Oxaliplatin neurotoxicity--no general ion channel surface-charge effect.

BACKGROUND: Oxaliplatin is a platinum-based chemotherapeutic drug. Neurotoxicity is the dose-limiting side effect. Previous investigations have reported that acute neurotoxicity could be mediated via voltage-gated ion channels. A possible mechanism for some of the effects is a modification of surface charges around the ion channel, either because of chelation of extracellular Ca2+, or because of binding of a charged biotransformation product of oxaliplatin to the channel. To elucidate the molecular mechanism, we investigated the effects of oxaliplatin and its chloride complex [Pt(dach)oxCl](-) on the voltage-gated Shaker K channel expressed in Xenopus oocytes. The recordings were made with the two-electrode and the cut-open oocyte voltage clamp techniques. CONCLUSION: To our surprise, we did not see any effects on the current amplitudes, on the current time courses, or on the voltage dependence of the Shaker wild-type channel. Oxaliplatin is expected to bind to cysteines. Therefore, we explored if there could be a specific effect on single (E418C) and double-cysteine (R362C/F416C) mutated Shaker channels previously shown to be sensitive to cysteine-specific reagents. Neither of these channels were affected by oxaliplatin. The clear lack of effect on the Shaker K channel suggests that oxaliplatin or its monochloro complex has no general surface-charge effect on the channels, as has been suggested before, but rather a specific effect to the channels previously shown to be affected.

Oxaliplatin degradation in the presence of important biological sulphur-containing compounds and plasma ultrafiltrate.

Oxaliplatin undergoes extensive non-enzymatic chemical transformation in the body. Complexes with sulphur-containing compounds have previously been found in plasma from patients treated with oxaliplatin. We have studied the kinetics for the reactions between oxaliplatin and cysteine, methionine, and glutathione, by determination of the degradation of oxaliplatin using liquid chromatography with UV-detection. We also studied the degradation of oxaliplatin in plasma ultrafiltrate (PUF). For the degradation of oxaliplatin in the presence of glutathione, methionine, and cysteine, the second-order rate constants were 4.7M(-1)min(-1) (95% confidence interval [C.I.], 4.4-5.0M(-1)min(-1)), 5.5M(-1)min(-1) (95% C.I., 5.2-5.7M(-1)min(-1)), and 15M(-1)min(-1) (95% C.I., 14-17M(-1)min(-1)), respectively. The reaction rate was much faster than previously reported kinetics for cisplatin. The degradation rate of oxaliplatin in PUF was biphasic. The rate constant for the first phase varied from 9.5x10(-3) to 0.13min(-1) and for the second phase from (1.7 to 1.8)x10(-3)min(-1) in PUF from five healthy volunteers. The first hours of the degradation of oxaliplatin in PUF are accounted for by the degradation of oxaliplatin in a cocktail of sodium chloride and sulphur-containing compounds at physiological plasma concentrations. In conclusion, the rate of the reaction of oxaliplatin with three sulphur-containing compounds was faster for oxaliplatin than what is previously known for cisplatin. This may be important with respect to differences in the cellular effects of cisplatin and oxaliplatin treatment.

Inhibition of thioredoxin reductase but not of glutathione reductase by the major classes of alkylating and platinum-containing anticancer compounds.

Mammalian thioredoxin reductase (TrxR) is important for cell proliferation, antioxidant defense, and redox signaling. Together with glutathione reductase (GR) it is the main enzyme providing reducing equivalents to many cellular processes. GR and TrxR are flavoproteins of the same enzyme family, but only the latter is a selenoprotein. With the active site containing selenocysteine, TrxR may catalyze reduction of a wide range of substrates, but can at the same time easily be targeted by electrophilic compounds due to the extraordinarily high reactivity of a selenolate moiety. Here we addressed the inhibition of the enzyme by major anticancer alkylating agents and platinum-containing compounds and we compared it to that of GR. We confirmed prior studies suggesting that the nitrosourea carmustine can inhibit both GR and TrxR. We next found, however, that nitrogen mustards (chlorambucil and melphalan) and alkyl sulfonates (busulfan) efficiently inhibited TrxR while these compounds, surprisingly, did not inhibit GR. Inhibitions were concentration and time dependent and apparently irreversible. Anticancer anthracyclines (daunorubicin and doxorubicin) were, in contrast to the alkylating agents, not inhibitors but poor substrates of TrxR. We also found that TrxR, but not GR, was efficiently inhibited by both cisplatin, its monohydrated complex, and oxaliplatin. Carboplatin, in contrast, could not inhibit any of the two enzymes. These findings lead us to conclude that representative compounds of the major classes of clinically used anticancer alkylating agents and most platinum compounds may easily target TrxR, but not GR. The TrxR inhibition should thereby be considered as a factor that may contribute to the cytotoxicity seen upon clinical use of these drugs.

Synthesis and cytotoxicity of the dihydrated complex of oxaliplatin.

A new way of synthesizing the dihydrated oxaliplatin complex (DOC) is presented and its cytotoxicity is compared to that of oxaliplatin and cisplatin. By hydrolyzing oxaliplatin in aqueous sodium hydroxide at 70 degrees C, DOC was formed in less than 1 h. Cytotoxicity was studied in the non-small cell lung cancer cell line A549 using the fluorescent microculture cytotoxic assay. Oxaliplatin and cisplatin had similar cytotoxicity profiles, whereas DOC was found to be considerably more toxic. The cytotoxicity of oxaliplatin might, at least in part, be mediated through the formation of DOC.

Hydrolysis of oxaliplatin-evaluation of the acid dissociation constant for the oxalato monodentate complex.

Alkaline hydrolysis of the platinum anticancer drug oxaliplatin gives the oxalato monodentate complex and the dihydrated oxaliplatin complex in two consecutive steps. The acid dissociation constant for the oxalato monodentate intermediate was determined by a kinetic approach. The pK(a) value was estimated as 7.23. The monodentate intermediate is assumed to rapidly react with endogenous compounds, resulting in a continuous conversion of oxaliplatin via the monodentate form.

Alkaline hydrolysis of oxaliplatin--isolation and identification of the oxalato monodentate intermediate.

The alkaline degradation of the chemotherapeutic agent oxaliplatin has been studied using liquid chromatography. The oxalato ligand is lost in two consecutive steps. First, the oxalato ring is opened, forming an oxalato monodentate intermediate, as identified by electrospray ionization mass spectrometry. Subsequently, the oxalato ligand is lost and the dihydrated oxaliplatin complex is formed. The observed rate constants for the first step (k(1)) and the second step (k(2)) follow the equation k(1) or k(2) = k(0) + k(OH(-) )[OH(-)], where k(0) is the rate constant for the degradation catalyzed by water and k(OH(-) ) represents the second-order rate constant for the degradation catalyzed by the hydroxide ion. At 37 degrees C the rate constants for the first step are k(OH(-) ) = 5.5 x 10(-2) min(-1) M(-1) [95% confidence interval (CI), 2.7 x 10(-2) to 8.4 x 10(-2) min(-1) M(-1)] and k(0) = 4.3 x 10(-2) min(-1) (95% CI, 4.0 x 10(-2) to 4.7 x 10(-2) min(-1)). For the second step the rate constants are k(OH(-) ) = 1.1 x 10(-3) min(-1) M(-1) (95% CI, -1.1 x 10(-3) to 3.3 x 10(-3)) min(-1) M(-1) and k(0) = 7.5 x 10(-3) min(-1) (95% CI, 7.2 x 10(-3) to 7.8 x 10(-3) min(-1)). Thus, the ring-opening step is nearly six times faster than the step involving the loss of the oxalato ligand.

New insights into the biotransformation and pharmacokinetics of oxaliplatin.

Oxaliplatin is used primarily in the treatment of metastatic colorectal cancer. In this minireview, we discuss potentially important biotransformation pathways in light of its short elimination half-life in vivo. We also highlight new information achieved using a selective analytical technique to measure intact oxaliplatin in pharmacokinetic studies (comprising intravenous, intraperitoneal, and intrahepatic administration) and compare to results obtained by measurements of total platinum. The use of selective analytical techniques is strongly recommended giving kinetic parameters of the parent compound and not only to a complex mixture of platinum containing endogenous compounds.

Cisplatin and oxaliplatin toxicity: importance of cochlear kinetics as a determinant for ototoxicity.

BACKGROUND: Cisplatin is a cornerstone anticancer drug with pronounced ototoxicity, whereas oxaliplatin, a platinum derivative with a different clinical profile, is rarely ototoxic. This difference has not been explained. METHODS: In HCT-116 cells, cisplatin (20 microM)-induced apoptosis was reduced by a calcium chelator from 9.9-fold induction (95% confidence interval [CI] = 8.1- to 11.7-fold), to 3.1-fold induction (95% CI = 2.0- to 4.2-fold) and by superoxide scavenging from 9.3-fold (95% CI = 8.8- to 9.8-fold), to 5.1-fold (95% CI = 4.4- to 5.8-fold). A guinea pig model (n = 23) was used to examine pharmacokinetics. Drug concentrations were determined by liquid chromatography with post-column derivatization. The total platinum concentration in cochlear tissue was determined by inductively coupled plasma mass spectrometry. Drug pharmacokinetics was assessed by determining the area under the concentration-time curve (AUC). Statistical tests were two-sided. RESULTS: In HCT-116 cells, cisplatin (20 microM)-induced apoptosis was reduced by a calcium chelator from 9.9-fold induction (95% confidence interval [CI] = 8.1- to 11.7-fold to 3.1-fold induction) (95% CI = 2.0- to 4.2-fold) and by superoxide scavenging (from 9.3-fold, 95% CI = 8.8- to 9.8-fold, to 5.1-fold, 95% CI = 4.4- to 5.8-fold). Oxaliplatin (20 microM)-induced apoptosis was unaffected by calcium chelation (from 7.1- to 6.2-fold induction) and by superoxide scavenging (from 5.9- to 5.6-fold induction). In guinea pig cochlea, total platinum concentration (0.12 vs 0.63 microg/kg, respectively, P = .008) and perilymphatic drug concentrations (238 vs 515 microM x minute, respectively, P < .001) were lower after intravenous oxaliplatin treatment (16.6 mg/kg) than after equimolar cisplatin treatment (12.5 mg/kg). However, after a non-ototoxic cisplatin dose (5 mg/kg) or the same oxaliplatin dose (16.6 mg/kg), the AUC for perilymphatic concentrations was similar, indicating that the two drugs have different cochlear pharmacokinetics. CONCLUSION: Cisplatin- but not oxaliplatin-induced apoptosis involved superoxide-related pathways. Lower cochlear uptake of oxaliplatin than cisplatin appears to be a major explanation for its lower ototoxicity.

Oxaliplatin degradation in the presence of chloride: identification and cytotoxicity of the monochloro monooxalato complex.

PURPOSE: To study the degradation of oxaliplatin in chloride media and evaluate the cytotoxicity of oxaliplatin in normal and chloride-deficient medium. METHODS: The products of the reaction of oxaliplatin with chloride were separated on a Hypercarb S column with a mobile phase containing 40% methanol in 0.05 M ammonia and subjected to electrospray ionization mass spectrometry. The cytotoxicity of oxaliplatin in normal and chloride-deficient medium was evaluated by 30-min incubations on human colon adenocarcinoma cells (HT-29). RESULTS: We identified a new intermediate degradation product, the monochloro monooxalato complex ([Pt(dach)oxCl]-) and the final product. the dichloro complex (Pt(dach)Cl2), by liquid chromatography-mass spectrometry. [Pt(dach)oxCl]- was found as the negative ion, M-, at m/z 431, and the positive ion, [M+2H]+, m/z 433. Pt(dach)Cl2 was found as the negative ion, [M-H]-, m/z 377, and the positive ion, [M+NH4]+, m/z 396. The fast initial degradation of oxaliplatin can be coupled to the fast formation of [Pt(dach)oxCl]-. In the cytotoxic assay, the cell survival was not affected by the chloride levels. CONCLUSIONS: [Pt(dach)oxCl]-, a new transformation product of oxaliplatin, has been identified. Its in vitro cytotoxic effect does not appear to exceed that of oxaliplatin.