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[CANCER RESEARCH 50, 6971-6975. November 1. 1990] Cytotoxicity, Radiosensitization, Antitumor Activity, and Interaction with Hyperthermia of a Co(III) Mustard Complex1 Beverly A. Teicher,2 Michael J. Abrams, Kristina W. Rosbe, and Terence S. Herman Dana-Farber Cancer Institute [B. A. T., K. W. R., T. S. H.J and Joint Center for Radiation Therapy fB. A. T., T. S. H.], Boston, Massachusetts 02II5, and Johnson-Matthey, Inc., West Chester, Pennsylvania 19380 [M. J. A.] ABSTRACT A complex of Co(III) with a nitro group and a bis(2-chloroethyl)amine moiety was prepared in an effort to develop a new anticancer agent with radiosensitizing capabilities, direct antitumor activity, and the ability to interact positively with clinically relevant hyperthermia temperatures. The activity of this drug was compared to a similar Co(III) complex, nitro-bis(2,4-pentanedionato)(pyridine)cobalt(III) (Co(Py)], which bears a pyridine moiety mustard of bis(2-chloroethyl)amine and should have no alkylating abilities. In EMT6 cells nitro-bis(2,4-pentanedionatoXbis(2-chloroethyl)amine)cobalt(III) (Co(BCA)j was significantly more cytotoxic than <'o(Py) and both drugs were more toxic toward normally oxygenated than hypoxic cells. Hyperthermia (42°C,1 h) increased the slope of the concentration-dependent survival curve for Co(BCA) but not for Co(Py) in normally oxygenated EMT6 cells. Co(BCA) was an effective radiosensitizer of hypoxic EMT6 cells in vitro, producing a dose-modifying factor of 2.40. In the human squamous cell line SCC-25 and the nitrogen mustard-resistant subline SCC-25/HN2 Co(BCA) was more cytotoxic than Co(Py), and the lethality of Co(BCA) was only minimally diminished in the SCC-25/HN2 line. In mice bearing the I 1210 leukemia i.p., Co(BCA) had a broad range of therapeutically effective dosage and produced a >60-day increase in life span at a dose 20-fold less than was lethally toxic. In addition, in the FSalIC murine fibrosarcoma, Co(BCA) produced a tumor growth delay of 9.4 days at 75 mg/kg i.p. daily x 5, but Co(Py) produced a delay of only 2.9 days at 50 mg/kg daily x 5 and was lethally toxic above this dose. These results indicate that Co(BCA) has significant antineoplastic effects in vitro and in vivo and interacts positively with both radiation and mild hyperthermia. Its broad therapeutic dose range further suggests potential clinical utility. INTRODUCTION Complexes of many transition metals have demonstrated cytotoxicity in cell culture and/or antitumor activity in tumorbearing animals (1-4). Only complexes of platinum, however, are currently in routine clinical use (5-10). A wide variety of metal complexes have also been shown to be radiation-sensitizers of mammalian and/or bacterial cells. These include com plexes of Ag(I), Cu(I), Cu(II), Zn(II), Hg(II), Rh(II), Ru(II), Pd(II), Pt(II), Ru(III), Co(III), and Fe(III) (11). We have pre viously examined the in vitro radiosensitizing potential of 100 ¿¿M concentrations of a series of 12 Co(III) complexes ranging in structure from hexanitro-Co(III) to hexammine-Co(III) (12). At this relatively low drug concentration, a wide range of radiosensitization activity was observed. Although the most active complexes in this study contained coordinated nitro ligands, the presence of nitro ligands on the complex was not a requirement for radiosensitizing capability (12). Complexes of Co(III) are kinetically inert octahedral coor dination complexes. This inertness is due to the </6 low spin electron configuration of trivalent cobalt (13). Kinetically inert Received 4/16/90; accepted 8/1/90. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1This work was supported by NIH Grant R01-CA36508. 2To whom requests for reprints should be addressed, at Dana-Farber Cancer Institute, 44 Binney St., Boston, MA 02115. transition metal complexes [e.g., Rh(III) and Cr(III)] undergo the water exchange reaction relatively slowly, with half-lives of about 1 day (14). In contrast, complexes of kinetically labile metal ions [e.g., Mn(II), Cr(II)] undergo this reaction with halflives of less than l s (14). The biological consequence of kinetic inertness is that many Co(III) complexes will remain intact when added to a culture medium or injected into animals and should arrive at their cellular targets with their ligand configu ration intact. In an effort to further increase antitumor activity and the radiosensitizing potential of Co(III) complexes, we have syn thesized a complex of Co(III) with an alkylating moiety. In this report we detail the synthesis, in vitro cytotoxicity, and initial in vivo antitumor studies with Co(BCA),3 which bears both a nitro group and a bis(2-chloroethyl)amine moiety. To evaluate the effect of the bifunctional alkylating group on Co(BCA), an analogous Co(III) complex bearing a pyridine moiety instead, Co(Py), was prepared and compared to Co(BCA). MATERIALS AND METHODS Preparation of Co(BCA). To prepare Co(BCA), 1.5 g of Na[Co(NO2)2(acetylacetonate)2] (4.6 mmol) were dissolved in water (60 ml) and the red-brown solution was filtered to remove residual insoluble material (15). To this stirred solution were added bis(2-chloroethyl)amine-HCl (0.82 g, 4.6 mmol) in water (10 ml) and sodium bicarbonate (0.39 g, 4.6 mmol) in water ( 10 ml). The solutions of BCAHC1 and NaHCOj were combined immediately before addition to the reaction mixture. After 30 min the brown microcrystalline product was collected, washed with ether, and dried in vacuo. The yield was 0.45 g (24% based on Co). The calculated atomic weight percentage compo sition for C,4H23CL2CoNo2O6 is C, 37.78%; H, 5.22%; N, 6.29%; Cl, 15.93%. Actual measurements yielded C, 37.92%; H, 5.10%; N, 6.54%; Cl, 16.22%. Co(Py) was prepared analogously and characterized as described previously (15). These materials are readily soluble in polar organic solvents. Both cobalt complexes are prepared freshly just prior to use in cell culture or animals. Mustargen (nitrogen mustard) was obtained from the Dana-Farber Cancer Institute pharmacy. Nor-nitrogen mustard [bis(chloroethyl)amine] was purchased from Aldrich Chemical Co (Milwaukee, WI). These agents were prepared freshly just prior to use in cell culture or animals. Cell Lines. The EMT6 mammary tumor cell line has been widely used for the study of antineoplastic agents (16-18). The experiments described here were performed using asynchronous EMT6 monolayers in exponential growth, in Waymouth's medium supplemented with antibiotics (Grand Island Biological Co., Grand Island, NY) and 15% newborn calf serum (Hyclone Laboratories, Logan, UT). This cell line has a plating efficiency of 65-80% and a doubling time of 16-19 h in vitro (\S). The SCC-25 human squamous cell carcinoma cell line retains an epithelioid appearance and grows without the aid of a feeder layer (19). It has a plating efficiency of 15-20%, as judged by plating single colonies on plastic. The cells were grown on Dulbecco's modified 3The abbreviations used are: Co(BCA), nitro-bis(2,4-pentanedionato)(bis(2chloroethyl)amine)cobalt(III); BCA, bis(2-chloroethyl)amine-HCl; Co(Py), nitrobis(2,4-pentanedionato)(pyridine)cobalt(III); PBS, phosphate-buffered 0.9% sa line. 6971 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1990 American Association for Cancer Research. ANTITUMOR Co(l][) MUSTARD COMPLEX Eagle's medium supplemented with 5% fetal bovine serum and anti biotics. For the SCC-25 line, hydrocortisone (0.4 Mg/ml) was included in the medium (20). The nitrogen mustard-resistant subline SCC-25/HN2 was developed through escalating selection pressure methodology (21). Nearly con fluent 100-mm1 dishes were treated with approximately the concentra tion of nitrogen mustard that would kill 90% of the cells in 1 h, washed 3 times with PBS, and covered with fresh medium plus serum. The dose of nitrogen mustard treatment was escalated at a rate of 15-20%/ week, and the cells were treated weekly unless there was no evidence of cell growth between treatments. After 14 months of treatment, clones were developed and screened for resistance to nitrogen mustard for doubling times similar to that of SCC-25 parent cell line (21). Other characteristics of the SCC-25/HN2 line have been described (22). Cytotoxicity Studies. EMT6 cells, SCC-25 cells, or SCC-25/HN2 cells in exponential growth were exposed for l h to concentrations ranging from 5 to 500 //M of the cobalt complexes, nitrogen mustard, or BCA. The cells were then washed 3 times with PBS and cloned as described previously (12, 16, 23). Hyperthermia Studies. Exponentially growing EMT6 cells were ex posed to the study drugs at 37°Cor 42°Cfor 1 h. Heating was mean ±SE for the treatment group compared to the control. Tumor volume was calculated as a hemiellipsoid. Untreated tumors reach 500 mm1 in 14.0 ±0.3 days (27, 28). In Vivo Data Analysis. Data on the delay of tumor growth were analyzed with a BASIC program for the Apple II + microcomputer. The program derived the best-fit curve for each set of data and then calculated the median, mean, and SE for individual tumor volumes and the day on which each tumor reached 500 mm\ RESULTS The structures of Co(BCA) and Co(Py) are shown in Fig. 1. The bulk of the pyridine ligand is similar to that of the bis-(2chloroethyl)amine ligand; however, the pyridine ligand does not have the bifunctional alkylating capability of the bis-(2-chloroethyl)amine. In vivo, Co(BCA) may be a prodrug, since in this complex the bis-alkylating moiety of Co(BCA) should be inac tivated via complexation of the nitrogen lone pair to the Co(III) ion. Intracellularly, free BCA should be released upon reduction of Co(BCA) to a Co(II) species. accomplished in a Plexiglas water tank with a continuous in flow and Co(BCA) was more cytotoxic toward EMT6 cells under both out-flow system controlled by a water temperature controller (Braun normally oxygenated and hypoxic conditions than was Co(Py) Thermomix 1460; Braun Instruments) (24). Cells underwent heating (Fig. 2). Both cobalt complexes were more cytotoxic toward in sealed plastic flasks (Falcon Plastics) containing 5 ml of complete medium plus drug. Water temperature could be maintained at ±0.10°C normally oxygenated EMT6 cells than toward hypoxic EMT6 cells. Co(BCA) at 100 MMfor 1 h killed about 1 log of normally (SD). oxygenated EMT6 cells. The concentration of Co(BCA) re Production of Hypoxia. To produce hypoxia, flasks containing EMT6 quired to kill 1 log of hypoxic EMT6 cells after a 1-h exposure cells in complete medium plus serum were fitted with rubber sleeve to the drug was about 140 ¿¿M. A drug concentration of about serum stoppers and exposed to a continuously flowing 95% nitrogen/ 5% CO2 humidified atmosphere for 4 h at 37°C(23). Parallel flasks 340 /UMCo(Py) for 1 h was required to produce 1 log of cell were maintained in 95% air/5% CO2. At this time, the drug (0.10 ml) kill of normally oxygenated EMT6 cells, while a concentration or vehicle (PBS, 0.10 ml) was added to the flasks by injection through of much greater than 500 UM Co(Py) for 1 h would have been the rubber stopper, without disturbing the hypoxia. needed to produce the same level of cell kill of hypoxic EMT6 Radiation Survival Studies. Hypoxia was produced as described cells. BCA shows no difference in cytotoxicity toward normally above. For radiation, the hypoxic flasks were placed in an insulated chamber at 37°Cfilled with 95% nitrogen/5% CO2 atmosphere. The cells were irradiated with a "7Cs radiation unit (Gammacell 40: Atomic Energy of Canada, Ltd.) at a dose rate of approximately 1.05 Gy/min. Drug-treated cells were irradiated so that the irradiation was complete 1 h after addition of the drug. X-ray doses of 5, 10, 15, and 20 Gy were used. The baseline oxygen enhancement ratio in the EMT6 cell line was 2.9 (12, 16). In Vitro Data Analysis. Quantitative analysis of dose-response curves for cell survival was performed using the log-probit iterative least squares method. This approach is a statistically rigorous and objective means for determining the slope of the dose-response curve. Correlation coefficients for the linear log-probit line and x2 analysis for goodness of fit were also calculated. The dose-modifying factors were calculated as the ratio of the slopes of the survival curves obtained with radiation alone and radiation in the presence of the test drug. In Vivo Studies. The LI210 leukemia was grown in DBA mice (The Jackson Laboratory, Bar Harbor, ME). For experiments, IO6 tumor cells were implanted i.p. on day 0. Drug treatment was begun on day 1 and drugs were administered daily for 5 days. Treatment outcome was assessed as increase in life span of treated compared to untreated control animals (25). The FSall fibrosarcoma (26) was carried in male C3H/He mice (The Jackson Laboratory). For the experiments, 2 x IO6tumor cells prepared from a brei of several stock tumors were implanted i.m. into the legs of male C3H/He mice 8 to 10 weeks of age. When the tumors were approximately 100 mm3 in volume, treatment was initiated. In those groups receiving drug, Co(BCA) (10, 25, 50, or 75 mg/kg) or Co(Py) (25, 50, or 75 mg/kg) in phosphate-buffered 0.9% saline (0.2 ml) was injected i.p. daily for 5 days. The progress of each tumor was measured thrice weekly. Tumor growth delay is the number of days for the tumors in treated animals to reach 500 mm,' compared to the tumor of untreated controls. Each treatment group had seven animals and the experiment was repeated 3 times. Days of tumor growth delay are the CH,CH,CÕ CH, HN IcH.CH.CI C-CH, NO, NO, Fig. I. Structures of Co(BCA) (left) and C'o(Py) (rijfAr). 1.0 t 0.01 0.001 50 10O 250 500 Drug Concentration, uM Fig. 2. Survival of EMT6 mouse mammary tumor cells treated with various concentrations of Co(BCA) for 1 h under normally oxygenated conditions (•)or hypoxic conditions (O) or treated with various concentrations of Co(Py) for 1 h under normally oxygenated conditions (•)or hypoxic conditions (C). The data are presented as means ±SE (bars) for three independent experiments. 6972 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1990 American Association for Cancer Research. ANTITUMOR Co(III) MUSTARD COMPLEX oxygenated or Hypoxie cells (data not shown). The cytotoxicity of both Co(BCA) and Co(Py) was increased when normally oxygenated EMT6 cells were exposed to the drugs at the clinically relevant hyperthermic temperature of 42°C(Fig. 3). The hyperthermia treatment itself resulted in a surviving fraction of about 0.65. Hyperthermia had a dosemodifying effect on the cytotoxicity Co(BCA); that is, there was an increase in the slope of the killing curve produced by Co(BCA) when the 1-h drug exposure was conducted under hyperthermic conditions. At a drug concentration of 100 //M Co(BCA), there was about a 7-fold increase in cytotoxicity at 42°Ccompared with 37°C,after correcting for 42°Cx l h lethality. On the other hand, the cytotoxicity of Co(Py) and hyperthermia appeared to be mainly additive. There was about a 1.8-fold increase in the cytotoxicity of Co(Py) in the presence of hyperthermia (42°C,1 h) over the entire concentration range of Co(Py) examined. Radiation survival curves for EMT6 cells in vitro in the presence and absence of the cobalt complexes are shown in Fig. 4. These results are drawn corrected for cytotoxicity of drug alone. The concentration of Co(BCA) and Co(Py) used for these radiation studies was 100 /J.Mand the drugs were present for l h prior to and during radiation delivery. Co(BCA) was an 0.001 so too 250 Drug Concentration, gM Fig. 3. Survival of EMT6 mouse mammary tumor cells treated with various concentrations of Co(BCA) for l h under normally oxygenated conditions at 37*C (•)or 42"C (O) or treated with various concentrations of Co(Py) for l h under normally oxygenated conditions at 37'C (•)or 42°C(D). The data are presented as means ±SE (bars) for three independent experiments. O.OO01 20 10 X-Ray Dose, Qray 20 Fig. 4. Radiation survival of EMT6 mouse mammary tumor cells in the presence of Co(BCA) or Co(Py). •.no drug, normally oxygenated cells; O, no drug, hypoxic cells: •100 /IM drug, normally oxygenated cells; D, 100 MMdrug, hypoxic cells. The data are presented as means ±SE (bars) for three independent experiments and are corrected for cytotoxicity of drug alone. effective radiosensitizer of hypoxic EMT6 cells, resulting in a dose-modifying factor of about 2.40. In contrast, Co(Py) was not an effective radiosensitizer. The presence of 100 /¿M Co(Py) did not significantly alter the slope of the radiation survival curve of hypoxic EMT6 cells. Neither Co(BCA) or Co(Py) had any significant effect on the radiation survival of normally oxygenated EMT6 cells. As shown in Fig. 5, the SCC-25/HN2 nitrogen mustardresistant subline of the SCC-25 human squamous carcinoma cell line is approximately 9-fold resistant to nitrogen mustard at a level of 50% cell kill. Co(BCA) was less toxic toward the SCC-25 and SCC-25/HN2 cell lines than toward the EMT6 murine cell line. The drug concentration which resulted in 1 log of cell killing by Co(BCA) upon exposure to the drug for 1 h was about 200 MMin the SCC-25 cell line and about 280 fiM in the SCC-25/HN2 cell line. Therefore, the SCC-25/HN2 cell line was only about 1.27-fold resistant to Co(BCA) at a level of 90% cell kill, compared to the SCC-25 cell line. The cytotoxicity of Co(Py) in these human tumor cell lines was somewhat less than that of Co(BCA), as was the case in EMT6 cells. Exposure to a concentration of about 465 pM Co(Py) for l h resulted in 1 log of cell killing in the SCC-25 cell line and exposure to a concentration of about 490 ^M for l h produced the same level of cell killing in the SCC-25/HN2 cell line. As expected, therefore, there was no significant difference in the sensitivity of these two cell lines to Co(Py). BCA is the mustard moiety present in the Co(BCA) complex. The SCC-25/HN2 cell line was about 2-fold more resistant to the cytotoxic actions of BCA than was the SCC-25 cell line. The concentrations of BCA which resulted in 1 log of cell killing in each cell line were about 140 and 280 Õ/Min the SCC-25 and SCC-25/HN2 cell lines, respectively. The in vivo antitumor activity of the cobalt complexes was assessed in L1210 leukemia (Table 1). LI210 cells (IO6) were implanted i.p. in DBA mice on day 0. Treatment was begun on day 1 and the drugs were administered daily for 5 days. Co(BCA) was administered over a wide dosage range and was highly effective against this tumor at doses from 5 to 50 mg/kg daily for 5 days, where mean survivals of >60 days were produced. The antitumor activity of Co(BCA) decreased at doses of 1 mg/kg and less and it was toxic at doses of 100 mg/ kg and greater. Co(Py) was ineffective at a dose of 5 mg/kg and showed moderate activity at doses of 10 and 50 mg/kg, but there were no 60-day survivors in any groups of animals treated with Co(Py). Nor-nitrogen mustard (BCA) was most effective at a dose of 50 mg/kg but was not nearly as successful as Co(BCA) and appeared to have a much steeper dose-response curve in this tumor system than did Co(BCA) or Co(Py). Using a similar experimental design, tumor growth delay studies were conducted in the FSallC fibrosarcoma (Table 2). Treatment was begun when the tumors were about 100 mm3 in volume and the drugs were administered daily for 5 days. Co(BCA) treatment resulted in measurable growth delays in the FSallC fibrosarcoma over the dosage range examined, from 10 to 75 mg/kg. Tumor growth delay continued to increase with increasing dose of the drug. At 75 mg/kg, Co(BCA) produced a growth delay of about 9.4 days. Co(Py) was only minimally effective against the FSallC fibrosarcoma at a dose of 50 mg/kg and at that dose was about 2.8-fold less effective than Co(BCA). DISCUSSION Complexes of a wide variety of metals have undergone preclinical testing as anticancer agents (1-3). Some of these new 6973 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1990 American Association for Cancer Research. ANTITUMOR Co(III) MUSTARD COMPLEX 1.0 Fig. 5. Survival of SCC-25 (•)and SCC25/HN2 (O) human squamous carcinoma cells exposed to various concentrations of nitrogen mustard, Co(BCA), Co(Py), or BCA. The data are presented as means ±SE (ears) for three independent experiments. 0.01 100 260 600 100 250 500 100 250 600 Drug Concentration, MM Table 1 Survival of mice bearing LI 210 leukemia treated with Co(BCA), Co(Py), or BCA Controls from four experiments survived 8.4 ±0.6 days. Tumor cells (10*) were implanted i.p. on day 0. Drugs were administered on days 1-5. the study contained coordinated nitro ligands, the presence of nitro ligands on the complex was not a requirement for radi osensitizing activity. All of the trinitrotiamine-containing com plexes were very effective radiosensitizers, whether the amine dose was NH3, CH.,NH2, or the chelating ligand diethyltriamine. DrugCo(BCA)Co(Py)BCADaily (mg/kg)250100SO2510510.5O.I1005010510050105Survival (days)4.4 Within the nitroamine series, increasing the number of nitro 1.015.6 ± ligands led to increased radiosensitizer activity. Co(BCA) is a 1.6>60>60>60>6044.5 ± Co(III) complex containing a nitroligand and a ligand capable of bifunctional alkylation upon release from the complex. The action of alkylating moiety in enhancing the cytotoxicity of this 2.422.3 ± new complex is evident in its increased cytotoxicity toward both 1.718.9 ± the EMT6 murine tumor cell line and the SCC-25 and SCC1.86.7 ± 25/HN2 human tumor cell lines, as compared to the pyridinecontaining analogue. The dose-modifying effect of hyperther0.529.0 ± 2.727.6 ± mia on the cytotoxicity of this complex is similar to patterns 2.48.9 ± observed with other alkylating agents and hyperthermia. 1.112.7 ± Whether the increase in cytotoxicity of Co(BCA) at elevated 1.538.8 ± temperature is due to increased intracellular drug levels, in 2.917.8 ± creased reactivity with DNA, or hyperthermia-induced inhibi 1.810.1 ± ±1.4T/C"Toxic1.8>7.1>7.1>7.1>7.15.32.62.2Toxic3.43.31.061.54.62.11.2 tion of DNA repair remains to be defined for this drug, as for * Survival of treated/survival of control. most other drugs which interact with hyperthermia (33). The Co(BCA) complex was an effective radiosensitizer of hypoxic EMT6 cells, with efficacy similar to that of cis-dinitroTable 2 Growth delay of the FSal 1Cfibrosarcoma produced by treatment with tetrammine-cobalt(III). The lack of radiosensitizing effect by Co(BCA) or Co(Py) Drugs were administered i.p. daily on days 7-11 after tumor cell implantation. the Co(Py) complex may be due to a failure of the complex to growth delay reach a position near enough to the DNA to allow the Co(III) DrugCo(BCA)Co(Py)Dose(mg/kg)10255075255075Tumor (days)3.1 moiety to participate in transfer functions. The particular con 1.04.4 ± figuration of this molecular may, itself, inhibit electron trans 1.28.2 ± fers. 1.69.4 ± Nitrogen mustard is transported into cells via the choline 1.51.2 ± transport system. The resistance of cells in culture to nitrogen ±0.72.9 mustard has often been attributed to a decrease in the transport 1.1Toxic ± of the drug into the cells (34). Co(BCA), Co(Py), and BCA may enter cells primarily by passive diffusion and, consequently, alteration in the choline transport system would not effect metal complexes, like the classical inorganic anticancer agent cellular sensitivity to these drugs. Co(BCA), therefore, would cis-diamminedichloroplatinum(II), have the potential to form be expected to be a mustard-containing antitumor agent which coordination bonds with cellular targets and would generally be is not cross-resistant with nitrogen mustard in the SCC-25/ classified an antitumor alkylating agents (1-4). Others, such as HN2 subline, as was essentially found in these studies. the chiral Co(III) Ã-rá-phenanthroline complexes, bind tightly Co(BCA) showed an unusually broad dosage range of activity and site-specifically to DNA (29-31). Some of these site-spe against both the LI210 leukemia and the FSal 1C fibrosarcoma. cific binding agents can be activated oxidatively to cleave the Co(BCA) was more efficacious and less toxic than Co(Py) in both murine tumor systems. Co(BCA) may be regarded as the DNA (32). lead complex for a new class of antitumor agents with both a The precise properties which result in optimal radiosensitizbifunctional organic alkylating capability and a metal compo ing ability in metal complexes are not known. We have previ ously examined the radiosensitizing potential of a series of 12 nent capable of radiosensization. Co(III) complexes, ranging in structure from hexanitro-Co(III) to hexammine-Co(III), in vitro (12). At the relatively low drug REFERENCES concentration of 100 ^M, a wide range of radiosensitization 1. Kopf-Maier, P. Cytostatische nicht-platinmetall-Komplexe: neue Perspek tiven fur die krebsbehandlung? Naturwissenschaften. 74: 374-382. 1987. activities was observed. Although the most active complexes in 6974 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1990 American Association for Cancer Research. ANTITUMOR Co(III) MUSTARD COMPLEX 2. Kopf-Maier, P., Neuse, E., Klapotke, T.. and Kopf, H. 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Cytotoxicity, Radiosensitization, Antitumor Activity, and Interaction with Hyperthermia of a Co(III) Mustard Complex Beverly A. Teicher, Michael J. Abrams, Kristina W. Rosbe, et al. Cancer Res 1990;50:6971-6975. Updated version E-mail alerts Reprints and Subscriptions Permissions Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/50/21/6971 Sign up to receive free email-alerts related to this article or journal. To order reprints of this article or to subscribe to the journal, contact the AACR Publications Department at [email protected]. To request permission to re-use all or part of this article, contact the AACR Publications Department at [email protected]. Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1990 American Association for Cancer Research.