Combined effect of cyclosporine and sirolimus on improving the longevity of recombinant adenovirus-mediated transgene expression in the retina

Objectives: To reevaluate the longevity and intraocular safety of recombinant adenovirus (rAd)-mediated gene delivery after subretinal injection, and to prolong transgene expression through the combination of 2 synergistic immunosuppressants. Methods: An rAd vector carrying green fluorescent protein (GFP) gene was delivered subretinally in the rat eye. The GFP expression was monitored in real time by fundus fluorescent photography. Intraocular safety was examined by observation of changes of retinal pigmentation, cell infiltration in virus-contacted area, immunophenotyping for CD4(+) and CD8(+) cytotoxic T lymphocytes, and CD68(+) macrophages, histologic findings, and dark-adapted electroretinography. Two synergistic immunosuppressants, cyclosporine and sirolimus, were used alone or in combination to prolong transgene expression by temporary immunosuppression. Results: The GFP expression peaked on day 4, dramatically decreased on day 10, and was not detectable on day 14. The decreased GFP expression was coincident with cell infiltration in virus-contacted area. Immunostaining showed that the infiltrating cells were CD4(+) and CD8(+) cytotoxic T lymphocytes and CD68(+) macrophages. Clumped retinal pigmentation and decreased b wave of dark-adapted electroretinogram were observed at 3 to 4 weeks after injection. Histologic examination confirmed rAd-induced retinal degeneration. Transient immunosuppression by cyclosporine and sirolimus, either alone or in combination, improved transgene expression, with the combination being the most efficient. The combined immunosuppression attenuated but did not retard the rAd-induced retinal damage. Conclusions: Transgene expression mediated by rAd after subretinal delivery is short-term and toxic to the retina. Combination of cyclosporine and sirolimus may act as an immunosuppressive adjunct to prolong rAd-mediated gene transfer. Clinical Relevance: The intraocular safety of rAd should be carefully considered before clinical trials are performed.

CEDIA (R) sirolimus assay compared with HPLC-MS/MS and HPLC-UV in transplant recipient specimens

The role of the therapeutic drug monitoring laboratory in support of immunosuppressant drug therapy is well established, and the introduction of sirolimus (SRL) is a new direction in this field. The lack of an immunoassay for several years has restricted the availability of SRL assay services. The recent availability of a CEDIA (R) SRL assay has the potential to improve this situation. The present communication has compared the CEDIA (R) SRL method with 2 established chromatographic methods, HPLC-UV and HPLC-MS/MS. The CEDIA (R) method, run on a Hitachi 917 analyzer, showed acceptable validation criteria with within-assay precision of 9.1% and 3.3%, and bias of 17.1% and 5.8%, at SRL concentrations of 5.0 mu g/L and 20 mu g/L, respectively. The corresponding between-run precision values were 11.5% and 3.3% and bias of 7.1% and 2.9% at 5.0 mu g/L and 20 mu g/L, respectively, The lower limit of quantification was found to be 3.0 mu g/L. A series of 96 EDTA whole-blood samples predominantly from renal transplant recipients were assayed by the 3 methods for comparison. It was found that the CEDIA (R) method showed a Deming regression line of CEDIA = 1.20 X HPLC-MS/MS - 0.07 (r = 0.934, SEE = 1.47), with a mean bias of 20.4%. Serial blood samples from 8 patients included in this evaluation showed that the CEDIA (R) method reflected the clinical fluctuations in the chromatographic methods, albeit with the variable bias noted. The CEDIA (R) method on the H917 analyzer is therefore a useful adjunct to SRL dosage individualization in renal transplant recipients.

Relationships between sirolimus dosing, concentration and outcomes in renal transplant recipients

Aim To explore relationships between sirolimus dosing, concentration and clinical outcomes. Methods Data were collected from 25 kidney transplant recipients (14 M/11 F), median 278 days after transplantation. Outcomes of interest were white blood cell (WBC) count, platelet (PLT) count, and haematocrit (HCT). A naive pooled data analysis was performed with outcomes dichotomized (Mann-Whitney U-tests). Results Several patients experienced at least one episode when WBC (n = 9), PLT (n = 12), or HCT (n = 21) fell below the lower limits of the normal range. WBC and HCT were significantly lower (P < 0.05) when sirolimus dose was greater than 10 mg day(-1), and sirolimus concentration greater than 12 mu g l(-1). No relationship was shown for PLT and dichotomized sirolimus dose or concentration. Conclusions Given this relationship between sirolimus concentration and effect, linked population pharmacokinetic-pharmacodynamic modelling using data from more renal transplant recipients should now be used to quantify the time course of these relationships to optimize dosing and minimize risk of these adverse outcomes.

Generalized, pruritic, ulcerating maculopapular rash necessitating cessation of sirolimus in a liver transplantation patient

The use of sirolimus as an alternative to calcineurin antagonists has enabled the continuation of immunosuppression in patients with renal impairment with preservation of kidney function. Sirolimus is generally well tolerated, with the main causes of cessation of therapy related to its effect on blood lipid profile as well as leukopenia and thrombocytopenia. We report a case of a debilitating ulcerating maculopapular rash necessitating cessation of the drug in a liver transplantation patient. A 56-year-old Caucasian liver transplantation patient presented with a diffuse, debilitating rash attributed to sirolimus use. This ultimately necessitated cessation of the immunosuppressant with subsequent resolution of her symptoms. From a review of the current literature, this is a highly unusual adverse reaction to sirolimus.

Comparison of the reintroduced MEIA (R) assay with HPLC-MS/MS for the determination of whole-blood sirolimus from transplant recipients

Therapeutic monitoring with dosage individualization of sirolimus drug therapy is standard clinical practice for organ transplant recipients. For several years sirolimus monitoring has been restricted as a result of lack of an immunoassay. The recent reintroduction of the microparticle enzyme immunoassay (MEIA (R)) for sirolimus on the IMx (R) analyser has the potential to address this situation. This Study, using patient samples, has compared the MEIA (R) sirolimus method with an established HPLC-tandem mass spectrometry method (HPLC-MS/MS). An established HPLC-UV assay was used for independent cross-validation. For quality control materials (5, 11, 22 mu g/L), the MEIA (R) showed acceptable validation criteria based on intra-and inter-run precision (CV) and accuracy (bias) of < 8% and < 13%, respectively. The lower limit of quantitation was found to be approximately 3 mu g/L. The performance of the immunoassay was compared with HPLC-MS/MS using EDTA whole-blood samples obtained from various types of organ transplant recipients (n = 116). The resultant Deming regression line was: MEIA = 1.3 x HPLC-MS/MS+ 1.3 (r = 0.967, s(y/x) = 1) with a mean bias of 49.2% +/- 23.1 % (range, -2.4% to 128%; P < 0.001). The reason for the large and variable bias was not explored in this study, but the sirolimus-metabolite cross-reactivity with the MEIA (R) antibody could be a substantive contributing factor. Whereas the MEIA (R) sirolimus method may be an adjunct to sirolimus dosage individualization in transplant recipients, users must consider the implications of the substantial and variable bias when interpreting results. In selected patients where difficult clinical issues arise, reference to a specific chromatographic method may be required.

Bayesian population pharmacokinetic analysis of sirolimus

Objective: It is usual that data collected from routine clinical care is sparse and unable to support the more complex pharmacokinetic (PK) models that may have been reported in previous rich data studies. Informative priors may be a pre-requisite for model development. The aim of this study was to estimate the population PK parameters of sirolimus using a fully Bayesian approach with informative priors. Methods: Informative priors including prior mean and precision of the prior mean were elicited from previous published studies using a meta-analytic technique. Precision of between-subject variability was determined by simulations from a Wishart distribution using MATLAB (version 6.5). Concentration-time data of sirolimus retrospectively collected from kidney transplant patients were analysed using WinBUGS (version 1.3). The candidate models were either one- or two-compartment with first order absorption and first order elimination. Model discrimination was based on computation of the posterior odds supporting the model. Results: A total of 315 concentration-time points were obtained from 25 patients. Most data were clustered at trough concentrations with range of 1.6 to 77 hours post-dose. Using informative priors, either a one- or two-compartment model could be used to describe the data. When a one-compartment model was applied, information was gained from the data for the value of apparent clearance (CL/F = 18.5 L/h), and apparent volume of distribution (V/F = 1406 L) but no information was gained about the absorption rate constant (ka). When a two-compartment model was fitted to the data, the data were informative about CL/F, apparent inter-compartmental clearance, and apparent volume of distribution of the peripheral compartment (13.2 L/h, 20.8 L/h, and 579 L, respectively). The posterior distribution of the volume distribution of central compartment and ka were the same as priors. The posterior odds for the two-compartment model was 8.1, indicating the data supported the two-compartment model. Conclusion: The use of informative priors supported the choice of a more complex and informative model that would otherwise have not been supported by the sparse data.

Chemoradiation therapy is effective for the palliative treatment of malignant dysphagia

Between 1993 and 2001, 106 patients with esophageal cancer were reviewed at a multidisciplinary clinic and treated with palliative intent by chemoradiation therapy. This study assesses the palliative benefit on dysphagia and documents the toxicity of this treatment. The study population comprised 72 men and 34 women with a median age of 69 years. Patients were treated with a median radiation dose of 35 Gy in 15 fractions with a concurrent single course of 5 FU-based chemotherapy. Dysphagia was measured at the beginning and completion of treatment and at monthly intervals until death, using a modified DeMeester (4-point) score. Treatment was well tolerated, with only 5% of patients failing to complete therapy. The treatment-related mortality was 6%. The median survival for the study population was 7 months. The median baseline score at presentation was 2 (difficulty with soft food). Following treatment, 49% of patients were assessed as having a dysphagia score of 0 (no dysphagia). Seventy-eight per cent had an improvement of at least one grade in their dysphagia score after treatment. Only 14% of patients showed no improvement with treatment. Fifty-one per cent maintained improved swallowing until the time of last follow-up or death. This single-institution study shows that chemoradiation therapy administered for the palliation of malignant dysphagia is well tolerated and produces a sustainable normalization in swallowing for almost half of all patients.