of a thumbtip were surgically removed as (2, 4). Repeated injections of these materials resulted in completely as possible. A small piece of the tumor, about 1 marked inhibition of growth of Sarcoma 180 implanted s.c., mm in diameter, together with 0.05 ml of 7-day-old Sarcoma with a high rate of complete regression. However, when tested 180 ascites tumor were implanted s.c. into the groin region, against autochthonous grafts of spontaneous mammary adeno- avoiding the site of operation. This was accomplished by carcinoma in mice, none of these polysaccharides was of any aspirating the Sarcoma 180 ascites into a tuberculin syringe value in inhibiting tumor growth producing regression (6). fitted with an injection needle with a caliber large enough to Sarcoma 180 is a long-term transplanted allogeneic tumor and, accommodate a solid tumor graft. The tumor graft was then although our experiments with this sarcoma have been carried inserted at the distal end of the needle, and was released under out in a tumor-host system (which ensured 100% takes with a the skin along with Sarcoma 180 by simply pushing the uniformly rapid rate of tumor growth), subtle differences plunger. All the tumors removed were histologically confirmed to be mammary adenocarcinoma. Received October 25, 1972; accepted December 22, 1972. After the operation, each mouse was treated daily with i.p. APRIL 1973 645 Downloaded from cancerres.aacrjournals.org on September 26, 2021. © 1973 American Association for Cancer Research. Reiko Tokuzen and Waro Nakahara injection of distilled water solution of lentinan. from implantation to complete regression was about 33 days Lentinan, j3-(l-»3)glucan isolated from an edible Japanese (S.D., 21 to 45 days). The results are summarized in Table 1. mushroom, Lentinus edades, was chosen as a representative of At the time of death, mice treated with lentinan were the polysaccharides known to be active in inhibiting Sarcoma classified into 3 groups according to the fate of implanted 180 that has been implanted in mice. The dose used, 1 mg/kg tumor: complete regression, incomplete regression, and large daily for 10 days, was reported by Chinara et al. (1) to tumor. produce marked inhibition of Sarcoma 180 with a high rate of If the mice with incompletely regressed tumors had lived complete regression. longer, they might have been classified under "complete regression," because the tumors were gradually shrinking. As controls, autochthonous mammary tumor grafts mixed with Sarcoma 180 were implanted by the same method as Histologically, these small tumors were mainly Sarcoma 180 above but did not receive lentinan treatment ("No Lentinan" with granulation tissue and large necrosis, but islets of controls). adenocarcinoma grafts also remained in the tumors. The cause All mice were kept in individual cages, maintained under the of death for these animals was local recurrence at the site of identical laboratory conditions, and carefully inspected once a operation and new tumors that developed elsewhere in the week. Any tumor nodules palpated were noted, including mammary gland. implanted tumors (autografi plus Sarcoma 180); local re In spite of lentinan treatment, 8 out of 37 mice died with a currence of tumors at the site of operation and new tumors large tumor at the site of implantation and with much tumor apart from the site of operation appeared often. ascites. Seven of these large tumors were histologically Careful autopsy was performed on all mice that died. confirmed to be Sarcoma 180, but 1 tumor was mammary adenocarcinoma. These diagnoses were made on sections of a part of each large tumor. Without examinations of complete RESULTS serial sections, it was impossible to say whether any adenocarcinoma existed in a sarcoma or vice versa. Tumors in Mice Treated with Lentinan. The implanted The postoperative survival in the complete regression group tumors, composed of autochthonous graft and Sarcoma 180, with lentinan treatment was not as prolonged as we had enlarged to approximately the size of a small fingertip by the expected. This was probably due to the local recurrence of the end of the 1st week. During this time, as we had already tumors at the site of operation and/or of new mammary reported, there was extensive outpouring of lymphoid cells, tumors that developed at other sites and that grew very mainly plasma cells and macrophages, in the immediate rapidly. vicinity of the grafts. By the end of the 3rd week the tumors "No Lentinan" Controls. The mice without lentinan were gradually shrinking. This stage was characterized by treatment all died with large tumors at the site of implanta histologically small remains of autochthonous grafts of tion, and some also with tumor ascites. The tumors were mammary adenocarcinoma in the usual acinus form and diagnosed as Sarcoma 180, and no remains of the autografts of Sarcoma 180 (Fig. 1) that were surrounded by granulation mammary adenocarcinoma could be found in them by routine tissue and large necrotic areas. By the end of 5 or 6 weeks histological examination. A few of these controls developed after implantation, the tumor had undergone complete local recurrence at the site of operation and some new tumors regression in as many as 23 out of 37 mice. The average time elsewhere, but they were all small at the time of death. Fig. 1. Granulation tissue containing remains of autografi (adenocarcinoma) weeks after mixed implantation with coma 180 in lentinan-treated mouse. X 100. 646 CANCER RESEARCH VOL. 33 Downloaded from cancerres.aacrjournals.org on September 26, 2021. © 1973 American Association for Cancer Research. Suppression of Autochthonous Tumor Grafts Table 1 Effect of ¡entinanon autografa of implanted spontaneous mammary tumors mixed with Sarcoma 180 Autografi + 180Complete Sarcoma recurrenceNo. tumorNo. of of of postoperative mice%Lentinan, micemg/kgformice %New survival(days)62.8(28-135)° 1 10 days (37 mice) regression 23 62.2 14 73.4 12 [incomplete regression 616.28 5 83.3 1 16.7 34.0(26-51) LargetumorLarge 21.66100Local4Control5 62.5 50.066.6Av.53.6(34-96)43.2 (6 mice) tumorNo. 2 33.34%63.2 (34-61) a Nos. in parentheses, standard deviation. DISCUSSION sufficient systemic resistance to prevent the development of mammary tumors elsewhere. The local reaction induced against an allogeneic tumor graft (Sarcoma 180) was capable of so involving the implanted autochthonous tumor graft mixed with it that the mixed graft ACKNOWLEDGMENTS became completely suppressed in the majority of instances. We are indebted to Dr. Hiroshi Nagasawa, for supplying the This failure of autochthonous graft was not due to its spontaneous mammary tumor-bearing mice, and to Dr.
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