Antitumor effects of recombinant human interleukin 1 alpha in RIF-1 and Panc02 solid tumors

The antitumor effects of recombinant human interleukin 1 alpha (IL-1) were determined in RIF-1 and Panc02 murine solid tumors. Acute tumor hemorrhage was observed in both models as early as 3 h after a single 25 micrograms/kg IL-1 treatment and was quantitated by the intratumor accumulation of 59Fe-labeled erythrocytes (RBC). The IL-1-mediated hemorrhagic response was maximal 6-12 h after treatment and greater in Panc02 tumors than in RIF-1 tumors. Hemorrhagic responses to RIF-1 tumors growing in athymic nude mice were similar to those seen for RIF-1 tumors in C3H/HeJ mice. This acute vascular injury was accompanied by progressive edema in tumors but not in skin or muscle. In RIF-1 tumors, the extracell water volume at 12 h after IL-1 (395 microI/g) was nearly twice that in untreated controls (215 microI/g). IL-1 also produced marked reductions in tumor blood flow as early as 1 h after treatment. Maximal blood flow restriction was seen at 6 h after IL-1. Although restricted blood flow was observed in tumors for up to 48 h, IL-1 effects on muscle, liver, and skin blood flow were transient with recovery by 12 h after treatment. IL-1 (up to 0.4 ng/ml for 72 h) was not toxic to RIF-1 tissue culture cells in vitro, but 0.2 ng/ml IL-1, for 20 h, reduced the clonogenicity of RIF-1 cells in primary explant cultures by approximately 50%. In vivo, the clonogenic cellularity of RIF-1 tumors was reduced by 70%, 24 h after a single 25 micrograms/kg treatment. Increased clonogenic cell proliferation was observed at 24 h, and rapid repopulation of the clonogenic cell population was seen by 48 h. Although IL-1 transiently slowed the growth of RIF-1 tumors, no significant regrowth delay was observed. In Panc02 tumors, cell proliferation was also inhibited after IL-1. Recovery, however, was delayed and occurred more slowly than in RIF-1 tumors. Significant growth inhibition and regrowth delay (5 days) was observed in Panc02 tumors after a single IL-1 treatment. The results of these studies show that IL-1 has significant effects on the pathophysiology of both RIF-1 and Panc02 tumors in vivo. Further, our results indicate that these effects may be mediated through the activation of a non T-cell, adherent cell population residing in the tumor at the time of IL-1 treatment.     

Enhancing effect of misonidazole on the response of the RIF-1 tumour to cyclophosphamide.

The effect of misonidazole (MISO) on the cytotoxicity of cyclophosphamide (CY) was investigated in the mouse. The response of the RIF-1 tumour was measured by growth delay and by cell survival in a cloning assay. MISO enhanced the cytotoxicity of CY. For single treatment, enhancement was maximal when MISO was given 30 min to 2 h before CY. The enhancement ratio (i.e. the dose of CY alone divided by the dose of CY with MISO required to cause the same response) increased with increasing dose of MISO up to 250 mg/kg, but decreased with increasing dose of CY above 50 mg/kg. For 5 daily treatments, enhancement increased with CY dose up to approximately 25 mg/kg/injection. Survival of marrow stem cells was measured using the spleen-colony assay. MISO did not enhance significantly the cytotoxicity of CY at doses under 100 mg/kg. Enhancement was seen at higher doses, but the effect was less than in tumours. CY reduced the number of circulating white blood cells. Neutrophils were most severely depleted. The WBC count was slightly lower when CY was given in combination with MISO than after CY alone, but the effect could be accounted for by direct MISO cytotoxicity. These experiences suggest that a therapeutic gain may be achieved if MISO is combined with doses of CY in the clinical range. From experiments performed to investigated the possible mechanisms involved, we conclude that for the RIF-1 tumour the major effect of MISO is to inhibit the repair from CY-induced potentially lethal damage.     

Quantitative T1rho magnetic resonance imaging of RIF-1 tumors in vivo: detection of early response to cyclophosphamide therapy.

This study compares two potential magnetic resonance imaging (MRI) indices for noninvasive early detection of tumor response to chemotherapy: the spin-lattice relaxation in the rotating frame (T1rho) and the transverse relaxation time (T2). Measurements of these relaxation parameters were performed on a s.c. murine radiation-induced fibrosarcoma (RIF-1) model before and after cyclophosphamide treatment. The number of pixels exhibiting T1rho values longer than controls in viable regions of the tumor increased significantly as early as 18 h after drug administration and remained elevated up to 36 h after treatment (P < 0.005). Although a trend of increasing T2s relative to controls was noted in viable regions of the tumor 36 h after treatment, the changes were not statistically significant. Histological examination indicated a decrease in mitotic index that paralleled the changes in T1rho. We conclude that T1rho measurements may be useful for noninvasive monitoring of early response of tumors to chemotherapy.     

Effect of misonidazole or metronidazole pretreatment on the response of the RIF-1 mouse sarcoma to melphalan, cyclophosphamide, chlorambucil and CCNU.

The effect has been studied of adding either misonidazole (MISO) or metronidazole (METRO) to cytotoxic drug treatment of C3H mice bearing the RIF-1 sarcoma. The nitroimidazoles were injected 30 min before the cytotoxic drugs at a dose of 2 . 5 mmol/kg. Both clonogenic-cell survival and growth delay were measured as indicators of tumour response and depression in WBC count and acute lethality were used to indicate normal-tissue response. For melphalan, neither pretreatment agent produced any change in tumor response. For cyclophosphamide, no change was produced by METRO but a minimal increase in tumour response occurred with MISO. An enhancement of cell killing by CCNU was seen with MISO pretreatment, but there was no increase in tumour growth delay. METRO, however, did not enhance tumour response by either endpoint. WBC depression by CCNU was not enhanced by MISO pretreatment, and there was no significant reduction in the acute LD50. This indicates a therapeutic advantage from the addition of MISO to CCNU in this model system. For chlorambucil, considerable enhancement of tumour response followed either MISO or METRO pretreatment (dose-modifying factors of 2 . 0 and 1 . 4 respectively). However, the modification by MISO of normal-tissue response to chlorambucil was also enhanced by about a factor of 2, with no therapeutic gain.     

Application of 23Na MRI to monitor chemotherapeutic response in RIF-1 tumors

Effects of an alkylating anticancer drug, cyclophosphamide (Cp), on 23Na signal intensity (23Na SI) and water apparent diffusion coefficient (ADC) were examined in subcutaneously-implanted radiation-induced fibrosarcoma (RIF-1) tumors by 23Na and 1H magnetic resonance imaging (MRI). MRI experiments were performed on untreated control (n = 5) and Cp-treated (n = 6) C3H mice, once before Cp injection (300 mg/kg) then daily for 3 days after treatment. Tumor volumes were significantly lower in treated animals 2 and 3 days posttreatment. At the same time points, in vivo MRI experiments showed an increase in both 23Na SI and water ADC in treated tumors, whereas control tumors did not show any significant changes. The correlation between 23Na SI and water ADC changes was dramatically increased in the Cp-treated group, suggesting that the observed increases in 23Na SI and water ADC were caused by the same mechanism. Histologic sections showed decreased cell density in the regions of increased 23Na and water ADC SI. Destructive chemical analysis showed that Cp treatment increased the relative extracellular space and tumor [Na+]. We conclude that the changes in water ADC and 23Na SI were largely due to an increase in extracellular space. 23Na MRI and 1H water ADC measurements may provide valuable noninvasive techniques for monitoring chemotherapeutic responses.     

Effects of hyperglycemia on oxygenation,radiosensitivity and bioenergetic status of subcutaneous RIF-1 tumors

Since tissue oxygen tension is a balance between delivery and consumption of oxygen, considerable effort has been directed at increasing the former and/or decreasing the latter. Techniques to decrease the rate of cellular oxygen consumption (increasing the distance oxygen can diffuse into tissues) include increasing glycolysis by administering supra-physiologic levels of glucose. We have examined the effect of hyperglycemia produced by intravenous glucose infusion on the tissue oxygenation and radiation response of subcutaneously implanted murine radiation induced fibrosarcomas (RIF-1). A 0.3 M glucose solution was delivered via tail vein injection according to a protocol that maintained glucose at a plasma concentration of 17+/-1 mM. The effect of this treatment on radiation response (clonogenic and growth delay studies), tumor oxygenation (needle electrode pO2 and 2-[2-nitro-1H-imidazol-1-yl]-N-(2,2,3,3,3-pentafluoropropyl) acetamide (EF5) binding), and tumor bioenergetics and pH (31P NMR spectroscopy) was examined. Systemic measurements included hematocrit and blood glucose and lactate concentrations. The results of these studies suggest that these subcutaneously implanted RIF-1 tumors are both radiobiologically and metabolically hypoxic and that intravenous glucose infusion is not an effective method of modifying this metabolic state.     

Comparison between tumor pH and cell sensitivity to heat in RIF-1 tumors

We have performed a direct comparison between tumor pH and cell survival in heated RIF-1 tumors growing intradermally in thighs of C3H mice. The pH of individual tumors was measured by inserting a microelectrode into the tumor center. After pH measurements, the tumor-bearing legs were heated in a water bath (44.6 degrees C, 30 min). We then excised a small piece of tumor tissue (20-30 mg) from the area where the tip of the microelectrode had been placed and cell survival was determined by in vitro cloning. In some animals, 3 or 5 g/kg of glucose were injected i.p. 1.5 h before heating, to decrease the tumor pH. The average pH before heating was 6.83, 6.68, and 6.51, respectively, for tumors in untreated animals, those given 3 g/kg glucose, or those given 5 g/kg glucose. After heating, the average surviving fractions were 9.91 x 10(-2), 4.72 x 10(-2), and 5.43 x 10(-3), respectively. The cell yield did not differ significantly among the three treatments. As the tumor pH decreased, the surviving fraction also decreased for each treatment group. The correlation coefficient between tumor pH and log surviving fraction was highly significant for heat plus 5 g/kg glucose and all the treatment groups. The slope of the regression line obtained by a least squares method was steepest for all the treatment groups, followed by the heat plus 5 g/kg glucose and heat plus 3 g/kg glucose groups. The smallest slope of the regression line was noted for tumors treated by heat alone. The study shows that the tumor sensitivity to heat is enhanced when the tumor pH is lower and that adaptation of cells to low pH conditions may play a role in determining the relationship between pH and cell kill in RIF-1 tumors.     

31P-nuclear magnetic resonance studies of the effect of recombinant human interleukin 1 alpha on the bioenergetics of RIF-1 tumors

The effect of a single injection of human recombinant interleukin 1 alpha (IL-1 alpha) on s.c. RIF-1 tumors in mice was studied by in vivo 31P nuclear magnetic resonance spectroscopy. Spectra were obtained before and up to 24 h after IL-1 alpha. At 2, 4, 6, and 8 h after IL-1 alpha injection, RIF-1 tumors exhibited a reduction in bioenergetic status compared to untreated controls. The Pi to beta-nucleoside triphosphate and the phosphomonoester to beta-nucleoside triphosphate ratios increased, while the phosphocreatine to Pi and phosphodiester to phosphomonoester ratios decreased. Tumor blood flow, estimated by 86RbCl uptake, decreased within 30 min after IL-1 alpha treatment. Minimum perfusion was detected at 4 h, with recovery between 6 and 12 h after IL-1 alpha treatment. Histological sections of the RIF-1 tumors revealed intravascular congestion by 2 h, extravascular hemorrhage by 4 h, and necrosis by 12 h after treatment with IL-1 alpha. The time course of bioenergetic changes in RIF-1 tumors determined by 31P-NMR spectroscopy was found to parallel the reduction and subsequent recovery of tumor blood flow.     

Reoxygenation in the RIF-1 tumor

The proportion of hypoxic cells in the RIF-1 tumor was examined for 13 days following a 15 Gy conditioning dose. The paired survival curve technique indicated that 100% of the surviving cells were hypoxic immediately following this treatment. However, within 1 hour, only about 50% remained hypoxic, this proportion continued to drop to about 10% but did not reach the pretreatment level of 1.1% for the duration of the study.     

Phase I trial of misonidazole (NSC#261037) plus cyclophosphamide in solid tumors

Misonidazole, a hypoxic cell sensitizer, enhances the antitumor effects of cyclophosphamide in preclinical studies. Several studies also showed increased cytotoxicity for normal tissues. We undertook a phase I study of this combination. The regimen consisted of oral administration of misonidazole at one of two dose levels, 1 g/m2 and 2 g/m2, followed by an intravenous (IV) injection of cyclophosphamide four hours later. The cycle was repeated every twenty-one days. The dose of misonidazole remained constant for each regimen, but the dose of cyclophosphamide ranged from 0.4 g/m2 to 1.3 g/m2. Thirty-eight trials in 35 patients with advanced solid tumors were considered evaluable. Dose-limiting toxicity was granulocytopenia at 1 g/m2 of cyclophosphamide without significant thrombocytopenia or anemia. Peripheral neuropathy was negligible. Two patients received cumulative doses of 8 and 16 g/m2 of misonidazole without neurotoxicity. One patient developed hemorrhagic cystitis. Nausea and vomiting was mild to moderate. Possible evidence of tumor stabilization was seen in three patients, and one patient had a mixed response. The mean serum half-life for misonidazole was 11.3 hours (range, 8.4 to 20.0) and for cyclophosphamide 8.3 hours (range, 3.2 to 15.5), both within the previously reported ranges. In conclusion, it appears that this combination is well tolerated and that misonidazole does not significantly potentiate myelotoxicity caused by cyclophosphamide or alter its pharmacokinetics. The recommended starting doses for misonidazole and cyclophosphamide in phase II trials using this schedule of administration should be 2 g/m2 and 1 g/m2, respectively, with escalation for cyclophosphamide to individual tolerance.     

Drug resistance following irradiation of RIF-1 tumors: influence of the interval between irradiation and drug treatment

RIF-1 tumors contain a small number of cells (1 to 100 per 10(6) cells) that are resistant to 5-fluorouracil, methotrexate, or adriamycin. The frequency of drug-resistant cells among individual untreated tumors is highly variable. Radiation, delivered in vivo at doses of 3 to 12 Gy, increases the frequency of methotrexate- and 5-fluorouracil-resistant cells, but not the frequency of adriamycin-resistant cells. The magnitude of induction of 5-fluorouracil and methotrexate resistance shows a complex dependence on the radiation dose and on the interval between irradiation and assessment of drug resistance. For a dose of 3 Gy, induced 5-fluorouracil and methotrexate resistance is seen only after an interval of 5 to 7 days, whereas for a dose of 12 Gy, high levels of induced resistance are observed 1 to 3 days after irradiation. The maximum absolute risk for induction of resistance is 4 per 10(4) cells per Gy for methotrexate, and 3 per 10(6) cells per Gy for 5-fluorouracil. These results indicate that tumor hypoxia may play a role in the increased levels of drug resistance seen after irradiation, and that both genetic and environmental factors may influence radiation-induction of drug resistance. These studies provide essential data for models of the development of tumor drug resistance, and imply that some of the drug resistance seen when chemotherapy follows radiotherapy may be caused by radiation-induced drug resistance.     

Reoxygenation of the RIF-1 tumor after fractionated radiotherapy

The regrowth delay assay was used to assess hypoxic fraction in the RIF-1 tumor. Results approximating those of earlier paired survival curve data were obtained for previously untreated tumors and tumors treated with a single dose of 15 Gy. Further studies showed that after 5 daily fractions of 5 Gy, the hypoxic fraction returned to approximate pretreatment values within 24 hr.     

Does cyclophosphamide (CPM) improve survival rates in patients with solid tumors?

Cyclophosphamide (CPM) was first synthesized in 1958 by Arnold and Bourseaux as part of a program to develop new anticancer agents with fewer side effects and greater efficacy than those in existence. There are now many reports of good response rates obtained with CPM in patients with both hematological and solid malignant conditions, and it has been widely accepted as an important component of multi-drug regimens. What remains to be proven, however, is whether CPM significantly improves survival expectancy when used either alone or in combination with other agents. It should be remembered that evidence of efficacy as a single agent in terms of response rate or even complete remission may not be translated into improved survival when that agent is incorporated into a multi-drug regimen. Before a chemotherapeutic agent is evaluated as part of a multi-drug regimen there should be adequate evidence of efficacy against the relevant tumor. In addition, the agent must be used at a dosage that has been established as adequate. Despite these safeguards, it is not necessarily true that the addition of the drug either alone or as part of a multi-drug regimen will improve survival. Further use of the agent alone or in combination would then be illogical, just as it would be to use agents that have been shown to be ineffective, or to use an agent at too low a dose. The purpose of this article is to review the accumulated experience with CPM in both adult and pediatric oncology in the treatment of solid tumors with the above considerations in mind.     

FLOT-1在人类实体肿瘤中的研究进展

浮舰蛋白-1（flotillin-1,FLOT-1）属于脂筏标记蛋白,参与多种细胞生理活动,与许多肿瘤发生发展密切相关。FLOT-1在乳腺癌、食管鳞状癌、肾细胞癌（renal cell carcinoma,RCC）、移行细胞癌（transitional cell carcinoma,TCC）、非小细胞肺癌（nonsmall cell lung cancer,NSCLC）、肝细胞癌等肿瘤中均高表达,并与临床分期、转移、浸润和预后相关。本文综述了FLOT-1的结构和功能,及其在多种人类实体肿瘤中的作用。      

Reoxygenation in the RIF-1 tumor after chemotherapy.

The proportion of hypoxic cells in the RIF-1 tumor was observed for 24 hr after treatments with bleomycin, cisplatin, cyclophosphamide, and mitomycin C. The assays were based upon the paired survival curve method for determining the hypoxic fraction using irradiation of aerobic and artificially hypoxic tumors. It was observed that at 1/2 hr after bleomycin, hypoxic fraction was elevated but returned to baseline levels by 2 hr. Following cisplatin, hypoxic fraction did not rise above baseline. However, after cyclophosphamide, hypoxic fraction was elevated and did not return to baseline over the 24-hr observation period of this study. At 1/2 hr after mitomycin treatment, the hypoxic fraction was raised but within 1 hr returned to baseline. These results indicate that tumor reoxygenation varies after different drug treatments and that the determination of drug specificity for aerobic versus hypoxic cells may be strongly biased by the choice of the time after treatment for making the determination.     

活化的激酶C受体1与实体肿瘤

活化的激酶C受体1(RACK1)在多种肿瘤中有不同表达,因其含有的7个WD重复序列可与多种分子相互结合,RACK1除了可以激活蛋白激酶C,还在许多肿瘤相关信号通路中发挥作用.随着研究的不断深入,RACK1有望在肿瘤诊断、治疗以及预后等多方面发挥积极作用.     

19F magnetic resonance spectroscopy studies of the metabolism of 5-fluorouracil in murine RIF-1 tumors and liver

The metabolism of 5-fluorouracil (5FU) in tumors and livers of RIF-1 tumor-bearing C3H mice given i.p. injections of 5FU was serially monitored by 19F magnetic resonance spectroscopy. The levels of 5FU and fluoronucleotide detected in the tumors after a dose of 130 mg/kg (n = 13) were less than one-third of those after 260-mg/kg 5FU (n = 14). During the days after these doses, tumor size decreased by 24 +/- 3 and 52 +/- 6 SEM%, respectively. A second 130-mg/kg dose, given at day 7 after the first 130-mg/kg dose, resulted in still lower tumor fluorine levels and little change in tumor size. There was a significant correlation between the magnetic resonance spectroscopy-detected fluoronucleotide levels and the shrinkage of tumors after the 260-mg/kg dose (r = 0.44; P = 0.024). In mouse liver, the degradation of 5FU to alpha-fluoro-beta-ureidoprobionic acid and alpha-fluoro-beta-alanine after the 260-mg dose (n = 13) was slower than after a dose of 130 mg/kg (n = 14). For the respective doses, the half-life of 5FU was 59 +/- 7 versus 28 +/- 2 SEM min (P less than 0.0001). There was a negative correlation between the levels of 5FU catabolite (alpha-fluoro-beta-ureidoprobionic acid and alpha-fluoro-beta-alanine) in liver and fluoronucleotide in tumor (r = -0.80; P = 0.0020), which indicates that the degradation in liver and the activation of 5FU in tumor are competing processes.     

Sensitivity of normal mouse marrow and RIF-1 tumour to hyperthermia combined with cyclophosphamide or BCNU: a lack of therapeutic gain.

The effect of simultaneous whole-body heat (45 min 41 degrees C) on cyclophosphamide (CTX) and BCNU toxicity to normal mouse marrow stem cells and to the RIF-1 tumour in C3H/He mice has been studied. Marrow stem-cell survival was assayed by the spleen-colony technique at both 2 and 24 h after treatment, and also by following peripheral WBC count during the weeks after treatment. Heat potentiation of CTX toxicity to marrow stem cells was similar at both times and 24 h after treatment heat was dose-modifying with a DMF of 2.0. The heat potentiation of BCNU toxicity to stem cells was much greater at 24 h than at 2 h, and at 24 h had a DMF of 2.1. Peripheral WBC counts supported the results from 24 h assay for both drugs. RIF-1 tumour response was assayed by clonogenic cell survival measured 24 h after treatment, and by growth delay. For clonogenic tumour-cell survival after CTX, heated and unheated curves were parallel at doses above 75 mg/kg, yielding DMFs varying between 1.9 and 1.4 according to dose. DMFs for BCNU were also dose-dependent, lying between 2.0 and 1.6, the RIF-1 tumour being much less sensitive to BCNU than to CTX. Growth-delay data agreed with clonogenic cell survival. Therapeutic ratios for the combination of heat with CTX or BCNU fell in the range 0.91--0.69, according to dose, i.e. no gain or even therapeutic loss under the conditions of this study.