RT Journal Article
SR Electronic
T1 Short-term oxaliplatin exposure according to established hyperthermic intraperitoneal chemotherapy (HIPEC) protocols lacks effectiveness <em>in vitro</em> and <em>ex vivo</em>
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 709055
DO 10.1101/709055
A1 Markus W. Löffler
A1 Nick Seyfried
A1 Markus Burkard
A1 Benedikt Oswald
A1 Alexander Tolios
A1 Can Yurttas
A1 Franziska Herster
A1 Joseph Kauer
A1 Tarkan Jäger
A1 Karolin Thiel
A1 Sebastian P. Haen
A1 Hans-Georg Rammensee
A1 Sascha Venturelli
A1 Matthias Schwab
A1 Alfred Königsrainer
A1 Stefan Beckert
YR 2019
UL http://biorxiv.org/content/early/2019/07/31/709055.abstract
AB Cytotoxicity of oxaliplatin-containing solutions (OCS), sampled during patient treatment with hyperthermic intraperitoneal chemotherapy (HIPEC), was assessed by well-established continuous impedance-based real-time cell analysis (RTCA) ex vivo. HIPEC treatment was replicated by exposing OAW-42 cancer cells to OCS for 30 or 60 minutes at 42 °C. In contrast to previous observations with continuous exposure, where cytotoxicity was proven, identical OCS obtained during HIPEC did not induce cell death reproducibly and showed strongly attenuated effects after only 30 minutes of application. Based on these unexpected findings, spike-ins of oxaliplatin (OX) into peritoneal dialysis solution (PDS) or dextrose 5 % in water (D5W) were used to replicate HIPEC conditions, as used in either our own protocols or the recently presented randomized controlled PRODIGE 7 trial, where OX HIPEC for 30 minutes failed to produce survival benefits in colorectal carcinoma patients. With OX-spiked into D5W or PDS at identical concentrations as used for PRODIGE 7 or conforming with own HIPEC protocols, we did not observe the expectable cytotoxic effects in RTCA, after replicating OX HIPEC for 30 minutes. These results were corroborated for both solvents at relevant drug concentrations by classical end-point assays for cytotoxicity in two cancer cell lines. Further results suggest that penetration depth, drug dosage, exposure time and drug solvents may constitute critical factors for HIPEC effectiveness. Accordingly, we witnessed substantial cell shrinkage with both PDS and D5W, potentially contributing to reduced drug effects. Based on these results, intensified pharmacological research seems warranted to establish effective HIPEC protocols.Key PointsOxaliplatin (OX)-containing solutions obtained during patient treatment with Hyperthermic intraperitoneal chemotherapy (HIPEC) unexpectedly showed low cytotoxicity in an impedance-based ex vivo cytotoxicity cell assay.OX cytotoxicity under HIPEC conditions could be enhanced by extending drug exposure to one hour by an impedance-based ex vivo cytotoxicity cell assay.HIPEC failed to show survival benefits in the randomized controlled PRODIGE 7 trial and was questioned in the aftermath.Clinically relevant OX concentrations applied in conjunction with hyperthermia (42 °C) for 30 minutes, as used either at our own medical center or according to the PRODIGE 7 trial, proved predominantly ineffective, when used according to HIPEC routines in an impedance-based in vitro cytotoxicity cell assay.Respective findings were corroborated in two different cell lines and by two established end-point assays, showing that 50 % cell death could not be reached by the same HIPEC treatment with OX, in contrast to continuous drug exposure.As potentially relevant factor, the thickness of the exposed cell layer was identified, requiring at least ~100 µm penetration depth for our model to indicate effectiveness.Additionally, we show relevant cell shrinkage by two drug diluents used either at our own medical center or according to the PRODIGE 7 trial, potentially associated with fluid shifts out of the cell and impaired drug effects.Our own as well as recent findings by Ubink et al. (Br J Surg. 2019. doi: 10.1002/bjs.11206) support the notion that lacking effectiveness of OX HIPEC may explain the negative PRODIGE 7 trial results.CRCColorectal cancerCRSCytoreductive surgeryCTBCellTiter-Blue® assayCtrl.ControlD5WDextrose 5 % in water solutionFSC-AForward scatter areaHIPECHyperthermic intraperitoneal chemotherapyi.v.IntravenousLC5050 % cell viability thresholdMEMMedium for cell cultureMMCMitomycin CmMMillimolarµMMicromolarnCINormalized cell indexOCSOxaliplatin-containing solutions (obtained during HIPEC treatment)OSOverall survivalOvCaOvarian cancerOXOxaliplatinPBSPhosphate-buffered salinePCRPolymerase chain reactionPDSPeritoneal dialysis solutionPMPeritoneal metastasesRCTRandomized controlled trialRLRinger’s lactateRTCAReal time cell analysisSRBSulforhodamine BSDStandard deviationTCATichloroacetic acid