Objectives
The use of taxanes (paclitaxel and docetaxel) in pregnant cancer patients is increasing. We aimed to compare their transplacental transfer using the gold standard human placental perfusion model, to guide drug selection.
Study Design
Term placentas were perfused with paclitaxel or docetaxel and 2 different albumin concentrations. Main transfer parameters such as fetal transfer rate (FTR), clearance index, and placental uptake of taxanes were assessed.
Results
Twelve placentas were perfused, 6 with paclitaxel and 6 with docetaxel. Mean FTR of paclitaxel decreased significantly from 5.67 ± 0.02% in low albumin conditions to 1.72 ± 0.09% in physiological albumin conditions. Similarly, mean clearance index decreased significantly from 0.22 ± 0.02 to 0.09 ± 0.01. Regarding docetaxel, mean FTR were similar in low albumin and physiological conditions (5.03 ± 0.60% and 4.04 ± 0.22%, respectively) while mean clearance index decreased significantly from 0.18 ± 0.02 to 0.13 ± 0.01. Taxanes accumulation in cotyledon was similar for docetaxel and paclitaxel: 4.54 ± 1.84% vs 3.31 ± 1.88%, respectively.
Conclusion
Transplacental transfer and placental accumulation of paclitaxel and docetaxel were low and similar, especially in physiological conditions of albumin. Further studies are warranted to optimize the selection of a taxane in pregnant cancer patients.
Cancer is the second leading cause of mortality in young women aged 25-44 years in Western countries. About 1/1000 to 1/2000 pregnant patients are diagnosed with a malignancy, breast and gynecological cancers being the most common. This incidence is expected to increase, given the trend for women to delay childbearing.
Chemotherapy may be indicated in pregnant patients, with the need to balance both the maternal and fetal prognoses. Parameters that should be taken into account include gestational age, cancer stage and aggressiveness, patients’ wishes, and potential toxicities of active anticancer treatments.
Chemotherapy is contraindicated during organogenesis since its use increases the rates of miscarriages, intrauterine death, and malformations. However, during the second and third trimesters of pregnancy, several drugs such as doxorubicin and epirubicin may be used with limited short-term toxicity. Of note, the transplacental transfer of doxorubicin and epirubicin averages 3-4%.
Conversely, other agents such as the humanized IgG1 trastuzumab may induce significant fetal toxicity, probably consecutive to a large transplacental transfer.
Among active drugs in breast, gynecological, and lung neoplasms that may occur during pregnancy, taxanes display a favorable toxicity profile during the second and third trimesters. Indeed, paclitaxel and docetaxel are inhibitors of the depolymerization of microtubules, and target the beta-tubulin. Taxanes subsequently induce the formation of deformed tubulin structure that finally induces apoptosis. Maternal side effects (mainly alopecia, nausea and vomiting, hematological toxicity) seem to be mild and manageable, as seen in nonpregnant women. To date, fetal side effects are scarcely documented, mainly represented by intrauterine growth restriction, anhydramnios, and cytopenia. This toxicity profile seems to be nevertheless acceptable, for both drugs.
However, no data are available regarding the comparative use of paclitaxel and docetaxel in pregnant patients. Although a low transplacental transfer rate was previously suggested for paclitaxel, nothing is known about docetaxel.
Given the lack of objective data that may guide the selection of a taxane for the treatment of pregnant patients, we aimed to comparatively assess the transplacental transfer of paclitaxel and docetaxel in different albumin conditions, using the ex vivo gold standard method of human perfused cotyledon, and therefore, to determine the placental accumulation of taxanes.
Materials and Methods
Materials
Term placentas (37-41 weeks+5 days of gestation) from uneventful pregnancies were immediately collected (n = 12) after vaginal delivery or cesarean section.
The enrolled patients did not receive any medication except for epidural analgesia or oxytocin during labor, and did not present any vascular disease such as diabetes mellitus, preeclampsia, or intrauterine growth restriction.
Each patient gave informed written consent. The study was approved by the local ethics committee.
The maternal and fetal solutions were prepared with Earle medium containing 2 or 30 g/L of bovine serum albumin (Euromedex, Souffelweyersheim, France). Paclitaxel (Taxol; Bristol-Myers-Squibb, Rueil-Malmaison, France) and docetaxel (Taxotere; Sanofi-Aventis, Paris, France) were provided by the Department of Pharmacy of the Cochin Teaching Hospital in an injectable form (syringe). The targeted maternal concentrations of paclitaxel and docetaxel were around the average plasmatic maximal concentration, ie, 1530 and 4150 ng/mL for an infusion of 90 mg/m 2 of paclitaxel and for an infusion of 100 mg/m 2 of docetaxel, respectively.
Methods
Placental perfusion
Collected placentas after cesarean section or vaginal delivery were subsequently perfused in an open, double circuit, according to a validated method. Perfusion experiments were started within 45 minutes after delivery.
After a visual examination to confirm vascular integrity of both maternal and fetal sides, a distal branch of a fetal artery and its associated vein that were supplying a peripheral cotyledon were cannulated. The fetal circulation was established at a flow rate of 6 mL/min with a peristaltic pump. After confirmation of absence of vascular leakage, the perfused area progressively whitened, which allowed visualization of the selected cotyledon.
The perfusion was initiated by inserting 2 catheters into the intervillous space on the maternal side. The maternal circulation was established at a flow rate of 12 mL/min with another peristaltic pump. The pH of the maternal and fetal reservoirs was adjusted to 7.4 and 7.2, respectively.
Parameters such as perfusion pressure in the fetal circuit and potential fluid leakage from the fetal or the maternal circulation are usually monitored to check the validity of the technique.
Hence, the balance of flows in the fetal circuit was controlled all along the perfusion procedure by checking the fetal artery and the fetal venous pressures. Given the fact that fetal artery flow was known (induced by the peristaltic pump), if a difference of pressure was noticed between the fetal artery and the fetal vein, the balance of flows was subsequently found incorrect and the perfusion procedure was discontinued (presence of leakage). If the balance was correct (same fetal artery and venous pressures), the procedure was pursued. Similarly, maternal circuit was monitored using the same parameters.
Usually, the pressure values are between 40-60 mm Hg for the fetal circulation, and between 10-20 mm Hg for the maternal circulation.
A freely diffusing marker, antipyrine (Ap), was added in the maternal medium (final concentration: 20 mg/L), then, paclitaxel (or docetaxel) was subsequently added into the same compartment. The perfusion duration was 90 minutes given the fact that the aim of our study was to only investigate the kinetic of transplacental transfer of taxanes. Indeed, the goal of the perfusion was to reach the plateau of placental transfer and then to stop the perfusion. Thus, a 90-minute perfusion is usually sufficient to access this kinetic of transplacental transfer.
Samples from the maternal reservoir were subsequently collected at 0, 30, 60, and 90 minutes whereas samples from venous fetal circulation were collected every 5-90 minutes to assess paclitaxel and docetaxel concentrations.
Ap concentrations were measured by high-performance liquid chromatography using ultraviolet detection at 290 nm. The mobile phase was a mixture of 0.05 mol/L phosphate buffer (pH 3)/methanol/tetrahydrofuran (75/25/0.9, vol/vol/vol). Standard curves were prepared with Ap concentrations ranging from 50-20,000 μg/L. Both within-day and between-day accuracies and precision were <10%, and the lower quantification limit of Ap was 50 μg/L.
Two transport parameters were assessed during the study, and calculated as follows: fetal transfer rate (FTR) = (Cf/Cm) × 100, where Cf is the venous fetal concentration of drug and Cm is the maternal concentration of the same drug. The result is given as a percentage: clearance index = FTR (drug)/FTR (Ap), where FTR (drug) is the FTR of the studied drug and FTR (Ap) is the FTR of Ap.
Another parameter was assessed during our experiments: the placental uptake ratio, calculated as follows: [cotyledon taxane molecule quantity (g)/maternal taxane molecule quantity (g)] × 100.
Otherwise, we determined our success rate of placental perfusion and explained the different causes of failure, as recommended by a recent consensus.
Study of placental accumulation–quantitative analyze of taxanes
At the end of taxane perfusion, the cotyledon was perfused with a cold phosphate-buffered saline solution during 10 minutes (washing solution) and subsequently frozen in liquid nitrogen. All the cotyledons were subsequently stored at −80°C until sample processing before taxane analysis.
Once the extraction was achieved, quantitative analyze of docetaxel and paclitaxel was done using reverse-phase high-performance liquid chromatography (Agilent Technologies, Waldbronn, Germany) coupled with tandem mass spectrometry (Quattro-LCZ triple quadrupole mass spectrometer, Waters, Milford, MA). Briefly, 0.1 mL of cabazitaxel (200 ng/mL), used as internal standard, was added to aliquots (0.2 mL) of standards, controls, and samples before liquid-liquid extraction with ter-butyl-methyl-ether (1 mL). After vortex mixing and centrifugation at 13,000 rpm for 1 minute, organic supernatant (∼1 mL) was transferred and evaporated at 40°C using a SpeedVac concentrator (Thermo Scientific, Rockford, IL). Remnant was diluted in 0.1 mL of solvent analysis (acetonitril/methanol/0.05% formic acid; 30/30/40, vol/vol/vol) then drawn in a glass microvolume vial, before liquid chromatography coupled to tandem mass spectrometry analysis of 10 μL aliquot. Analytes were detected in the positive electrospray ion mode using tandem mass spectrometry with selective reaction monitoring. Mass transitions monitored were mass-to-charge ratio (m/z) 808.0→225.7 for docetaxel and m/z 854.0→285.7 for paclitaxel using collision energy of 15, 18, and 12 eV, respectively. Analytes were quantified using Masslynx software (Micromass, Manchester, United Kingdom). The method was linear in the 2.5-2500 ng/mL and 1.0-1000 ng/mL concentration range for docetaxel and paclitaxel, respectively. The lower limit of quantification was 0.8 ng/mL and 1.2 ng/mL for docetaxel and paclitaxel, respectively.
Data and statistical analysis
Descriptive statistics (mean ± SEM and median [range]) were used to describe analytical data. To compare the transplacental transfer of paclitaxel and docetaxel with 2 different albumin conditions, 1-way analysis of variance was applied.
Finally, to study the correlation between transplacental transfer and placental accumulation, a Spearman test was applied.
Results
The median age of patients at delivery was 33.6 years (range, 25−42). All deliveries occurred at term (ie, 37−41 weeks+5 days), with a median term of delivery of 39.6 weeks of gestation (range, 37.1−41.7). Median neonate heights and weights were 48.5 cm (range, 46−52) and 3380 g (range, 2620−3920), respectively. Four deliveries were performed by cesarean section, and the 8 remaining deliveries were vaginal.
Twelve placentas were successfully perfused during 90 minutes. Two placentas were cannulated but important leaks led us to abort the procedure before isolating the cotyledon. One cotyledon was successfully perfused and isolated, but during the procedure, pressure monitoring was not correct and the procedure was subsequently stopped. Finally, 3 placentas were excluded at the final checkpoint, ie, during the validation with Ap (the FTR of Ap was not correct).
Transplacental Transfer of Ap
The total mean FTR of Ap was 29.3% in all experiments. Individual mean FTRs in 2 g/L of albumin solution were 23%, 27%, and 34% for placentas perfused with paclitaxel, and 32%, 37%, and 22% for placentas perfused with docetaxel, respectively.
Individual mean FTRs in albumin physiological conditions (30 g/L of albumin solution) were 24%, 29%, and 21% for placentas perfused with paclitaxel, and 42%, 20%, and 40% for placentas perfused with docetaxel, respectively.
These results demonstrated the overlap between the maternal and the fetal circulation, and thereby led to a validation of each experiment.
Transplacental transfer of paclitaxel with 2 g/L and 30 g/L of albumin
Six placentas were perfused with paclitaxel, among them 3 placentas in 2 g/L albumin conditions and 3 placentas in 30 g/L albumin concentration. Mean maternal concentration of paclitaxel was similar as the targeted maximal concentration. All individual paclitaxel maternal concentrations were relatively stable during the experiment.
The kinetics of individual paclitaxel FTR during the perfusion procedure is represented in Figure 1 , A. Mean FTR of paclitaxel decreased from 5.67 ± 0.02% to 1.72 ± 0.09% in low and physiological albumin conditions, respectively.
The kinetics of individual paclitaxel clearance index during the perfusion procedure is represented in Figure 1 , B. Mean clearance index decreased from 0.22 ± 0.02 to 0.09 ± 0.01 in low and physiological conditions, respectively.
All the data regarding paclitaxel transplacental transfer are summarized in the Table .
| Characteristic | Paclitaxel + low albumin | Paclitaxel + physiological conditions | Docetaxel + low albumin | Docetaxel + physiological conditions |
|---|---|---|---|---|
| Molecular weight, g/mol | 854 | 807.9 | ||
| Plasmatic protein binding, % | 89-98 | >95 | ||
| Perfusion medium | Earle + albumin (2 g/L) | Earle + albumin (30 g/L) | Earle + albumin (2 g/L) | Earle + albumin (30 g/L) |
| Fetal transfer rate (%), mean ± SEM | 5.67 ± 0.02 | 1.72 ± 0.09 | 5.03 ± 0.60 | 4.04 ± 0.22 |
| Clearance index, mean ± SEM | 0.22 ± 0.02 | 0.09 ± 0.01 | 0.18 ± 0.02 | 0.13 ± 0.01 |
| Placental uptake (%), mean ± SEM | — | 3.31 ± 1.88 | — | 4.54 ± 1.84 |
| No. of perfused placentas | 3 | 3 | 3 | 3 |
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