Transplantation of human tumors in nude mice.

Ninety-one human tumors, including various common carcinomas, low-grade malignant tumors, and benign tumors, were transplanted into athymic nude mice. Tumor take was confirmed histologically for 22 neoplasms at the initial transplantation, and 14 serially transplantable tumors were established, including some hitherto unestablished or unreported, such as lung and hepatic cell carcinomas. Among the 91 tumors were 21, 14, and 13 carcinomas of the lung, stomach, and breast, respectively. Transplantability was highest in lung carcinomas (10/21), followed by gastric carcinomas (2/14) and breast carcinomas (1/13). Morphology of original tumors was retained well in most transplanted tumors, but desmoplastic or scirrhous tumors, such as gastric and breast carcinomas, tended to become medullary with a decrease in amount of tumor stroma. The ability to produce mucin in gastric carcinomas or melanin in malignant melanoma was maintained in serially transplantable tumors. In addition, ectopic production of adrenocorticotropin and beta melanocyte-stimulating hormone continued in a transplanted small cell carcinoma of the lung. Preliminary results were obtained on hormone dependency of the transplantable breast carcinoma and on alpha1-fetoprotein in the transplantable hepatic cell carcinoma.     

Metastasis of human tumors in athymic nude mice

The incidence of metastasis of xenogeneic tumors transplanted to nude mice is controversial. We studied 106 malignant human tumor lines in a total of 1,045 nude mice, and observed metastasis in only 14 instances (1.3%), involving 11 different tumor lines. Three of the lines showed repeated metastasis. Breast tumor lines metastasized with significantly greater frequency than other tumor types. None of the sarcoma lines metastasized. Tumors derived from human metastases were no more prone to metastasizing in nude mice than were tumors derived from primary sites. However, deep penetration of the body wall during growth of the tumor transplant was highly correlated with metastasis (p less than 0.001). Such factors as nude mouse health, tumor size and growth rate, and age and sex of the host mouse were not correlated with metastasis. Serial passage in nude mice did not select for a more malignant tumor line, since the incidence of metastasis did not differ at various passage levels. Thus, metastasis of human malignant tumors in nude mice would appear to depend primarily upon the site of tumor growth in the nude mouse, and upon the intrinsic metastasizing capability of the tumor line employed.     

Interleukin 17, a T-cell-derived cytokine, promotes tumorigenicity of human cervical tumors in nude mice.

Interleukin (IL) 17 is a proinflammatory cytokine secreted mainly by activated human memory CD4 T cells that induces IL-6, IL-8, and nitric oxide. Because IL-6 and IL-8 have been implicated in the pathogenesis of cervical cancer, we investigated the action of IL-17 on human cervical tumor cell lines in vitro and in vivo. We showed that in vitro, IL-17 increases IL-6 and IL-8 secretion by cervical carcinoma cell lines at both protein and mRNA levels. No direct effect of IL-17 on in vitro proliferation of cervical tumor cell lines could be demonstrated. However, two cervical cell lines transfected with a cDNA encoding IL-17 exhibited a significant increase in tumor size as compared to the parent tumor when transplanted in nude mice. This enhanced tumor growth elicited by IL-17 was associated with increased expression of IL-6 and macrophage recruitment at the tumor site. A potential role of IL-17 in modulation of the human cervical tumor phenotype was also supported by its expression on the cervical tumor in patients with CD4 infiltration. IL-17 therefore behaves like a T-cell-specific cytokine with paradoxical tumor-promoting activity. This may partially explain previous reports concerning the deleterious effect of CD4 T cells in cancer.     

Combined immunochemotherapy of human solid tumors in nude mice

In vivo immunochemotherapy of human solid tumors was studied in a nude mouse model. Large tumors (3 to 6 cm3) were induced by s.c. injection of the acute lymphoblastic leukemia T-cell line CEM. Transient tumor inhibition could be achieved by intratumoral injection of an intact-ricin immunotoxin that specifically recognizes a determinant CD5 (T,p67) expressed on the cell surface. Injection of the in vitro active cyclophosphamide congener mafosfamid had little effect on the progression of tumor growth. A combination regimen of immunotoxin and mafosfamid induced the most dramatic antitumor effect; a 72 to 100% reduction in tumor volume was observed within 3 to 4 days posttreatment. However, tumors relapsed within 5 to 13 days. Persistent, tumor regression was observed only when protocols using successive injections of combined immunotoxin/mafosfamid were used. All seven mice undergoing this treatment had a precipitous decrease in tumor size, and 86% survived greater than 30 days posttreatment. No residual tumor was detectable on Day 30 in five of seven mice. Regression was partly attributed to the selective activity of immunotoxin, since successive injections of an irrelevant control immunotoxin coupled to ricin in combination with mafosfamid did not reduce tumor size. Thus, we have demonstrated that a combination of anticancer chemotherapy and immunotoxin therapy yielded a greater antitumor effect than either therapy alone.     

Vascular structure of five human malignant melanomas grown in athymic nude mice.

The vascular structure of 5 human malignant melanomas grown in athymic nude mice was characterized. The vessels were filled with a radio-opaque medium administered via the abdominal aorta of the mice. X-ray images, obtained from 720 micrometers-thick tumour sections, provided qualitative information on the vascular structure of the tumours. Histograms for vessel length, surface, and volume as a function of vessel diameter were obtained by stereological analysis of 2 micrometers-thick sections. The volume fraction of necrotic tissue in the tumours was also determined by stereological analysis. The 5 melanomas exhibited individual, characteristic vascular structures as well as individual, characteristic necrotic fractions. The total vessel length ranged from 32 +/- 2 to 80 +/- 4 mm, the total vessel surface from 1.6 +/- 0.1 to 3.8 +/- 0.2 mm2, and the total vessel volume from 0.009 +/- 0.001 to 0.022 +/- 0.002 mm3--all values per mm3 histologically intact tumour tissue. The necrotic fractions ranged from 30 +/- 1 to 49 +/- 4%, and tended to be higher in the poorly than in the well-vascularized melanomas. The volume doubling times ranged from 4.2 to 21.6 days. Melanomas with short volume-doubling times had lower necrotic fractions and tended to be better vascularized than those with long volume-doubling times.     

Tumor-bone interaction induced by transplantable human tumors in nude mice

Tumor-bone interactions were experimentally studied using four transplantable human urogenital tumors in nude mice. The method consisted of subcutaneously (SC) inoculating tumor cells over the calvaria in nude mice after the periosteum has been disrupted. This resulted in a local tumor causing fragmentation of the bone. The degree of tumor-bone interaction also varied with the type of implanted tumors as shown in radiographic and histologic examinations. All tumors were associated with histologic patterns of classical bone remodeling, including bone destruction with osteoclast proliferation and reactive new bone formation. The evidence presented here suggests that the majority of tumor-bone interaction showed a combination of both features, bone destruction and new bone formation, and the mechanisms whereby tumors interact with bone may vary with the biological properties of the tumor. Our new system would be suitable for studying the biology of local interaction between bone and tumor cells and searching out a method to protect the bone from cancer cells.     

Antitumor activity of ME2303, a fluorine-containing anthracycline, against human tumors implanted in nude mice

A fluorine-containing anthracycline, ME2303, given intravenously once a week for 4 weeks at the maximum tolerated doses showed better therapeutic effects against 2 gastric, 3 lung and 2 human breast tumor xenografts than did adriamycin (ADM) at the maximum tolerated dose. Among the tumors, ME2303 showed a better effect against St-40, a well-differentiated human gastric adenocarcinoma, against which ADM showed only a marginal effect. Likewise, ME2303 was more effective against Lu-24 human small cell carcinoma and MX-1 human medullary tubular adenocarcinoma than ADM. Notably, the Lu-24 tumor, developed in nude mice, disappeared after the treatment in 3 out of 6 mice. ME2303 would be an interesting compound for phase I and II clinical studies in the future.     

Regression of human tumors established in nude mice after continuous infusion of thymidine.

A system for the continuous infusion of thymidine solutions in nude mice has been developed. High doses (0.5 to 1.0 ml/mouse/hr) of a 28.5-mg/ml thymidine solution (444 to 888 mg/kg/hr) can be administered continuously for 96 to 140 hr. The preliminary results indicate that it is possible to induce total tumor regression of human heterotransplants established in nude mice of one human teratocarcinoma, five different human melanomas, and one human lung carcinoma and to inhibit to a large degree the growth of two human breast carcinomas by multiple (two to eight) cycles of infusion. The life span of the thymidine-treated animals has been significantly increased compared to that of control animals.     

Antimetastatic intraoperative chemotherapy of human colon tumors in the livers of nude mice.

We have developed a new antimetastatic chemotherapeutic strategy for combination with hepatic resection of human colon cancers in a high-metastasis nude mouse model. The new procedure involves i.p. administration of 5-fluorouracil (5-FU) 2 h before hepatic resection of the human colon tumors, with therapy continued postoperatively for 4 consecutive days. We termed this strategy neo-neoadjuvant chemotherapy. The regime significantly prolonged animal survival compared with preoperative 5-FU neoadjuvant therapy, 5-FU postoperative adjuvant therapy, surgery alone, 5-FU without surgery, or the untreated control. The median survival of neo-neoadjuvant i.p. 5-FU-treated group was 81 days, compared with 27 days for the control group (P < 0.009). The median survival of animals in the neoadjuvant group was 37 days (P < 0.021 compared with the control group). There was also a significant difference between the median survival of neo-neoadjuvant, and the neoadjuvant group (P < 0.031). When all animals in the control group had died, 70% of animals with neo-neoadjuvant and 60% of animals with neoadjuvant 5-FU were still alive (P < 0.003 and P < 0.011, respectively). When all animals with neoadjuvant 5-FU treatment had died, 70% of animals with neo-neoadjuvant treatment were still alive (P < 0.003). Survival of all other treatment groups, including 5-FU without surgery, surgery alone, and adjuvant postoperative chemotherapy, was not significantly different from the untreated control group. Two animals in the neo-neoadjuvant group were free of tumors when sacrificed at days 154 and 165 post surgery. Whereas 100% of animals in the control, 90% in the 5-FU alone, 70% in the surgery alone, 60% in the 5-FU adjuvant, and 40% in the neoadjuvant groups had metastases in the lymph nodes draining the liver, only 10% of animals in the neo-neoadjuvant group had metastases. These data suggest that the neo-neoadjuvant therapy increased survival by preventing metastasis of cancer cells not removed in the liver resection procedure. The results of this study indicate that the neo-neoadjuvant treatment strategy for resection of colon cancer liver metastasis should be explored clinically.     

Comparative growth of human tumors in pharmacologically immunosuppressed,immune-deprived,cyclosporin a-treated and nude mice

The growth of three human tumor xenografts, namely an Ewing sarcoma, a colon carcinoma and an osteosarcoma, was compared in nude and conditioned mice. Conditioning protocols included (1) immune deprivation (thymectomy, lethal irradiation and cytosine arabinoside pretreatment); (2) immunosuppression with procarbazine, cyclophosphamide and antithymocyte serum; and (3) continuous administration of cyclosporin A. Similar tumor growth was seen in nude mice, immune-deprived mice and mice treated with the medium and high-dose immunosuppressive protocol. Cyclosporin A allowed only very modest tumor growth. In the main comparative experiment with the Ewing sarcoma and the colon carcinoma, overall survival was lowest with nude mice (17%), higher with immune-deprived mice (61%) and best with immunosuppressed mice (81 and 87%). For the screening of anticancer agents the immunosuppressive protocol consisting of synergistic procarbazine, cyclophosphamide and antithymocyte serum may be added to the already available models. It allows adequate tumor growth with good animal survival and does not require operative procedures and irradiation.     

One hundred and twenty-seven cultured human tumor cell lines producing tumors in nude mice.

One hundred and twenty-seven cultured human tumor cell lines produced tumors after sc inoculation of 1-20 million cells into nude mice. They included 56 carcinoma lines, 14 sarcoma lines, and 57 lines from miscellaneous tumors and were all glucose-6-phosphate dehydrogenase type B. Twenty-nine percent of the lines produced tumors of 1 cm3 size within 1 month and 41% in the second month after inoculation. The histopathology correlated with the human tumor of origin in all cases.     

Long-term observation on transplanted tumors of human intestinal mucoid adenocarcinoma in nude mice

Long-term observation on transplanted tumor of human intestinal mucoid adenocarcinoma in nude mice is reported. The transplanted tumor which has been passed 57 passages in 151 nude mice with transplant survival rate of 99.34% in more than 5 years is the tumor with the utmost number of passages domestically undertaken as yet. During the long-term observation, the transplanted tumor was not only identical to the primary tumor in histopathology, ultrastructure and chromosome pattern, but also constant in biological characteristics, such as secretion of mucin and production of carcinoembryonic antigen. These tumor cells were able to combine with CEA mcAb specifically by obviously showing the scanning image of transplanted tumor of human intestinal mucoid adenocarcinoma in nude mice with 131I-CEAmcAb. The roles of the transplanted tumor model in cancer research are discussed.     

Detection of metastatic tumors in nude mice: brief communication.

Homozygous nude (nu/nu) mice were inoculated ip with either highly malignant human bladder transitional cell carcinoma or human prostate adenocarcinoma. These animals were subsequently given injections of normal human T-lymphocytes to restore the known T-lymphocyte deficiency present in homozygous nude mice. Metastatic spread of the prostate and bladder carcinomas was evident in mice given human T-lymphocytes. Although tumor growth was observed at the sites of tumor inoculation, no tumor spread was observed in mice not receiving T-lymphocytes.     

Rottlerin suppresses growth of human pancreatic tumors in nude mice,and pancreatic cancer cells isolated from Kras mice

The purpose of the study was to examine the molecular mechanisms by which rottlerin inhibited growth of human pancreatic tumors in Balb C nude mice, and pancreatic cancer cells isolated from Kras^G12D mice. AsPC-1 cells were injected subcutaneously into Balb c nude mice, and tumor-bearing mice were treated with rottlerin. Cell proliferation and apoptosis were measured by Ki67 and TUNEL staining, respectively. The expression of components of Akt, Notch, and Sonic Hedgehog (Shh) pathways were measured by the immunohistochemistry, Western blot analysis, and/or q-RT-PCR. The effects of rottlerin on pancreatic cancer cells isolated from Kras^G12D mice were also examined. Rottlerin-treated mice showed a significant inhibition in tumor growth which was associated with suppression of cell proliferation, activation of capase-3 and cleavage of PARP. Rottlerin inhibited the expression of Bcl-2, cyclin D1, CDK2 and CDK6, and induced the expression of Bax in tumor tissues compared to untreated control. Rottlerin inhibited the markers of angiogenesis (Cox-2, VEGF, VEGFR, and IL-8), and metastasis (MMP-2 and MMP-9), thus blocking production of tumorigenic mediators in tumor microenvironment. Rottlerin also inhibited epithelial–mesenchymal transition by up-regulating E-cadherin and inhibiting the expression of Slug and Snail. Furthermore, rottlerin treatment of xenografted tumors or pancreatic cancer cells isolated from Kras^G12D mice showed a significant inhibition in Akt, Shh and Notch pathways compared to control groups. These data suggest that rottlerin can inhibit pancreatic cancer growth by suppressing multiple signaling pathways which are constitutively active in pancreatic cancer. Taken together, our data show that the rottlerin induces apoptosis and inhibits pancreatic cancer growth by targeting Akt, Notch and Shh signaling pathways, and provide a new therapeutic approach with translational potential for humans.     

Calorie restriction: effect on growth of human tumors heterotransplanted in nude mice

The presence of 4 human malignant tumors (1 breast, 1 lung, and 2 colon carcinomas) growing subcutaneously as heterotransplants in nude mice did not significantly affect the body weights of adult animals until the tumors reached very large dimensions (tumor wt greater than 15% of the body wt). However, a colon carcinoma (HT 29) induced a cessation of the natural rate of body weight increase when it grew in young adults (animals weighing approximately equal to 25 g which will gain 6 g or approximately equal to 25% body wt in 1 mo). Calorie restriction at all the levels tested (8, 6, 4, and 2 g/day/mouse) with standard pelletized mouse food produced both weight loss in the animals (with and without tumor) and a lowering of the growth rate of all the 4 tumors tested growing at a subcutaneous site and/or under the kidney capsule. Each tumor responded differently to the calorie restriction. The 4 tumors tested grew equally in both male and female nude mice. Young animals weighing 20 g inoculated with a fifth tumor (MeWo melanoma) exhibited tumor growth inhibition proportional to restriction of calorie intake. Their survival, however, did not improve.     

Targeted therapy with a Salmonella typhimurium leucine-arginine auxotroph cures orthotopic human breast tumors in nude mice

We report here a modified auxotrophic strain of Salmonella typhimurium that can target and cure breast tumors in orthotopic mouse models. We have previously reported development of a genetically modified strain of S. typhimurium, selected for prostate tumor targeting and therapy in vivo. The strain, termed S. typhimurium A1, selectively grew in prostate tumors in xenograft models causing tumor regression. In contrast, normal tissue was cleared of these bacteria even in immunodeficient athymic mice with no apparent side effects. A1 is auxotrophic (leucine-arginine dependent) but apparently receives sufficient nutritional support only from tumor tissue. The ability to grow in viable tumor tissue may account, in part, for the unique antitumor efficacy of the strain. In the present report, to increase tumor-targeting capability of A1, the strain was reisolated after infection of a human colon tumor growing in nude mice. The tumor-isolated strain, termed A1-R, had increased targeting for tumor cells in vivo as well as in vitro compared with A1. Treatment with A1-R resulted in highly effective tumor targeting, including viable tumor tissue and significant tumor shrinkage in mice with s.c. or orthotopic human breast cancer xerographs. Survival of the treated animals was significantly prolonged. Forty percent of treated mice were cured completely and survived as long as non-tumor-bearing mice. These results suggest that amino acid auxotrophic virulent bacteria, which selectively infect and attack viable tumor tissue, are a promising approach to cancer therapy.     

Radiotherapy of human xenograft NSCLC tumors in nude mice with a 90Y-labeled anti-tissue factor antibody

Tissue factor (TF) is a type I transmembrane protein and the initiator of the extrinsic blood coagulation pathway. TF plays a critical role in tumor development and its overexpression is observed in many tumors. To understand the prevalence and relative level of TF expression in non-small-cell lung cancer (NSCLC), we analyzed 50 NSCLC tumors by immunohistochemical staining and found that 88% of human NSCLC tumors overexpressed TF. We then generated a high affinity anti-TF antibody, TF278, which specifically binds TF on the surface of cells and is internalized upon binding. An 111In-labeled TF278 demonstrated favorable tumor accumulation in an SW-900 xenograft tumor model with a maximum mean percent of injected dose per gram of tissue (%ID/g) of 73.1% at 96 hours postinjection. In addition, we labeled the antibody with 90Y and tested its ability to inhibit the growth of tumors in an SW-900 xenograft tumor model in immunocompromised mice. The 90Y-TF278 slowed the growth of SW-900 tumors at a 50 microCi dose and completely regressed SW-900 tumors at a 150 microCi dose with little toxicity.