Simultaneous determination of the peptide-mitomycin KW-2149 and its metabolites in plasma by high-performance liquid chromatography.

A gradient high-performance liquid chromatographic (HPLC) method is described for the quantification of KW-2149 and its two major metabolites in plasma. The method involves a sample clean-up by solid-phase extraction on C18 columns, separation of the respective compounds by HPLC on a YMC ODS-AQ column (5-microm particle size, 150x6 mm I.D.), using a methanol-water gradient system as an eluent, and measurement by UV absorbance detection at 375 nm. The limits of quantitation were 10 ng/ml for KW-2149 and M-16, and 15 ng/ml for M-18. Recoveries from plasma were higher than 92% on C18 extraction columns. Intra-day precision, expressed as %C.V., was between 1.4 and 6.5%. Intra-day accuracy ranged from 94 to 107%. Precision and accuracy of variability of inter-assays increased somewhat; however, were still within acceptable ranges. The ability of the method to quantify KW-2149 and two major metabolites simultaneously, with precision, accuracy and sensitivity, make it useful in monitoring the fate of this new mitomycin in cancer patients.

Determination of the new anticancer agent KW 2149, 7-N-[2-[2-(gamma-L-glutamylamino)ethyl)dithio)ethyl]mitomycin C, an analogue of mitomycin C.

The new mitomycin 7-N-[2-[2-(gamma-L-glutamylamino)ethyl)dithio)ethyl] mitomycin C (KW 2149) (I) proved to be active against a wide variety of experimental tumours. In order to perform pharmacokinetic studies with the new drug in Phase I sessions, a fast and reliable method has been developed based on the data of previous assays for mitomycin C. XAD-2 was preferred for isolation of I from blood plasma. The recovery of I was 50% whereas that of mitomycin C was 85%. Optimal separation was obtained on octadecyl silica columns with methanol-water (45:55, v/v) as mobile phase, while ultraviolet absorbance detection was performed at 375 nm. The assay enabled determination of I in a plasma concentration range of 20-1000 ng/ml using porfiromycin as internal standard.

[Pharmacokinetics and local availability of mitomycin. The influence of vasoconstriction and chemoembolization]. / Pharmakokinetik und lokale Verfügbarkeit von Mitomycin. Einfluss von Vasokonstriktion und Chemoembolisation.

The influence of the vasopressin-pro-drug glycylpressine (GP) and of a chemoembolisation with Spherex starch-particles (SP) on the availability of mitomycin (CAS 50-07-7, mitomycin C, MMC) was investigated in 30 patients with liver metastases, MMC administration was performed after blocking of the common hepatic artery by different concentrations of SP and after different time-intervals of GP. The comparison of plasma-levels after bolus injection without GP and after administration of MMC after 2, 5 and 15 min of vasoconstriction and after chemoembolisation with 450 mg or 900 mg SP, respectively, showed a remarkable reduction in the systemic circulation of MMC in the blood vessel system at about 40% (GP) and 45% (SP). A statistically significant influence on the pharmacokinetics of MMC with regard to CO, Vd, T 1/2zp, AUC and Cl(tot) was found, but not in t1/2el, t1/2biol and Vl. Both methods cause a distinctly accelerated diffusion of MMC into the tissue of the tumor region by change of the hemodynamics, leading to lowered side-effects. Thus the clinical picture was improved by MMC.

[Targeting cancer chemotherapy for metastatic liver cancer--effects of DSM on hepatic hemodynamics and on clinical outcome].

The effects of intra-arterial infusion of degradable starch microsphere (DSM) on hepatic hemodynamics were studied in 22 patients with metastatic liver cancer and the clinical outcome with mitomycin C (MMC) combined with DSM was reported herein. Hepatic arterial blood flow, measured with a transit-time ultrasonic blood flow meter, changed 283 +/- 27 ml/min to 40 +/- 36 ml/min by an hepatic arterial infusion of DSM and, a mean occlusion time aS 24 +/- 11 min. Combined infusion with DSM and MMC reduced MMC levels in the peripheral blood at 0.0248 less than p less than 0.0421, compared with those by an infusion with MMC alone and consequently, these findings proved to result from intrahepatic accumulation of MMC. RI-angiography using 99mTc-macroaggregated albumin (99mTc-MAA) was performed to examine hemodynamic changes in the metastatic liver and, a tumor (T) to non-tumor (N) ratio of 99mTc-MAA accumulation increased 0.37 to 0.62 by combined use of DSM. Thus, an intra-arterial infusion combined DSM and MCC was performed for 22 patients with unresectable hepatic metastases. Tumor regression was observed in 16 patients (73%). Side effects possibly attributable to DSM was transient nausea and vomiting. These results show that combined use of DSM is effective for intra-arterial chemotherapy against metastatic hepatic cancer.

High mitomycin C concentration in tumour tissue can be achieved by isolated liver perfusion in rats.

To enable the treatment of hepatic metastasis with higher, theoretically more effective, doses of systemically toxic anticancer drugs, an isolated liver perfusion (ILP) technique was developed in WAG/Ola rats. First, in a toxicity study the maximally tolerated dose (MTD) of mitomycin C (MMC) was determined for a 25-min ILP and for hepatic artery infusion (HAI) after the administration of a bolus dose. The MTD in the ILP setting (4.8 mg/kg) was 4 times that using HAI (1.2 mg/kg). Subsequently, in a rat colorectal hepatic-metastasis model, concentrations of MMC in tumour, liver, plasma and perfusate were measured during a 25-min ILP to investigate the expected pharmacokinetic advantage of ILP. The mean plasma level determined after ILP (1.2 as well as 4.8 mg/kg MMC) was significantly lower (P less than 0.001) than that obtained following HAI. This may explain both the absence of severe systemic toxicity and the higher MTD in ILP-treated groups. No significant difference in mean tumour and liver tissue concentrations of MMC were found when the groups treated with 1.2 mg/kg drug via HAI vs ILP were compared. The mean MMC concentration in tumour tissue was significantly higher (almost 5 times; P less than 0.05) in rats treated by ILP with the MTD (4.8 mg/kg) than in those treated via HAI with the MTD (1.2 mg/kg). ILP of MMC can be safely performed using a dose 4 times higher than the MTD in the HAI setting, leading to an almost 5-fold concentration of MMC in hepatic metastasis. ILP of MMC may therefore represent a promising therapy for metastasis confined to the liver.

[A case of hepatocellular carcinoma treated by intrahepatic arterial administration of anticancer agents: serial determination of the concentration of the chemotherapeutic agents in serum and dialysate during hemodialysis].

The concentration of adriamycin (ADR) and mitomycin C (MMC) were measured in serum and dialysate in a patient with hepatocellular carcinoma complicating chronic renal failure. ADR at a dose of 20 mg and MMC at 10 mg were administered through the hepatic artery or peripheral vein during hemodialysis. ADR and MMC became undetectable after 24 hours and 2 hours, respectively. On the other hand, while MMC was detected in dialysate, but ADR was not. Consequently, it was speculated smaller molecular weight of MMC (334.34) than that of ADR (579.99) might be responsible for the appearance of MMC in dialysate. These results suggest that hemodialysis may be effective in reducing side effects of certain anticancer agents. Serum concentration of the anticancer agents injected through hepatic artery was similar to that injected intravenously.

Extracorporeal removal of anticancer drugs in hepatic artery infusion: the effect of direct hemoperfusion combined with venovenous bypass.

The effect of a new extracorporeal system combining direct hemoperfusion (DHP) with venovenous bypass was evaluated in the elimination of anticancer drugs in hepatic artery infusion. Adriamycin (3 mg/kg) and mitomycin C (1 mg/kg) were given to mongrel dogs through the hepatic artery with three different durations of 1, 10, and 20 minutes. Plasma drug levels were determined at the inlet and outlet of DHP and right external jugular vein (systemic level). Blood flow through DHP averaged 200 ml/min. In dogs without DHP (group I; n = 4), systemic levels of adriamycin and mitomycin C increased rapidly with 1-minute infusion, reaching the peak values of 6.61 +/- 2.44 (mean +/- SD) and 2.20 +/- 1.05 micrograms/ml, respectively. With DHP under single venous bypass (group II; n = 5), the peak values were reduced to 1.25 +/- 1.02 and 0.79 +/- 0.52 microgram/ml. Moreover, the peak levels were markedly reduced by DHP under hepatic venous isolation (group III; n = 6), the values being 0.41 +/- 0.15 and 0.13 +/- 0.07 microgram/ml with 1-minute infusion. The drug-removal rates were improved substantially in group III compared with group II. The longer the duration of infusion, the higher the removal rates tended to be in group III. These results indicate that effective elimination of anticancer drugs can be accomplished by this system during intraarterial chemotherapy of the liver.

Chemotherapy in a pregnant rat model. 1. Mitomycin-C: pregnancy-specific kinetics and placental transfer.

Although cancer complicates pregnancy infrequently, its occurrence jeopardizes maternal and fetal well-being. Treatment with chemotherapeutic agents may adversely affect rapidly dividing fetal tissue, while physiologic changes in pregnancy may alter maternal drug disposition. Previous work on placental transfer and pregnancy-specific kinetics of antineoplastic agents is limited, making the establishment of treatment guidelines for the pregnant cancer patient difficult. Using a pregnant rat model and sensitive HPLC methodology we quantitated the placental transfer and resulting fetal exposure of mitomycin-C (MMC), an alkylating agent. Following maternal dosing, the relative fetal exposure was 6.4%, indicating that MMC does cross the placenta, although to a limited degree. Significant pregnancy-specific alterations in drug disposition, including higher plasma concentrations and decreased clearances in pregnant animals, highlight the need for drug-level monitoring and possible dosage modification when these agents are used in pregnancy.

[Effect of degradable starch microspheres (DSM) on hepatic hemodynamics].

The effect of intra-arterial infusion of degradable starch microspheres (DSM) on hepatic arterial blood flow was measured using a transit-time ultrasonic blood flow meter. Before the infusion of DSM, the mean hepatic arterial flows were 316 +/- 79 ml/min. After infusion, the mean flows decreased to 43 +/- 86 ml/min (86%). RI-angiography using 99mTc-macroaggregated albumin (MAA) was performed to measure hemodynamic changes in the liver. T/N ratio of 99mTc-MAA accumulation was increased from 0.37 to 0.62. Based on the peripheral MMC blood levels after combined infusion with DSM and MMC, the mean AUC with MMC plus DSM was decreased to 55% of that of AUC with MMC alone. These results show that combined use of DSM is effective for intra-arterial chemotherapy against the hepatic cancer.

[Preventive effects of intra-arterial infusion chemotherapy of liver metastasis in non-resectable pancreatic cancer].

This is a study which evaluates intra-arterial chemotherapy for non-resectable carcinoma of the pancreas without liver metastasis (locally advanced). Twenty-one patients received ia-chemotherapy (MA-CP: Methotrexate + Angiotensin II----Citrovorum factor + Prostaglandin E1). They showed a 39% 1-year survival rate and 92% of 1-year cumulative rate of survival without liver metastasis. These incidences were significantly higher than those of (13%, 50%) 52 patients who underwent systemic chemotherapy (control). As the reason for such significant differences, we suspected that a high dose of chemotherapeutic view after MTX had been infused into the pancreatic-supplying arteries. This speculation was confirmed by the serial determination of the MTX concentration in the portal blood.

Pharmacokinetic analysis in intraperitoneal hyperthermic chemotherapy of far-advanced gastric cancer patients.

The pharmacokinetics of mitomycin C (MMC) has been evaluated in 10 far-advanced gastric cancer patients by means of an intraperitoneal hyperthermic perfusion (IPHP) using MMC to prevent and/or treat peritoneal dissemination. Further, misonidazole (MIS), which was used as a thermosensitizer, also was evaluated. The IPHP was performed for 120 min right after surgical procedure, using a closed-circuit with an inflow perfusate temperature from 46.3 approximately 47.5 degrees C and an outflow temperature of 44.0 approximately 46.0 degrees C. The MMC level in the perfusate was 10 micrograms/ml at the onset of IPHP and thereafter decreased to 1.7 +/- 0.4 micrograms/ml minutes later. The average AUC (area under the curve) in the perfusate was 3.3 micrograms/ml during the IPHP. The MMC level in the peripheral blood was 0.15 +/- 0.04 micrograms/ml 30 min after start of the IPHP and increased to 0.18 +/- 0.05 micrograms/ml at the end of IPHP. Additionally, the MIS level in the peripheral blood was 64.7 +/- 10.3 micrograms/ml 60 min after the IPHP, 60.6 +/- 9.2 micrograms/ml at 120 min, and then decreased to 32.8 +/- 10.3 micrograms/ml at 12 hours after IPHP. The AUC was calculated as being 975 micrograms.hr/ml. Again, the level in the portal vein blood was calculated from the peritoneal permeability and the peripheral blood level of the drug. From data, it was concluded that the perfusate and portal vein blood level of MMC, the peripheral blood level of MIS, as well as the perfusate temperature, are important in providing a favorable, safe, antitumoral treatment.

[Blood levels of mitomycin C in patients given by various routes of administration].

Twenty mg of mitomycin C was administered to patients with colo-rectal cancer through various routes of administration. The values of pharmacokinetic parameters were as follows Intra-arterial administration (3 patients with rectal cancer): T1/2 alpha 8.4 min, T1/2 beta 63.8 min, maximum concentration (5 min after ia injection) 0.83 microgram/ml, AUC0-120 25.0 micrograms/ml/min. Intra-portal administration (7 patients with colon cancer): T1/2 alpha 6.1 min, T1/2 beta 61.0 min, maximum concentration (5 min after iv.) 0.75 microgram/ml, AUC0-120 25.3 micrograms/ml/min. Intra-peritoneal administration (14 patients with colon cancer): maximum concentration (20-30 min after i.p.) 0.23 microgram/ml, AUC0-120 21.1 micrograms/ml/min. Intra-pelvic administration (7 patients with rectal cancer): maximum concentration (20-30 min after i.p.) 0.06 micrograms/ml, AUC0-120 7.3 micrograms/ml/min. The reduction curve of intra-arterial administration closely resembles to that of intra-portal administration. Maximum blood level and AUC of intracavitary administration were relatively low compared with those of administration into blood vessels. Bone marrow suppression was seen in patients given mitomycin C into blood vessels and wound healing disturbance such as anastomotic leakage were noted in patients by intra-cavitary administration.

Clinical trial with surgery and intraperitoneal hyperthermic perfusion for peritoneal recurrence of gastrointestinal cancer.

To treat six patients with peritoneal recurrence after radical operation for gastrointestinal cancer, an intraperitoneal hyperthermic perfusion (IPHP), combined with surgical resection of recurrent tumors, intestinal by-pass anastomosis, or both, was carried out. Immediately after complete resection of the intraperitoneal recurrent tumors, a 2- to 3-hour IPHP was performed under hypothermic general anesthesia at about 32 degrees C, using a perfusate containing 10 micrograms/ml or 20 micrograms/ml of mitomycin C (MMC) warmed at the inflow temperature of 46.6 degrees C to 46.9 degrees C. The apparatus used for IPHP was designed for intraperitoneal perfusion as a closed circuit. Although five of the six patients had a malignant peritoneal effusion at the time of admission, the effusion disappeared soon after IPHP, and no cancer cell was present in the lavage from Douglas' pouch. The other patient had a recurrent tumor at the anastomotic region after low anterior resection for rectal cancer and complete resection of the recurrent tumor, combined with IPHP, was carried out. One patient with a recurrent gastric cancer died of hepatic metastasis and cancerous pleuritis 5 months after this treatment, and the other five are in good health 12.8 +/- 5.1 months after IPHP. On the other hand, five patients with intra-abdominal recurrent gastric cancer, who received only surgical treatment within the same period of time, died 3.0 +/- 2.1 months after the surgery. Postoperatively, in the six patients with IPHP, transitory hepatic dysfunction, hypoproteinemia, and thrombocytopenia occurred. These results show that IPHP using MMC combined with surgery is a safe, reliable treatment for patients with peritoneal recurrence of gastrointestinal cancer.

Phase-system switching as an on-line sample pretreatment in the bioanalysis of mitomycin C using supercritical fluid chromatography.

One of the problems of the application of supercritical fluid chromatography (SFC) in bioanalysis is the fact that many sample pretreatment procedures deliver the solutes of interest in a polar solvent, which upon injection will dramatically disturb the phase system characteristics of the SFC system. The phase-system switching approach, recently introduced for liquid chromatography-mass spectrometry, can be used to avoid this problem. Plasma samples containing the thermolabile and pH-sensitive cytostatic drug Mitomycin C (MMC) were injected onto a short precolumn. After washing and drying of the precolumn the compound of interest was desorbed using a supercritical fluid and analyzed by SFC. Up to 1 ml of plasma containing 20 ng of MMC has been analyzed in this way.

Behavior and resorption of mitomycin C following multiple intravesical administrations.

Although several pharmacological data of intravesical mitomycin C (MMC) are available now, data about resorption following subsequent intravesical instillations with different instillation times are lacking. We have analyzed MMC concentrations in blood plasma and urine following eight subsequent instillations, with 0.5- and 1.0-h instillation times. The relationships between urinary pH, urinary MMC concentrations, and MMC blood plasma concentrations were determined, as well as the stability of MMC in urine at pH 5-8 at 37 degrees C. An average of 40.3 and 46.4% of the total parent drug was recovered for the 0.5- and 1.0-h instillations, respectively. Blood plasma concentrations of MMC could be measured in nearly all patients and were independent of instillation times, urine concentration, or urinary pH. Resorption of MMC and recovery was stable during eight subsequent instillations. It was demonstrated that MMC can be degradated in urine at physiological conditions (pH less than 6; 37 degrees C). However, neither an influence of prolongation of the instillation time on MMC recovery from urine, nor a significant correlation between urinary pH and urinary MMC concentrations could be demonstrated. Since MMC can be degradated at pH less than 6 within 0.5 h, buffering of instillation fluids containing MMC is recommended. Reuse of instilled MMC, therefore, cannot be advised.

[Repeated intra-arterial chemotherapy combined with mitomycin C and degradable starch microspheres in inoperable metastatic hepatic cancer].

Intra-arterial chemotherapy combined with mitomycin C (MMC) and degradable starch microspheres (DSM) was prescribed repeatedly for 14 patients with inoperable hepatic metastasis. This intra-arterial treatment was repeated 2.7 times on the average. The average dose of DSM and MMC in a single infusion was 685 +/- 201 mg and 13.8 +/- 3.8 mg, respectively. To analyse the degradation time of DSM, MMC concentrations in the peripheral blood were measured by HPLC method. RI-angiography using 99mTc macroaggregated albumin(MAA) was performed to estimate the hemodynamic changes in the liver. Antitumor efficacy was evaluated in terms of tumor regression measured by CT scan. An objective tumor response was observed in 9/14 patients(64.2%): CR 1/14; PR 6/14; MR 2/14. Elevated CEA levels were decreased in 11/12 patients (91.7%). Based on the peripheral MMC blood levels after combined infusion with DSM and MMC, an occlusion of intrahepatic vessels with DSM continued at least for 30 minutes. Again, RI-angiography with 99mTc-MAA plus DSM revealed the increased accumulation in the tumor, compared to 99mTc-MAA only. Side effects possibly attributable to DSM were observed in 14/38 treatments, though they were slight and temporary. Thus, these results indicate that this combined intra-arterial infusion of DSM and MMC achieve, higher regional selectivity.