Twenty-One Novel EGFR Kinase Domain variants in Patients with Nonsmall Cell Lung Cancer.

Somatic sequence variants in the epidermal growth factor receptor (EGFR) kinase domain are associated with sensitivity to tyrosine kinase inhibitors (TKIs) in patients with nonsmall cell lung cancer (NSCLC). Patients exhibiting sequence variants in this domain that produce kinase activity enhancement, are more likely to benefit from TKIs than patients with EGFR wild-type disease. Although most NSCLC EGFR-related alleles are concentrated in a few positions, established protocols recommend sequencing EGFR exons 18-21. In this study, 21 novel somatic variants belonging to such exons in adult Argentinean patients affected with NSCLC are reported. Of these, 18 were single amino acid substitutions (SASs), occurring alone or in combination with another genetic alteration (complex cases), one was a short deletion, one was a short deletion-short insertion combination, and one was a duplication. New variants and different combinations of previously reported variants were also found. Moreover, two of the reported SASs occurred in previously unreported positions of the EGFR kinase domain. In order to characterize the new sequence variants, physicochemical, sequence and conformational analyses were also performed. A better understanding of sequence variants in NSCLC may facilitate the most appropriate treatment choice for this complex disease.

Ocular and systemic toxicity of intravitreal topotecan in rabbits for potential treatment of retinoblastoma.

Treatment of intraocular retinoblastoma with vitreous seeding is a challenge. Different routes of chemotherapy administration have been explored in order to attaining pharmacological concentrations into the posterior chamber. Intravitreal drug injection is a promissing route for maximum bioavailability to the vitreous but it requires a well defined dose for achieving tumor control while limited toxicity to the retina. Topotecan proved to be a promising agent for retinoblastoma treatment due to its pharmacological activity and limited toxicity. High and prolonged concentrations were achieved in the rabbit vitreous after 5 µg of intravitreal topotecan. However, whether a lower dose could achieve potentially therapeutic levels remained to be determined. Thus, we here study the pharmacokinetics of topotecan after 0.5 µg and the toxicity profile of intravitreal topotecan in the rabbit eye as a potential treatment of retinoblastoma. A cohort of rabbits was used to study topotecan disposition in the vitreous after a single dose of 0.5 µg of intravitreal topotecan. In addition, an independent cohort of non-tumor bearing rabbits was employed to evaluate the clinical and retinal toxicity after four weekly injections of two different doses of intravitreal topotecan (Group A, 5 µg/dose; Group B, 0.5 µg/dose) to the right eye of each animal. The same volume (0.1 ml) of normal saline was administered to the left eye as control. A third group of rabbits (Group C) served as double control (both eyes injected with normal saline). Animals were weekly evaluated for clinical and hematologic values and ocular evaluations were performed with an inverse ophthalmoscope to establish potential topotecan toxicity. Weekly controls included topotecan quantitation in plasma of all rabbits. Electroretinograms (ERGs) were recorded before and after topotecan doses. One week after the last injection, topotecan concentrations were measured in vitreous of all eyes and samples for retinal histology were obtained. Our results indicate that topotecan shows non linear pharmacokinetics after a single intravitreal dose in the range of 0.5-5 µg in the rabbit. Vitreous concentration of lactone topotecan was close to the concentration assumed to be therapeutically active after 5 h of 0.5 µg intravitreal administration. Eyes injected with four weekly doses of topotecan (0.5 or 5 µg/dose) showed no significant differences in their ERG wave amplitudes and implicit times in comparison with control (p > 0.05). Animals showed no weight, hair loss or significant changes in hematologic values during the study period. There were no significant histologic damage of the retinas exposed to topotecan treatments. After intravitreal administration no topotecan could be detected in plasma during the follow-up period nor in the vitreous of treated and control animals after 1 week of the last injection. The present data shows that four weekly intravitreal injection of 5 µg of topotecan is safe for the rabbit eye. Despite multiple injections of 0.5 µg of topotecan are also safe to the rabbit eye, lactone topotecan vitreous concentrations were potentially active only after 5 h of the administration. We postulate promising translation to clinics for retinoblastoma treatment.

Pharmacokinetic analysis of topotecan after superselective ophthalmic artery infusion and periocular administration in a porcine model.

PURPOSE: To characterize the vitreous and plasma pharmacokinetics of topotecan after ophthalmic artery infusion (OAI) subsequent to superselective artery catheterization and to compare it with periocular injection (POI). METHODS: The ophthalmic artery of 4 pigs was catheterized and 1 mg of topotecan infused over a period of 30 minutes. The contralateral eye was subsequently used for administering topotecan by POI. Serial vitreous specimens were obtained by microdialysis and plasma samples collected and assayed for total and lactone topotecan. RESULTS: Maximum total topotecan concentration in the vitreous (median, range) was significantly higher after OAI compared with POI (131.8 ng/mL [112.9-138.7] vs. 13.6 ng/mL [5.5-15.3], respectively; P < 0.005). Median vitreous exposure calculated as area under the curve for total topotecan attained after OAI was significantly higher than after POI (299.8 ng·hour/mL [247.6-347.2] and 48.9 ng·hour/mL [11.8-63.4], respectively; P < 0.05). The vitreous to plasma exposure ratio was 29 after OAI and 3.4 after POI. Systemic exposure for total topotecan was low after both modalities of administration, with a trend to be lower after OAI compared with POI (10.6 ng·hour/mL [6.8-13.4] vs. 18.7 ng·hour/mL [6.3-21.7]; P = 0.54). CONCLUSION: Superselective OAI resulted in significantly higher vitreous concentrations and exposure and a trend toward lower systemic exposure than POI.

Developing a real-world database for oncology: a descriptive analysis of breast cancer in Argentina.

Introduction: Registries based on Real-World Data (RWD) are those obtained outside of systematised and randomised clinical trials. They allow the collection of information from a large number of patients and enable the participation of a significant number of professionals. PrecisaXperta is a web platform developed for this purpose with more than 2 years of operation, parameterised for oncology. Its design allows the construction of an epidemiological database in real time and exportable for processing. Objective: To describe the characteristics and operation of this online data recording tool, explain how it was developed and analyse the quality of the information recorded, taking as an example the data obtained for breast cancer. Materials and methods: Physicians, computer scientists and data science analysts participated in the development. Patient data, history, educational level, diagnosis, staging, molecular markers, quality of life, types of treatments, progression and response, imaging, complications, adverse events are some of the fields included. Data treatment in terms of encryption, anonymisation, protection and validation is also explained. The selected breast cancer data for description were processed with medium-level statistical programmes, since the number required to apply Big Data engines is not yet available. Results: From a total of 6,892 solid tumours, 1,892 were breast cancer and 1,654 were selected that complied with a data set minimum elaborated ad hoc. Cases from 13 provinces showed a geolocation bias according to the place of practice of the professionals in the collaborative network. The predominant lack of data was detected in molecular markers (ki67) and correlativity in some lines of treatment. Inconsistencies in dates and therapeutic schemes were also detected. Data curation made it possible to exclude them. The age of the patients was 55.3 ± 11.88 years. At the time of diagnosis, the predominance was in stage I: 36.48% and II 30.06%, with positive hormone receptors in 1,424 (89.96%) cases. The predominant treatments were hormonal (61.54%) and target directed with 30.85% for HER2(+) and 39.14% for HER2(-) accompanied in most cases (85.9%) by some period of chemotherapy. Immunotherapy was much less represented (0.36%). Data were processed, homogenised, pooled and presented and made accessible in a form suitable for application to RWD analyses. Conclusions: PrecisaXperta fulfils this purpose of systematising the information to facilitate its loading with its simple and intuitive interface. From the analysis of the data obtained in breast cancer, it is clear that some fields should be mandatory in order to improve the quality of the information. The results describing the registered breast cancers give us a surface view of the affected population and prepare us to design future studies when we have local Big Data. This type of development, with continuous improvements and online results, will allow with its dissemination, that the participating professionals have information of what happens in the real world, having available in a democratic way, the epidemiology to be able to study, publish and investigate with these data.

Pharmacokinetic analysis of topotecan after intra-vitreal injection. Implications for retinoblastoma treatment.

Topotecan is a promising drug with activity against retinoblastoma, however, attaining therapeutic concentrations in the vitreous humor is still a challenge for the treatment of vitreous seeds in retinoblastoma. Our aim was to characterize topotecan pharmacokinetics in vitreous and aqueous humor, and to assess the systemic exposure after intra-vitreal injection in rabbits as an alternative route for maximizing local drug exposure. Anesthetized rabbits were administered intra-vitreal injections of 5 microg of topotecan. Vitreous, aqueous, and blood samples were collected at pre-defined time points. A validated high-performance liquid chromatography assay was used to quantitate topotecan (lactone and carboxylate) concentrations. Topotecan pharmacokinetic parameters were determined in vitreous, aqueous and plasma using a compartmental analysis. Topotecan lactone concentrations in the vitreous of the injected eye were about 8 ng/mL 48 h after drug administration. The median maximum vitreous, aqueous and plasma total topotecan concentrations (C(max)) were 5.3, 0.68 and 0.21 microg/mL, respectively. The C(max) vitreous/aqueous of treated eyes and the C(max) vitreous/plasma were approximately 8 and 254, respectively. Total topotecan exposure (AUC) in the vitreous of the injected eye was 50 times greater than the total systemic exposure. These findings suggest that intra-vitreal administration of only 5 microg of topotecan reaches significant local levels over an extended period of time while minimizing systemic exposure in the rabbit. Intra-vitreal topotecan administration offers a promising alternative route for enhanced drug exposure in the vitreous humor with potential application for treatment of vitreal seeds in retinoblastoma while avoiding systemic toxicities.

Pharmacokinetic study of the variability of indinavir drug levels when boosted with ritonavir in HIV-infected children.

The aim of this work is to: (1) assess therapeutic drug monitoring of indinavir (IDV) during clinical routine practice in HIV-infected children, whose antiretroviral treatment includes IDV boosted with ritonavir (RTV), and (2) describe a possible relationship between IDV pharmacokinetics and MDR1 genotypes. In 21 ambulatory pediatric patients receiving IDV plus RTV, IDV plasma levels and MDR1 genotypes on exon 26 (C3435T) were determined. Nine of the 21 patients initially receiving 250 mg/m(2) IDV yielded trough levels below 0.10 microg/ml (median: 0.21, range: 0.04-1.31 microg/ml). When the dosage was increased to 400 mg/m(2) IDV plus 100 mg/m(2) RTV b.i.d., all, except 1 patient, achieved levels above 0.10 microg/ml. Pharmacokinetic analysis showed higher volume of distribution median values related to the C/C genotype in comparison with C/T or T/T genotypes for exon 26 (4.57 vs. 1.20 and 1.50 l/kg, respectively; p = 0.002). Although a higher median value of clearance was observed with the C/C genotype, the difference was not statistically significant (1.43 vs. 0.27 and 0.42 l/h, respectively; p = 0.052). These results may be explained by a reduced absorption of the drug, related with lower plasma IDV levels in patients carrying the C/C genotype in exon 26.

Lymphocyte P-glycoprotein variability in healthy individuals.

The aim of the present work was to describe the distribution of lymphocyte P-glycoprotein activity on a population of healthy individuals, taking also into account sex and age. P-glycoprotein activity in lymphocytes was measured by the Rhodamine 123 efflux assay using flow cytometry, in the presence and absence of verapamil, a P-glycoprotein inhibitor. We obtained a range of P-glycoprotein activity from 1.04 to 3.79. The distribution of the activity in the population studied was better described by a bimodal model, according with the Kolmogorov-Smirnov test. The frequency adjusted to the following equation: F = 0.70 N (2.11; 0.43) + 0.30 N(3.29; 0.26), in which 0.70 and 0.30 represented the proportion of each group, and 0.43 and 0.26 were the standard deviations of the activity of each group, respectively. The study of the relationship between subjects' age and P-glycoprotein activity showed no statistical significance. When healthy volunteers were separated according to sex, similar distributions were observed, although for men an increase in proportion of higher P-glycoprotein function group was observed. The variability observed in the population studied was important, with some volunteers with very scarce activity and some with a fourfold higher activity. Characterization of P-glycoprotein functionality in the population represents a useful contribution to the beginning of pharmacological treatments that consider its effect on pharmacokinetics and pharmacodynamics of individualized patients.

Topotecan vitreous levels after periocular or intravenous delivery in rabbits: an alternative for retinoblastoma chemotherapy.

PURPOSE: To determine the extent and the mechanism by which topotecan, a candidate agent for the treatment of retinoblastoma, gains access to the vitreous when administered by periocular injection or intravenous infusion. METHODS: In vivo experiments were conducted in which albino rabbits received 1 mg topotecan by periocular injection (POI group; n = 30) or as a 30-minute intravenous infusion (IV group; n = 16). Plasma and vitreal topotecan concentrations were analyzed during the 10 hours after administration. A population pharmacokinetic model was fit to the data. Additionally, periocular injections were performed postmortem to study the effect of removing the blood vasculature barrier. RESULTS: Potentially active lactone topotecan levels were detected in the vitreous in the POI and IV groups. Both administration schedules induced high total topotecan plasma exposures because of absorption from the periocular depot, though plasma lactone area under the curve (AUC) was significantly higher in the IV group. Similar vitreal concentrations were found in treated and control eyes in the POI group. The transfer from the periocular compartment to the vitreous was negligible. The absence of drug levels in the control eye of the postmortem-injected rabbits confirmed the systemic delivery of topotecan. Local toxicity was not observed. CONCLUSIONS: As a consequence of a favored passage across the blood-retinal barrier, considerable topotecan vitreous levels were detected in a rabbit model after systemic or periocular administration. Transscleral entry in vivo was constrained by rapid clearance from the administration site.

Nimodipine restores the altered hippocampal phenytoin pharmacokinetics in a refractory epileptic model.

The present work was undertaken to examine the central pharmacokinetics of phenytoin (PHT) in an experimental model of epilepsy, induced by administration of 3-mercaptopropionic acid (MP), and possible participation of P-glycoprotein in this model of epilepsy. Repeated seizures were induced in male Wistar rats by injection of 3-MP (45 mg kg(-1), i.p.) during 10 days. Control rats (C) were injected with saline solution. In order to monitor extracellular PHT levels, either a shunt microdialysis probe or a concentric probe was inserted into carotid artery or hippocampus, respectively. All animals were administered with PHT (30 mg kg(-1), i.v.) 30 min after intraperitoneal administration of vehicle (V) or nimodipine (NIMO, 2 mg kg(-1)). No differences were found in PHT plasma levels comparing all experimental groups. In pre-treated rats with V, hippocampal PHT concentrations were lower in MP (maximal concentration, C(max): 2.7+/-0.3 microg ml(-1), p<0.05 versus C rats) than in C animals (C(max): 5.3+/-0.9 microg ml(-1)). Control rats pre-treated with NIMO showed similar results (C(max): 4.5+/-0.8 microg ml(-1)) than those pre-treated with V. NIMO pre-treatment of MP rats showed higher PHT concentrations (C(max): 6.8+/-1.0 microg ml(-1), p<0.05) when compared with V pre-treated MP group. Our results indicate that central pharmacokinetics of PHT is altered in MP epileptic rats. The effect of NIMO on hippocampal concentrations of PHT suggests that P-glycoprotein has a role in reduced central bioavailability of PHT in our epileptic refractory model.

Pharmacokinetic-pharmacodynamic modeling of diltiazem in spontaneously hypertensive rats: a microdialysis study.

INTRODUCTION: The aim of the present work was to study the applicability of a modified E(max) pharmacodynamic model for the pharmacokinetic-pharmacodynamic (PK-PD) modeling of diltiazem in spontaneously hypertensive rats (SHR) and normotensive Wistar Kyoto (WKY) rats. METHODS: A "shunt" microdialysis probe was inserted in a carotid artery of anaesthetized SHR and WKY rats for simultaneous determination of unbound plasma concentrations of diltiazem and their effects on mean arterial pressure (MAP) and heart rate (HR) after the intravenous application of 1 and 3 mg kg(-1) of the drug. Correlation between diltiazem plasma levels and their cardiovascular effects was established by fitting the data to a conventional and modified E(max) model. RESULTS: Volume of distribution and clearance of diltiazem was greater in SHR than in WKY animals. A proportional increase of area under curve with dose increment was observed in WKY animals but not in SHR. A good correlation between plasma unbound concentrations of diltiazem and their hypotensive and chronotropic effects was found in both experimental groups using both PK-PD models. The application of the modified E(max) model for PK-PD modeling of diltiazem allowed a more accurate and precise estimation of PK-PD parameters than the E(max) equation do. Chronotropic effect of 3 mg kg(-1) diltiazem was lower in SHR compared to WKY animals. Initial sensitivity (S(0)) to diltiazem chronotropic effect was greater in SHR with regards to WKY animals after administration of 1 mg kg(-1). S(0) to diltiazem hypotensive effect was greater in SHR with regards to WKY animals after administration of both doses of diltiazem. DISCUSSION: Microdialysis sampling is a useful technique for the pharmacokinetic study and pharmacokinetic-pharmacodynamic (PK-PD) modeling of diltiazem. The modified E(max) model allows an accurate estimation of drug sensitivity in conditions when maximal pharmacological response can not be attained. Genetic hypertension induced changes in the pharmacokinetic and PK-PD behavior of diltiazem suggesting that SHR is an interesting animal model for pre-clinical evaluation of calcium channel blockers.

Increased delivery of chemotherapy to the vitreous by inhibition of the blood-retinal barrier.

Treatment of retinoblastoma -a pediatric cancer of the developing retina- might benefit from strategies to inhibit the blood-retinal barrier (BRB). The potent anticancer agent topotecan is a substrate of efflux transporters BCRP and P-gp, which are expressed at the BRB to restrict vitreous and retinal distribution of xenobiotics. In this work we have studied vitreous and retinal distribution, tumor accumulation and antitumor activity of topotecan, using pantoprazole as inhibitor of BCRP and P-gp. We used rabbit and mouse eyes as BRB models and patient-derived xenografts as retinoblastoma models. To validate the rabbit BRB model we stained BCRP and P-gp in the retinal vessels. Using intravitreous microdialysis we showed that the penetration of the rabbit vitreous by lactone topotecan increased significantly upon concomitant administration of pantoprazole (P=0.0285). Pantoprazole also increased topotecan penetration of the mouse vitreous, measured as the vitreous-to-plasma topotecan concentration ratio at the steady state (P=0.0246). Pantoprazole increased topotecan antitumor efficacy and intracellular penetration in retinoblastoma in vitro, but did not enhance intratumor drug distribution and survival in mice bearing the intraocular human tumor HSJD-RBT-2. Anatomical differences with the clinical setting likely limited our in vivo study, since xenografts were poorly vascularized masses that loaded most of the vitreous compartment. We conclude that pharmacological modulation of the BRB is feasible, enhances anticancer drug distribution into the vitreous and might have clinical implications in retinoblastoma. CHEMICAL COMPOUNDS INCLUDED IN THIS MANUSCRIPT: Topotecan (PubChem CID: 60700) Pantoprazole sodium (PubChem CID: 15008962).

Pharmacokinetic-pharmacodynamic (PK-PD) modeling of cardiovascular effects of metoprolol in spontaneously hypertensive rats: a microdialysis study.

The present work addressed possible alterations in the pharmacokinetics and the in vivo pharmacodynamic of metoprolol (MET) in spontaneously hypertensive (SH) rats and Wistar Kyoto (WKY) animals by means of the microdialysis technique. The correlation between MET unbound plasma concentrations and its pharmacological effects, such as heart rate and blood pressure change, was also examined in SH and WKY rats by the application of a PK-PD model. MET dialysate concentrations and its chronotropic and blood pressure effect were determined during 3 h after the administration of 3 and 10 mg.kg(-1) of the drug. A PK-PD model with a separate effect compartment was used to analyse the data. A good correlation between plasma MET concentrations and its hypotensive and chronotropic effect was found in all experimental groups. Although a greater maximal effect (E(max)) for the antihypertensive effect of MET was observed in SH rats (WKY: E(max): -17+/-1 mmHg; SH: E(max): -28+/-4 mmHg; P<0.05 versus WKY rats), no differences were found in the concentration yielding half-maximal response (IC(50)) comparing SH (IC(50): 583+/-146 ng x ml(-1)) and WKY animals (IC(50): 639+/-187 ng x ml(-1)). The bradycardic effect of MET was greater in SH rats (E(max): -29+/-1%, P<0.05 versus WKY rats) than in WK animals (E(max): -22+/-2%), but no differences were observed in the IC(50) comparing both experimental groups (WKY: IC(50): 187+/-53 ng x ml(-1); SH: IC(50): 216+/-62 ng x ml(-1)). Pharmacokinetic analysis shows that the volume of distribution of MET was greater in SH rats (Vd: 3.4+/-0.5 l, P<0.05 versus WKY rats) with regard to Wistar Kyoto (WKY) animals (Vd: 1.9+/-0.2 l). The results suggest that the pharmacokinetic behaviour of metoprolol are modified in SH rats, resulting in an increased volume of distribution. A greater maximal efficacy to the hypotensive effect of metoprolol was observed in SH rats, suggesting participation of beta-adrenoceptors in the maintenance of the hypertension. Also, a greater chronotropic response to metoprolol was found in the hypertensive group compared with WKY animals, suggesting that, at least in part, the greater cardiac effect of metoprolol explained the enhanced hypotensive response of the beta blocker in the SH animals.

Tetronic® 904-containing polymeric micelles overcome the overexpression of ABCG2 in the blood-brain barrier of rats and boost the penetration of the antiretroviral efavirenz into the CNS.

AIM: To assess the involvement of ABCG2 in the pharmacokinetics of efavirenz in the blood-brain barrier (BBB) and investigate a nanotechnology strategy to overcome its overexpression under a model of chronic oral administration. Materials & methods A model of chronic efavirenz (EFV) administration was established in male Sprague-Dawley rats treated with a daily oral dose over 5 days. Then, different treatments were conducted and drug concentrations in plasma and brain measured. RESULTS: Chronic treatment with oral EFV led to the overexpression of ABCG2 in the BBB that was reverted after a brief washout period. Moreover, gefitinib and the polymeric amphiphile Tetronic(®) 904 significantly inhibited the activity of the pump and potentiated the accumulation of EFV in CNS. The same effect was observed when the drug was administered within mixed micelles containing TetronicT904 as the main component. CONCLUSION: Tetronic 904-containing polymeric micelles overcame the overexpression of ABCG2 in the BBB caused by chronic administration of EFV then boosting its penetration into the CNS.

Influence of MDR1 C1236T polymorphism on lopinavir plasma concentration and virological response in HIV-1-infected children.

BACKGROUND: Variability in MDR1 and PXR has been associated with differences in drug plasma levels and response to antiretroviral therapy. We investigated whether polymorphisms in MDR1 (T-129C, C1236T and C3435T) and PXR (C63396T) affect lopinavir plasma concentration and the virological or immunological response to HAART in HIV-1-infected children. METHODS: Genotypes were identified in 100 blood donors and 38 HIV-1-infected children. All children received HAART with lopinavir boosted with ritonavir (LPV/r) at the time of LPV plasma level quantification, before (Ctrough) and between 1 and 2h after (Cpost-dose) the administration of the next dose of drug. CD4(+) T-cell counts and plasma viral load were analyzed before and after the initiation of LPV/r. RESULTS: MDR1 1236T, MDR1 3435T and PXR 63396T alleles showed a frequency of ~50% while the MDR1 -129C allele only reached 5%. Children heterozygotes 1236CT showed a significantly lower LPV Cpost-dose than homozygotes 1236TT (median Cpost-dose=3.04 µg/ml and 6.50 µg/ml, respectively; p=0.016). Children heterozygotes 1236CT also had a lower decrease of viral load after 36 weeks of LPV/r exposure compared with homozygotes 1236CC (median viral load changes=-0.50 log 10 copies/ml and -2.08 log 10 copies/ml, respectively; p=0.047). No effect on the immunological response was observed for polymorphisms of MDR1 or PXR. CONCLUSIONS: Our results suggest that the MDR1 C1236T SNP significantly reduces LPV plasma concentration affecting the virological response to HAART. Heterozygotes 1236CT might have an altered level of P-gp expression/activity in enterocytes and CD4(+) T lymphocytes that limits the absorption of LPV leading to an impaired virological suppression.

Pharmacokinetic analysis of melphalan after superselective ophthalmic artery infusion in preclinical models and retinoblastoma patients.

PURPOSE: To characterize melphalan pharmacokinetics after superselective ophthalmic artery infusion (SSOAI) in animals and children with retinoblastoma. METHODS: Vitreous and plasma samples of five Landrace pigs were obtained over a 4-hour period after SSOAI of melphalan (7 mg). Melphalan cytotoxicity was evaluated in retinoblastoma cell lines with and without topotecan. Plasma samples were obtained from 17 retinoblastoma patients after SSOAI of 3 to 6 mg of melphalan to one (n=14) or two eyes (n=3). Correlation between plasma pharmacokinetics and age, dosage, and systemic toxicity was studied in patients. RESULTS: In animals, melphalan peak vitreous levels were greater than its IC50 and resulted in 3-fold vitreous-to-plasma exposure. In patients, a large variability in pharmacokinetic parameters was observed and it was explained mainly by body weight (P<0.05). A significantly higher systemic area under the curve was obtained in children receiving more than 0.48 mg/kg for bilateral tandem infusions (P<0.05). These children had 50% probability of grades 3-4 neutropenia. Plasma concentrations after 2 and 4 hours of SSOAI were significantly higher in these children (P<0.05). A synergistic cytotoxic effect of melphalan and topotecan was evident in cell lines. CONCLUSIONS: Potentially active levels of melphalan after SSOAI were achieved in the vitreous of animals. Low systemic exposure was found in animals and children. Doses greater than 0.48 mg/kg, given for bilateral tandem infusions, were associated with significantly higher plasma levels and increased risk of neutropenia. Synergistic in vitro cytotoxicity between melphalan and topotecan favors combination treatment.

Efavirenz is a substrate and in turn modulates the expression of the efflux transporter ABCG2/BCRP in the gastrointestinal tract of the rat.

The oral bioavailability of the antiretroviral efavirenz (EFV) undergoes high inter and intra-individual variability, this fact supporting its therapeutic drug monitoring. Previously, it was demonstrated that the encapsulation of EFV within polymeric micelles increases the oral bioavailability of the drug. The breast cancer resistant protein (BCRP, ABCG2) is known to be inhibited by EFV in vitro. Since ABCG2 is profusely expressed in the gastrointestinal tract, the aim of the present work was to thoroughly investigate whether the intestinal permeability of EFV is modulated by ABCG2. The functional role of ABCG2 in mediating the transport of EFV at the intestinal level was consistent with the following findings: (a) an ABCG2 inhibitor, fumitremorgin C (5-10µM), significantly potentiated the mucosal-to-serosal permeation of the drug in everted gut sacs; (b) a five-day oral treatment with 20mg/kg EFV promotes the over-expression of ABCG2 in about 100%, this phenomenon being accompanied by a clear decline in the intestinal permeability of the antiretroviral and (c) the normalization of the ABCG2 expression within 24h after the last administration of EFV was coincident with the recovery of the ability of the drug to permeate through the small intestine wall. Interestingly, no interactions between EFV and P-glycoprotein (ABCB1) were apparent. Since the intestinal permeability of a drug could be associated with its in vivo absorbability, we suggest that the oral absorption of EFV is affected by modifications in the ABCG2 intestinal expression contributing to the intra-individual bioavailability variations.

Episcleral implants for topotecan delivery to the posterior segment of the eye.

Purpose. Intravenous or periocular topotecan has been proposed as new treatment modality for patients with advanced intraocular retinoblastoma, but systemic topotecan lactone exposure induced by both approaches may cause toxicity. The purpose of this study was to develop a topotecan-loaded ocular delivery system to minimize systemic exposure and achieve selective transscleral penetration. Methods. Biocompatible polymer implants containing low (0.3 mg) or high (2.3 mg) topotecan load were manufactured and characterized in vitro. Adrenaline (500 mug) was coloaded to induce local vasoconstriction in vivo in 2 of 4 animal groups. Implants were inserted into the episclera of rabbits, and topotecan (lactone and total) concentrations in ocular tissues and plasma were determined over a period of 48 hours. Results. In vitro, implants released 30% to 50% of the loaded drug within 48 hours and 45% to 70% by day 10. In vivo, topotecan lactone was highly accumulated in locally exposed ocular tissues (ranging from 10(5) to 10(6) ng/g in sclera and choroid and 10(2) to10(3) ng/g in retina) over 48 hours with all the formulations studied. Low vitreous topotecan lactone levels (approximately 5 ng/mL) were found in animals receiving concomitant local vasoconstriction and high load implants. Topotecan lactone concentrations in plasma and in contralateral eyes were minimal or undetectable as a marker of tissue selectivity of the proposed strategy. Conclusions. These studies may contribute to improving the efficacy and safety of chemotherapy treatments for retinoblastoma and may support the role of the local vasculature and tissues promoting drug clearance and local accumulation during transscleral drug delivery.

Relationship between P-glycoprotein activity measured in peripheral blood mononuclear cells and indinavir bioavailability in healthy volunteers.

Indinavir, a HIV-1 protease inhibitor, showed large inter-individual pharmacokinetic variability. It has been proposed as a substrate of P-glycoprotein (P-gp), an efflux transporter, that may contribute to limit indinavir bioavailability. A liquid formulation of indinavir was developed from indinavir capsules in order to study indinavir pharmacokinetics in healthy volunteers. Compartmental and noncompartmental analysis of indinavir plasma concentrations showed high inter-individual variability in terms of area under the curve (AUC) and maximal plasma concentration (C(max)). A significant negative association between AUC normalized to body weight (AUC x weight) and lymphocyte P-gp activity, using Rh123 efflux assay, was observed (p = 0.008; r = -0.75). AUC normalized to elimination rate constant (AUC x beta) also showed a significant negative relationship with lymphocyte P-gp activity (p = 0.03, r = -0.64). Apparent clearance (CL/[F x weight]) and volume of distribution (VD/[F x weight]) showed a positive correlation with P-gp activity. Conversely, elimination rate constant did not correlate with P-gp activity. Although there is not enough evidence of a correlation between lymphocitary and intestinal function of P-gp, our results suggest a relationship between a P-gp phenotype marker, Rh123 efflux assay in lymphocytes, and indinavir bioavailability.

A phase I study of periocular topotecan in children with intraocular retinoblastoma.

PURPOSE: To identify the maximum tolerated dose and dose-limiting toxicity of periocular topotecan in patients with relapsed or resistant intraocular retinoblastoma who are facing imminent enucleation. METHODS: For this phase I study, a starting dose of 0.5 mg of periocular topotecan administered through a 25-gauge needle was given with intrapatient escalation at a rate of 0.5 mg/cycle according to toxicity, up to a maximum dose of 2 mg. Two courses separated by 2 weeks were scheduled. Plasma levels of topotecan were measured by high-performance liquid chromatography in patients with available intravenous catheters. RESULTS: Seven eyes of five patients were treated with a total of 14 courses of periocular topotecan. Only mild orbital edema occurred, and grade 1 vomiting developed in the first patient that was controlled with ondansetron for the following courses. Dose-limiting toxicity was not reached and the maximum tolerated dose was set at the target dose of 2 mg (n=5 eyes). Lactone topotecan systemic exposure was lower than 55 ng/mL x h and it correlated linearly with dose in this small cohort. Even though the study was not designed to assess response, one eye was preserved after a partial response, but the remaining six were enucleated, either after a short period of disease stabilization followed by further therapy with other agents in five patients or by rapidly progressive disease in one. CONCLUSIONS: The dose limiting toxicity was not reached. Up to 2 mg of periocular topotecan could be given safely, but further studies are necessary to determine its effect on retinoblastoma (ClinicalTrials.gov number, NCT00460876).

Application of microdialysis in clinical pharmacology.

Microdialysis has been developed during the last 25 years by several authors primarily to study brain function and changes in levels of endogenous compounds such as neurotransmitters or metabolites in different laboratory animals. However, in the last ten years microdialysis sampling has been introduced as a versatile technique in the clinical setting. Although, microdialysis sampling has been extensively used for metabolic monitoring in patients, it was also employed for the study of distribution of different therapeutic agents especially anti-infective and antineoplasic drugs. In addition, clinical effect of drugs in patients could be also determined by means of microdialysis. So, this article reviewed the vast applications of the microdialysis technique for the study of pharmacokinetic and pharmacodynamic properties of drugs in the clinical setting.