^153Sm—乙二胺四亚甲基膦酸对Walker256癌荷瘤大鼠骨侵袭骨溶解…

目的 研究＾１５３Ｓｍ－乙二胺四亚甲基膦酸（＾１５３Ｓｍ－ＥＤＴＭＰ）对荷瘤大鼠骨侵袭、骨溶解的抑制作用。方法 运用Ｘ线骨骼照片和胫骨病理切片,观察Ｗａｌｋｅｒ２５６癌大鼠模型的肿瘤骨侵袭和骨溶解情况。结果 荷瘤大鼠静脉注射＾１５３Ｓｍ－ＥＤＴＭＰ７４,１４８ＭＢｇ／ｋｇ后,遭受Ｗａｌｋｅｒ２５６癌侵袭及发生骨溶解的胫骨数减少近一半,３７ＭＢｇ／ｋｇ组对癌的骨侵袭和骨溶解仍有抑制作用。但各组大鼠的     

~(153)Sm-乙二胺四亚甲基膦酸对 Walker 256癌荷瘤大鼠骨侵袭骨溶解的抑制作用

目的研究１５３Ｓｍ－乙二胺四亚甲基膦酸（１５３Ｓｍ－ＥＤＴＭＰ）对荷瘤大鼠骨侵袭、骨溶解的抑制作用。方法运用Ｘ线骨骼照片和胫骨病理切片，观察Ｗａｌｋｅｒ２５６癌大鼠模型的肿瘤骨侵袭和骨溶解情况。结果荷瘤大鼠静脉注射１５３Ｓｍ－ＥＤＴＭＰ７４，１４８ＭＢｇ／ｋｇ后，遭受Ｗａｌｋｅｒ２５６癌侵袭及发生骨溶解的胫骨数减少近一半，３７ＭＢｇ／ｋｇ组对癌的骨侵袭和骨溶解仍有抑制作用。但各组大鼠的移植瘤重量没有差异。结论１５３Ｓｍ－ＥＤＴＭＰ能抑制Ｗａｌｋｅｒ癌模型大鼠骨侵袭和骨溶解，但对移植瘤生长无抑制作用，故其抗癌骨侵袭、骨溶解的作用不是通过抑制动物移植瘤生长来实现的。     

Pharmacokinetics of fotemustine and BCNU in plasma,liver and tumor tissue of rats bearing two lines of Walker 256 carcinoma

Summary The plasma and tissue pharmacokinetics of fotemustine (diethyl-1-[3-(2-chlorethyl)-3-nitrosoureido]-ethylphosphonate) and BCNU (1,3-bis-[2-chlorethyl]-1-nitrosourea) were investigated in healthy control rats and in animals bearing either the nitrosourea-sensitive line A (W256/A) or the nitrosourea-resistant line B (W256/B) of Walker 256 carcinoma. The antitumor activities of these nitrosoureas were similar following i.v. doses ranging from 10 to 40 mg/kg. For both drugs, the survival of tumor-bearing rats was lower in the W256/B than in the sensitive W256/A line. Some sex differences were observed, female rats being more responsive than males to both drugs. Nitrosourea concentrations were assayed in plasma and tissues by differential pulse polarography so as to assess whether the pharmacokinetics could explain the differences in antitumor activity. The antineoplastic effects of fotemustine seemed to be influenced by its pharmacokinetics. The plasma AUC of the intact nitrosourea was higher in females than in males. Fotemustine was cleared 2–5 times more slowly than BCNU from tumor tissue, and its clearance was higher in W256/B- than in W256/A-bearing rats. This suggests that the antitumor activity in the responsive line might partly be due to longer exposure of the growing tumor to the drug. The distribution volume of both nitrosoureas in plasma was higher in tumor-bearing animals than in healthy controls, indicating that the tumor tissue probably constitutes an additional distribution space.Abbreviations BCNU 1,3-bis-[2-chlorethyl]-1-nitrosourea - W256/A Walker carcinoma 256 line A - W256/B Walker carcinoma 256 line B     

Different sensitivity of two Walker 256 carcinoma lines to cyclophosphamide: correlation with drug distribution, biotransformation and macromolecule binding

Two closely related lines of the same Walker 256 carcinoma in Crl-CD/COBS rats, described as behaving differently as regards tumor growth and host reaction, show different chemotherapeutic sensitivity to cyclophosphamide (CPA). Line B, which induces early cachexia with marked anorexia, is only moderately sensitive to CPA, while line A, which causes mild anorexia and only terminal cachexia, shows marked responsiveness to CPA, cure being attained in 75% of animals treated with a single dose of 120 mg/kg and in 90-100% of those given 20 mg/kg every other day. Comparative studies in both tumor lines on the distribution of CPA in vivo and on its metabolism by the liver perfusion technique showed no appreciable differences between the two lines in the pharmacokinetics of the compound, but indicate a much greater metabolizing capacity of CPA in the Walker 256/A animals. In vitro metabolic and covalent binding studies confirm that the liver of the Walker 256/A group metabolizes and covalently binds twice as much CPA as the liver of the Walker 256/B group. Conversely to Walker B, microsomal preparations of the Walker tumor line A are able to metabolize CPA to intermediates which irreversibly bind to tissue macromolecules, suggesting an in situ activation of the compound in the sensitive Walker tumor.     

Mechanical analysis of tumor growth regression by the cyclooxygenase-2 inhibitor, DFU, in a Walker256 rat tumor model: importance of monocyte chemoattractant protein-1 modulation.

Cyclooxygenase (COX)-2 inhibition results in tumor regression; however, little is known about the mechanism. In the present study, using a Walker256 tumor model and a rat bone marrow-derived endothelial cell line TR-BME-2, we analyzed the effects of a new selective COX-2 inhibitor, 5,5-dimethyl-3-(3-fluorophenyl)-4-(4-methylsulphonyl)phenyl-2-(5H)-furanone (DFU), on the production of chemokines and growth factors and on the neovascularization. The oral administration of DFU (5 mg/kg/d) significantly suppressed the tumor growth with decreasing microvessel density in vivo, although it showed no direct inhibition of Walker256 cell proliferation in vitro. It was newly found that the recruitment of systemically injected TR-BME-2 cells into the tumor site was significantly inhibited by DFU treatment. In addition, we found that DFU significantly reduced the production of monocyte chemoattractant protein-1 (MCP-1) both in tumor tissues and in the systemic circulation (P < 0.001 and P < 0.001, respectively). Such reduction was not observed in other chemotactic factors, vascular endothelial growth factor and stromal cell-derived factor-1. The induced chemotaxis of TR-BME-2 by serum of tumor-bearing rats was significantly reduced in DFU-treated rat serum, although DFU showed no direct inhibition for TR-BME-2 cells, either cell growth or chemotaxis. Treatment with neutralizing antibodies to soluble mediators, including MCP-1, significantly suppressed the chemotaxis. Regarding the down-regulation machinery of MCP-1 production in vivo, tumor-associated macrophages seem to play crucial roles, because DFU eliminated MCP-1 production in the activated macrophages remarkably but not in Walker256 tumor cells in vitro. In conclusion, COX-2 inhibitor DFU exerts tumor regression activity in a Walker256 tumor model by suppressing MCP-1 production in tumor tissues and in the circulation.     

An experimental rat model of local bone cancer invasion and its responsiveness to ethane-1-hydroxy-1,1-bis(phosphonate)

Line A Walker carcinoma differs from line B in that it does not elicit hypercalcemia and hypercalciuria when implanted in rats at various sites (s.c, i.m., intraaortically). However, Walker 256/A, unlike line B, may invade the tibia when implanted i.m. in the adjacent gastrocnemius muscle. This invasion was evaluated by measuring the increased weight of the bone and decreased calcium concentration per unit weight of the tibia, by reduced opacity to X-ray, and by the presence of tumor cells in the compact bone cortex. Ethane-1-hydroxy-1,1-bis(phosphonate), a diphosphonate derivative, at a dose of 10 to 30 mg/kg/day s.c., prevented cancer cell invasion of the tibia as judged by the above criteria. This inhibition was obtained with no apparent effect on the growth of Walker 256/A carcinoma.     

Failure of systemic ketosis to control cachexia and the growth rate of the Walker 256 carcinosarcoma in rats

The Walker 256 carcinosarcoma was shown to lack the enzyme 3-ketoacid CoA transferase. This suggests that ketone bodies cannot be used as a major substrate for the energy metabolism of this tumour. Systemic ketosis (1-2 mM acetoacetate plus 3-hydroxybutyrate) was induced both in tumour-bearing and in non-tumour-bearing rats with a diet containing 70% medium chain triglyceride. However, in rats bearing the Walker 256 tumour, this dietary ketosis did not reduce the tumour growth rate nor did it prevent the subsequent decrease in host body weight. Host body nitrogen losses were similarly unaffected. The ketosis induced in tumour bearing rats was shown to be abnormal since the blood glucose concentration of ketotic, tumour-bearing rats was significantly higher compared with that of ketotic non-tumour bearing rats (5.2 +/- 0.4 mM cf 3.4 +/- 0.6 mM, P less than 0.01). These results may partly explain why systemic ketosis failed to alter the growth and cachexia induced by the Walker 256 carcinosarcoma.     

Host stromal bradykinin B2 receptor signaling facilitates tumor-associated angiogenesis and tumor growth

We evaluated the significance of the host kallikrein-kinin system in tumor angiogenesis and tumor growth using two rodent models genetically deficient in a kallikrein-kinin system. Inoculation of Walker 256 carcinoma cells into the s.c. tissues of the back of normal Brown Norway Kitasato rats (BN-Ki rats) resulted in the rapid development of solid tumors with marked angiogenesis. By contrast, in kininogen-deficient Brown Norway Katholiek rats (BN-Ka rats), which cannot generate intrinsic bradykinin (BK), the weights of the tumors and the extent of angiogenesis were significantly less than those in BN-Ki rats. Daily administration of B(2) receptor antagonists significantly reduced angiogenesis and tumor weights in BN-Ki rats to levels similar to those in BN-Ka rats but did not do so in BN-Ka rats. Angiogenesis and tumor growth were significantly suppressed in B(2) receptor knockout mice bearing sarcoma 180 compared with their wild-type counterparts. Immunoreactive vascular endothelial growth factor (VEGF) was localized in Walker tumor stroma more extensively in BN-Ki rats than in BN-Ka rats, although immunoreactive B(2) receptor also was detected in the stroma to the same extent in both types of rats. Cultured stromal fibroblasts isolated from BN-Ki rats and BN-Ka rats produced VEGF in response to BK (10(-8)-10(-6) m), and this stimulatory effect of BK was abolished with a B(2) receptor antagonist, Hoe140 (10(-5) m). These results suggest that BK generated from kininogens supplied from the host may facilitate tumor-associated angiogenesis and tumor growth by stimulating stromal B(2) signaling to up-regulate VEGF production mainly in fibroblasts.     

THE EFFECT OF PAEONIA RUBRA 801 ON THE OCCURRENCE OF LUNG METASTASES IN WALKER 256 TUMOR

The development of lung nodules from intravenously injected Walker 256 (W256) tumor cells was the model system for metastases formation in this experiment. The influence and inhibitory mechanisms of a synthetic Paecnia rubra 801 (P801) on occurrence of lung metastases were investigated. The results showed that treatment with P801 alone or in combination with decoction of Huangqi (Astragalus root) can significantly reduce the number of lung nodules. The mechanisms responsible for decreased colonyforming potential in the rats treated with P801 were investigated. ADP-induced or W256 tumor cell-induced platelet aggregation was markedly inhibited by P801. It appears that inhibitory effect of P801 on occurrence of W256 tumor lung nodules is mainly due to reduced blood coagulability and inhibition of platelet activation by tumor cells.     

Two lines of Walker carcinoma 256: their peculiarities and different interactions with the host

Two sublines of Walker 256 carcinoma have been characterized for their ability to metastasize and to induce cachexia. The invasive, metastasizing line A induced terminal anorexia in rats with a mean survival time of 27 +/- 1.5 days. The non-invasive line B induced early anorexia and cachexia with a mean survival time of only 15 +/- 1 days. At death, the line B tumor was still smaller than the line A one, and no metastases were detectable. These two sublines are discussed as a composite model for studying anorexia and cachexia together with invasion and metastasis.     

Osteoporosis-like Changes in Walker Carcinoma 256-Bearing Rats, Not Accompanied with Hypercalcemia or Parathyroid Hormone-related Protein Production

Walker carcinoma 256 (W256) was reported to induce hypercalcemia dependent on bone metastasis and/or parathyroid hormone-related protein (PTHrP) in the rat, providing a model of the humoral hypercalcemia of malignancy. In this study, after the subcutaneous inoculation of cells of the W256/S line, which is maintained in this laboratory, into young female Wistar Imamichi rats (6 weeks old), serum calcium and phosphorus levels changed only within the control range, whereas serum alkaline phosphatase activity and urinary calcium level significantly increased and urinary phosphorus decreased during the tumor growth, resulting in hypercalciuria and hypophosphaturia. W256/S did not express PTHrP-mRNA, whereas LLC-W256 cells did express it. Serum PTHrP level was not changed in W256/S-bearing rats. Osteoporosis-like changes, bone weight loss, low contents of bone calcium and phosphorus, and a decrease in the bone mineral density (BMD), were observed in the femur 14 days after the tumor inoculation. There was a pronounced decrease in the serum 17β-estradiol level during the tumor growth. The reduction of BMD of femurs in W256/S-bearing rats was significantly inhibited by treatment with salmon calcitonin or 17β-estradiol. On the basis of these results, W256/S carcinoma-bearing rats seem to be a useful model for osteoporosis of hypoovarianism.     

Degradation of insulin and glucagon by a factor associated with Walker 256 carcinosarcoma cells

Earlier studies from this laboratory reported that, in rats bearing the Walker 256 carcinosarcoma, circulating insulin levels were significantly reduced relative to non-tumor-bearing rats. The present study extends this observation to include a significantly (P less than 0.01) reduced plasma level of glucagon in rats bearing the tumor for both 7 and 10 days. In order to determine if the tumor itself somehow plays a role in the degradation of these protein hormones, either cultured Walker 256 tumor cells (in the case of the insulin studies) or cells from freshly excised tumor (for the glucagon studies) were incubated with 125I-labeled insulin or glucagon. Following the incubation period, the amount of TCA-precipitable radio-label remaining in the incubation medium was markedly reduced after exposure to cells. This suggests that the tumor cells have the capability of degrading both insulin and glucagon. In a separate series of experiments, it was found that medium, in which freshly excised tumor cells had been incubated previously and then discarded, retained a substance which degraded 125I-labeled glucagon and that this degradation of glucagon could be blocked by co-incubation with aprotinin, a protease inhibitor.     

Pharmacokinetics of nitrosoureas: levels of 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) in organs of normal and Walker 256/B carcinoma bearing rats after i.v. bolus

The disappearance of 1,3-bis(2-chlorethyl)-1-nitrosourea (BCNU) from plasma, liver, kidney, lung, brain, spleen, tumor tissue and epididymal adipose tissue of Walker 256/B carcinoma-bearing rats and healthy animals was measured by differential pulse polarography after i.v. bolus of the drug. Only BCNU, not its decomposition products, was detected by the polarographic assay. Levels of BCNU in liver of tumor-bearing animals were significantly lower (about 10 times) than those on healthy rats. A bi-exponential fit was used to calculate the kinetics of BCNU in plasma, kidney, lung and brain, but no difference could be found between healthy and Walker tumor-bearing rats. BCNU disappeared faster from adipose tissue of tumor-bearing animals than from normals.     

Reduced Walker 256 carcinosarcoma growth in vasopressin-deficient Brattleboro rats

The growth pattern of carcinosarcoma Walker 256 was studied in rats with different levels of vasopressin in the blood. The Brattleboro rats are unable to synthesize vasopressin in a consequence of deletion in the coding gene. Hybrids from crossbreeding of the mutant Brattleboro and normal WAG rats inherit the one intact vasopressin gene and hold nearly normal hormone level. It was found that non-strain-specific carcinosarcoma Walker 256 intensively grows in WAG rats and their offsprings from crossbreeding with Brattleboro rats, and tumor development is equally terminated in them by death. Carcinosarcoma grows less intensely in Brattleboro rats; the tumor nodes increased only within the first 2 weeks, after which, the tumor began to decrease and eventually disappeared. Infusion of exogenous vasopressin to Brattleboro rats intensifies a tumor growth in the first 2 weeks after the inoculation of Walker 256 cells; however, it does not prevent a following regression and resorption of tumors.     

Differential macromolecular leakage from the vasculature of tumors

Tumor-induced neovascularization is essential for invasion, metastases, and exponential growth of solid tumors. The authors studied the differences in macromolecular leakage from the neovasculature of a fast-growing, early-metastasizing tumor, the Walker 256 carcinosarcoma, and a slow-growing, nonmetastasizing tumor, a rat chondrosarcoma. A 1-mm3 piece of the Walker 256 carcinoma or the chondrosarcoma was implanted in the cremaster muscle of rats. Five days after surgery the cremaster muscle with the implanted tumor was placed in a special bath containing Krebs solution such that the circulation and nerves from the animal to the cremaster were intact. Fluorescein isothiocyanate-labeled rat serum albumin (FITC-RSA) was injected (intra-arterially) into each rat to permit visualization of the vasculature by fluorescent microscopy. A closed-circuit television system was used to quantitate macromolecular leakage as a change in interstitial fluorescent intensity. Data are given as a relative fluorescent intensity (mean +/- standard error of the mean) in an area of the cremaster with tumor-induced neovascularization. These studies demonstrated that the vasculature induced by rapidly growing Walker 256 carcinosarcoma leak albumin freely when compared with the vasculature induced by the slow-growing chondrosarcoma. Furthermore, there was a significant increase in fluorescent intensity (albumin leakage) in the Walker tumor from 1 minute (24 +/- 3.0) to 30 minutes (49 +/- 5.6). In the normal cremaster area there was a significantly lower fluorescent intensity in the interstitium and a very slight increase with time (4 +/- 1.5 at 1 minute vs. 7 +/- 1.4 at 30 minutes). One interpretation of these data is that the mechanisms responsible for protein leakage from the vasculature of the Walker tumor may be involved in the fast growth and metastases of this tumor as compared with slower-growing tumors such as the chondrosarcoma.     

Hyperthermia-induced vascular injury in normal and neoplastic tissue

The sequential morphologic alterations in normal skeletal muscle in rats, Walker 256 tumors in rats, and transmissible venereal tumors (TVT) in dogs following microwave-induced hyperthermia (43 degrees C and 45 degrees C for 20 minutes), were studied by histologic and ultrastructural examination. Normal muscle and Walker 256 tumors showed edema, congestion, and hemorrhage at 5 minutes post-heating (PH), followed by suppuration, macrophage infiltration, and thrombosis at 6 and 48 hours PH, and finally by regeneration and repair by 7 days PH. Vascular endothelial damage and parenchymal degeneration were present 5 minutes PH. Progressive injury occurred for at least 48 hours PH. Two hyperthermia treatments separated by a 30- or 60-min cooling interval, were applied to Walker 256 tumors in a subsequent study. Increased selective heating of tumor tissue versus surrounding normal tissue, and increased intratumoral steady state temperatures were found during the second hyperthermia treatment. Canine TVTs were resistant to hyperthermia damage. These results suggest that vascular damage contributes to the immediate and latent cytotoxic effects of hyperthermia in normal tissue and some types of neoplastic tissue, and that selective heating of neoplastic tissue occurs in tumor tissue with disrupted microvasculature.     

Naringin inhibits tumor growth and reduces interleukin-6 and tumor necrosis factor α levels in rats with Walker 256 carcinosarcoma

The flavonoid naringin is a polyphenolic compound that naturally occurs in citrus. Patients with cancer generally present features of malnutrition and cachexia. Levels of the proinflammatory cytokines tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) are raised in patients with cancer. This study was designed to analyze the in vivo effect of naringin in the therapeutic treatment of rats bearing Walker 256 carcinosarcoma (W256). Rats were treated intraperitoneally with different doses of naringin (10, 25 and 35 mg/kg), for 50 days. At 25 mg/kg, naringin inhibited tumor growth by ~75%. With this treatment, TNF-α and IL-6 levels decreased (p<0.05) in comparison with the control. In addition, two rats presented complete tumor regression. Inhibition of tumor growth, survival increase and the reduction of TNF-α and IL-6 levels in rats bearing W256 treated with naringin strongly suggest that this compound has potential as an anticarcinogenic drug.     

Liver-to-lung traffic of cancer cells

Following the injection of B16 melanoma cells into the portal veins of mice, all animals developed liver tumors, but only 16% developed lung tumors. Portal vein injections of radiolabelled B16 and Walker 256 cancer cells into mice and rats, respectively, revealed that all of the cells were temporarily arrested in the liver and most were then slowly released. Bioassays indicated that of 8 X 10(4) B16 cells released from the liver over 24 h after portal vein injections of 10(5) cells, only approximately 1% were delivered to the lungs in a viable state. Experiments made with radiolabelled B16 cells showed that transit through either the liver or lungs following portal vein or tail vein injections, respectively, resulted in massive death of cancer cells. It is suggested that the death of most circulating cancer cells passing through the first organ encountered after leaving their primary tumor, serves to severely limit their further direct spread to other organs. It is therefore expected that metastases to these other organs would, to a large extent, be generated by cancer cells from metastases in the "first organs" as distinct from direct seeding from cancer cells released from the primary tumor. If the results of the present experiments have general application, they serve to emphasize the importance of metastasis of metastases in the natural history of the spread of cancer. In a previous publication (Weiss, 1980), studies of lung-to-liver traffic of cancer cells in rats revealed that, after tail vein injections, most Walker 256 cells were temporarily arrested in the pulmonary vasculature and then slowly released. A large proportion of the released cancer cells were dead or lethally injured on release from the lungs, and this "first organ processing" apparently accounted for the comparative rarity of extrapulmonary tumors following tail vein injections. In this communication, the concept of "first organ processing" of circulating cancer cells is further examined with respect to the liver-to-lung traffic of B16 melanoma and Walker 256 cells injected into the portal veins of mice or rats respectively. Both of these cell types grow well in the lungs following tail-vein injection.     

Circadian variations in 32P uptake of DMBA-induced mammary tumour and Walker carcinosarcoma in rats.

The 32P uptake in a mammary tumour induced by DMBA and in the Walker 256 carcinosarcoma was measured by external GM -tubes. The uptake was significantly higher than in the skin. During exposure to a synchronized light regime a circadian variation was present in the 32P uptake of the hormone-dependent DMBA-induced tumour. The maximal 32P uptake was in the dark period, in which the highest temperature in the tumour has also been found (Møoller and Bojsen, 1975). In the hormone-independent Walker 256 carcinosarcoma there was no periodicity in 32P uptake. No variation in 32P uptake was registered in the skin of normal controls or in tumour-bearing rats.