Gerrit Los

Pentetration of carboplatin and cisplatin into rat peritoneal tumor nodules after intraperitoneal chemotherapy

SummaryPlatinum distribution was studied in rat peritoneal tumors after i.p. treatment with equimolar doses of carboplatin and cisplatin. Low platinum concentrations (4 ppm) were detected in the periphery of the tumor after carboplatin treatment, whereas no platinum was detected 0.5 mm in from the periphery. In contrast, after cisplatin treatment, high platinum concentrations (29 ppm) were measured in the periphery of the tumor and moderate concentrations (14 ppm) were measured in the center. Only following increased carboplatin doses were low platinum concentrations detectable in the tumor. The total platinum concentration in the tumors was determined after equimolar administration of both drugs. In all, 7 times more platinum was detected after cisplatin treatment than after carboplatin treatment, and 10 times more carboplatin than cisplatin had to be injected to obtain comparable platinum concentrations in the tumors. When single cells were incubated with equimolar concentrations of carboplatin and cisplatin, 6–7 times more platinum was found in cells treated with cisplatin. However, pharmacokinetic studies favored i.p. administration of carboplatin because the clearance of this compound from the peritoneal cavity, expressed ast1/2β, was lower than that of cisplatin (239 vs 78 min), resulting in an AUC in the peritoneal cavity for both total and ultrafiltered drug that was almost 3 times higher for carboplatin than cisplatin. The AUC for ultrafiltered carboplatin in plasma was 2-fold that for cisplatin (2,801±210 vs 1,334±431 μM m). The present study demonstrated that in spite of the pharmacological advantages of carboplatin, its capacity to penetrate into peritoneal tumors and tumor cells is far lower than that of cisplatin.

Platinum distribution inintraperitoneal tumors afterintraperitoneal cisplatin treatment

SummaryThe spatial distribution of platinum (Pt) in the kidney was studied by an autoradiographic technique, in which cisplatin (CDDP) was replaced by195mPt-labeled CDDP, and by proton-induced X-ray emission (PIXE). Although both studies demonstrated comparable spatial distribution patterns, PIXE had the advantage that Pt concentrations could be determined quantitatively, in contrast to the relative information obtained by the autoradiographic technique. Using PIXE, the distribution of Pt in i.p. tumors was studied after i.p. administration of CDDP. The highest Pt concentrations were always found on the periphery of tumors, indicating that the periphery was exposed to a higher drug concentration than the center. Dose was correlated to the concentration of CDDP at both the center and the periphery (r=0.99).The Pt concentration in the periphery was usually higher by a factor of 2–3 after i.p. administration than after i.v. treatment, whereas in the center of the tumor no concentration difference could be detected. The penetration depth of CDDP lay between 1 and 2 mm and was calculated from the differences in Pt concentration after i.p. and i.v. treatment. This indicates that the effective advantage of i.p. chemotherapy with CDDP in cases of cancers limited to the peritoneal cavity is accentuated at the periphery of the tumor.

Optimisation of intraperitoneal cisplatin therapy with regional hyperthermia in rats

The purpose of this study was to optimise intraperitoneal chemotherapy by combining this modality with regional hyperthermia. In vitro data demonstrated that both the uptake of cisplatin into CC531 tumour cells and cytotoxicity were increased at temperatures of 40 degrees C (factor 4) and 43 degrees C (factor 6) compared to 37 degrees C. The increase of intracellular platinum concentration correlated well with the decrease in survival of these cells. In vivo, rats were treated intraperitoneally with cisplatin (5 mg/kg) in combination with regional hyperthermia of the abdomen (41.5 degrees C, 1 h). The mean (S.D.) temperature in the peritoneal cavity was 41.5 (0.3) degrees C and outside the peritoneal cavity 40.5 (0.3) degrees C. Enhanced platinum concentrations were found in peritoneal tumours (factor 4.1) and kidney, liver, spleen and lung (all around a factor 2.0), after combined cisplatin-hyperthermia treatment. The platinum distribution in peritoneal tumours was more homogeneous after the combined treatment than after cisplatin alone, possibly due to increased penetration of cisplatin into peritoneal tumours. Pharmacokinetic data demonstrated an increased tumour exposure for unfiltered platinum in the peritoneal cavity (area under the curve [AUC] increased from 339 mumol/l/min to 486 mumol/l/min at 37 degrees C and 41.5 degrees C, respectively), and for total and ultrafiltered platinum in the blood. The AUC for total platinum increased from 97.9 to 325.8 mumol/min and for ultrafiltered platinum from 22.2 to 107 mumol/l/min at 37 degrees C and 41.5 degrees C respectively. The latter might be due to a slower elimination of platinum from the blood. The combined treatment, intraperitoneal cisplatin and regional hyperthermia, also increased toxicity. The thermal enhancement ratio (TER) using lethality as endpoint was 1.8.

Effects of temperature on the interaction of cisplatin and carboplatin with cellular DNA

Increased levels of cisplatin (cDDP)- and carboplatin (CBDCA)-DNA adducts were detected in cDDP (10 microM)- and CBDCA (6 mM)-treated CC531 cells when the temperature was raised from 37 degrees to 43 degrees. In the case of cDDP, increased DNA adduct formation was already detectable at 38.5 degrees; additional temperature steps led to further increases in DNA modification. Increased CBDCA-DNA adduct formation was observed only at temperatures higher than 40 degrees. In vitro studies on the interaction of CDDP and CBDCA with isolated salmon sperm DNA, however, demonstrated no significant differences in the DNA binding rate between 37 degrees and 43 degrees for cDDP and a minor effect for CBDCA only at 43 degrees, almost totally excluding a direct temperature effect on DNA platination in this temperature range. Furthermore, neither the stability of the formed platinum-DNA adducts nor the rate of adduct loss in CC531 cells was changed at higher temperatures. The observed difference in cellular adduct formation, however, could be related to increased uptake of cDDP and CBDCA into CC531 cells at higher temperatures. In the case of cDDP, a temperature shift from 37 degrees to 38.5 degrees resulted in a significantly higher intracellular platinum concentration (0.03 +/- 0.01 vs 0.071 +/- 0.021 micrograms platinum/10(6) cells, respectively); for CBDCA, temperatures > or = 41.5 degrees were needed to increase the platinum concentration significantly above 37 degree values (0.3 +/- 0.1 vs 0.6 +/- 0.1 micrograms platinum/10(6) cells, respectively). In addition, the increase in DNA adduct formation of cDDP and CBDCA at elevated temperatures was comparable with the increase in cDDP-DNA adducts after a cDDP concentration escalation at 37 degrees, indicating a concentration-dependent increase in cDDP-DNA adducts. It seems that heat affects primarily the cellular uptake of cDDP and CBDCA and not their covalent binding to DNA.

CELLULAR PHARMACOKINETICS OF CARBOPLATIN AND CISPLATIN IN RELATION TO THEIR CYTOTOXIC ACTION

We have studied the cellular pharmacokinetics of carboplatin (CBDCA), as part of the evaluation of the antitumor activity of CBDCA in cancers limited to the peritoneal cavity in comparison with cisplatin (cDDP). The uptake of CBDCA into L1210 (lymphosarcoma), CC531 (colonic carcinoma), COV413.B (human ovarian carcinoma) and NB1 (human neuroblastoma) cells was 1.5 to 13 times lower than the uptake of cDDP. The uptake of CBDCA into human ovarian carcinoma cells, taken directly from patients, was also 8-20 times lower than cDDP. Platinum concentrations, expressed as a percentage of the total intracellular Pt concentration, were similar for CBDCA and cDDP in cytosol and nucleus/membrane fractions. A second major difference between the drugs was their binding to DNA. Less CBDCA-DNA than cDDP-DNA adducts were formed after incubation at equimolar amounts of drug with isolated salmon sperm DNA (5-25 times less). A 16-69 times higher concentration of CBDCA than cDDP was needed to induce similar changes in cell growth activity (50% [3H]thymidine inhibition) in CC531 and COV413.B cells, indicating that equitoxicity can only be achieved when tumor cells are exposed to higher concentrations of CBDCA than cDDP. Similar toxicity was achieved in CC531 cells after incubation with a 16-fold higher CBDCA dose than cDDP. Comparable intracellular platinum concentrations, however, were obtained with a 10-fold higher CBDCA dose, suggesting that cellular pharmacokinetics of the drugs are different. Regarding drug uptake and pharmacokinetics the mechanism of action of CBDCA differed from cDDP at a cellular level.

The use of oxaliplatin versus cisplatin in intraperitoneal chemotherapy in cancers restricted to the peritoneal cavity in the rat

Pharmacokinetic studies demonstrated the advantage of intraperitoneal oxaliplatin (1-OHP) for cancers restricted to the peritoneal cavity. The area under the concentration X time curve (AUC) in the peritoneal cavity for both total and ultrafiltered drug was almost 2 times higher for 1-OHP than cisplatin (cDDP). The AUC for ultrafiltered 1-OHP in plasma was also a factor 4 higher than cDDP, indicating that peritoneal tumors received a higher exposure from 1-OHP than cDDP directly in the peritoneal cavity and indirectly via the systemic circulation. Total platinum concentrations in peritoneal tumors of rats were determined after i.p. administration of equimolar doses of 1-OHP and cDDP. In spite of the pharmacological advantages, no significant difference in platinum concentration was demonstrated. In addition, no difference in the distribution of platinum within peritoneal tumors was detected after i.p. treatment with equimolar doses, i.e., platinum concentrations were comparable both in the periphery, 29 +/- 4 ppm for cDDP and 22 +/- 8 for 1-OHP and in the center of the tumor, 18 +/- 3 for both drugs. When CC531 tumor cells were incubated in vitro with equimolar concentrations of 1-DHP and cDDP in vitro, 2 to 4 times less platinum was found in cells treated with 1-OHP, indicating that the uptake of 1-OHP differed from that of cDDP. Oxaliplatin was not cross resistant for cDDP in CC531.RL4 tumor cells, a cDDP resistant cell line, which may indicate its value in ovarian cancer patients who did not respond to earlier cDDP treatment.

Intraperitoneal tumor growth and chemotherapy in a rat model.

Animal models are important to evaluate new treatment modalities. In the present paper a new animal model is described, in which the effects of intraperitoneal (i.p.) administration of cytostatic drugs on cancers restricted to the peritoneal cavity can be studied. The tumor cell line used is a chemically induced carcinoma (CC531), sensitive in vitro to cisplatin (cDDP), carboplatin (CBDCA), 5-fluorouracil (5-FU), doxorubicin and mitoxantrone. Three to 5 weeks after i.p. inoculation of 2 x 10(6) CC531 cells, 80% of Wag/Rij rats develop small tumor nodules on peritoneal surfaces. Both tumor size and localization at this time are comparable to the human situation, especially to cases of minimal residual disease ovarian carcinoma. The model has been used to determine the usefulness of i.p. treatment in comparison to i.v. Changing the route of administration of cDDP from i.v. to i.p. increases tumor platinum concentrations and prolongs survival. The model offers the possibility to study drug pharmacokinetics and tumor drug penetration related to i.p. drug administration.

ADRIAMYCIN-LOADED ALBUMIN-HEPARIN CONJUGATE MICROSPHERES FOR INTRAPERITONEAL CHEMOTHERAPY

Adriamycin-loaded albumin-heparin conjugate microspheres ~~R-~C~S) were evaluated as possible intraperitoneal (i.p.) delivery systems for site-specific cytotoxic action. The biocompatibility of the microspheres after intraperitoneal injection was tested first. 1 day after i.p. administration of empty as well as drug-loaded AHCMS to male Balb/c mice, only a moderate increase in i.p. neutrophils was measured. 3 days after injection neutrophil levels were comparable with the controls. No significant increases in the numbers of other cell types were observed, indicating an acute inflammatory response which can be considered to be mild. Antitumour efficacy was tested in an L1210 tumour-bearing mouse model and in a CC531 tumour-bearing rat model. The use of ADR-AHCMS leads to longer survival times of mice and improved tumour growth deIay in rats, as compared with untreated controls and free drug treated animals. In both animal models higher adriamycin doses were initially tolerated if the drug was formulated in microspheres, although long-term adriamycin toxicity effects were evident in all treated groups. Doses and dosage schedules may be optimized to further reduce the toxic effects of the drug.

The influence of hyperthermia on the uptake of cisplatin in the rat cervical spinal cord

The influence of local hyperthermia on the uptake of cisplatin in the rat cervical spinal cord was investigated. After single intraperitoneal or intravenous injection of cisplatin (5 mg/kg body weight), the spinal cord region cervical 5-thoracic 2 was heated for 60 min at mean (S.D.) 41.2 (0.4) degrees C or 40 min 42.4 (0.3) degrees C using a 434 MHz microwave heating device. One day after treatment with either hyperthermia alone, cisplatin alone or the combination, none of the animals expressed neurological symptoms. The spinal cord was dissected and platinum levels were measured by flameless atomic absorption spectroscopy. No difference was found in uptake of platinum in the spinal cord between control- and heat treated animals. In a second series of experiments, the spinal cord was heated for 30-60 min. during a 2 h infusion of cisplatin. One day after treatment at 42.3 degrees C for 60 min, neither motor nor sensory functions were affected and platinum levels did not differ significantly between control and treated animals. Also, platinum levels measured in the spinal cord immediately after cisplatin infusion were not influenced by heat treatment at 42.1 or 43.0 degrees C for 30 min. However, after a heat dose of 60 min 43 degrees C, cisplatin uptake was significantly increased (P less than 0.001) by a factor of 2.8 (1.3). The data demonstrate that mild hyperthermia has no effect on the uptake of cisplatin in the spinal cord, while an injurious heat dose leads to a significant increase in cisplatin uptake. The present findings indicate that, in case of treatment of tumours of the central nervous system with hyperthermia and cisplatin, a treatment which might be toxic for the tumour is well tolerated by the normal nervous tissue.

Combination treatment of cis- and carboplatin in cancers restricted to the peritoneal cavity in the rat

In the present study, cisplatin (cDDP) and carboplatin (CBDCA) were combined in different in vitro and in vivo assays to determine whether combined cDDP and CBDCA treatment would eventually lead to a better antitumor response. Co-incubation of CC531 cells with cDDP and CBDCA led to higher intracellular Pt concentrations (30.5±3.4 ng Pt/106 cells) than did cDDP (16.9±9.4 ng Pt/106 cells) or CBDCA (1.28±0.72 ng Pt/106 cells) incubation alone. In survival assays an additive cell kill was seen after combined treatment with cDDP and CBDCA. DNA binding experiments using isolated salmon-sperm DNA exposed to the drugs separately or in combination were in agreement with the survival studies (for cDDP a binding of 12.42 μg Pt/mg DNA; for CBDCA, 0.49 μg Pt/mg DNA at 76 h). Toxicity studies in rats treated with cDDP plus CBDCA required a dose reduction for cDDP amounting to 20% of the MTD, whereas the CBDCA dose could be maintained. Pharmacokinetics studies showed higher AUCs andt1/2β in plasma as well as the peritoneal cavity after combined treatment with cDDP and CBDCA (both given i.p.) or following cDDP given i.p. and CBDCA given i.v. Pt concentrations in peritoneal tumors corresponded with these observations, with higher Pt concentrations following combined treatment than after single-agent injection. In addition, combined adminstration of cDDP i.p. and CBDCA i.v. led to higher Pt concentrations in peritoneal tumors than did administration of both drugs i.p. (3.93±0.9 vs 2.76±0.2 mg Pt/g tissue). The higher Pt concentrations in the peritoneal tumors after combined treatment was associated with a significantly better antitumor response in comparison with that observed after single-agent treatment (a growth delay of 30.2±5.6 days for cDDP i.p. plus CBDCA i.v. vs 16.1±5.4 days for cDDP alone and 10.8±4.2 days for CBDCA alone).