Rituximab Pharmacokinetics Research Articles

Differential impact of sex in young and elderly patients with DLBC: Correlation with rituximab (R) pharmacokinetics.

8570 Background: Sex and weight independently influence R clearance in elderly DLBCL pts (Mueller et al., Blood 2012). Methods: We analyzed the impact of sex on R pharmacokinetics and outcome of 1,222 elderly pts of the RICOVER-60, 823 young (18 to 60 years) aaIPI=0,1 pts of the MInT, and 375 aaIPI=2,3 pts of the Mega-CHOEP trials. R pharmacokinetics was determined by ELISA in 33 young and 49 elderly patients. Population pharmacokinetic modeling was performed with nonlinear mixed-effect modeling software (NONMEM VI). Results: R clearance was independent of tumor mass (IPI), but weakly correlated (0.2, R2linear=0.045) with increasing age in male, and moderately inversely correlated (-0.5, R2linear=0.207) with age in female DLBCL patients, resulting in similar R clearances in young female and male patients (9.88 vs. 10.38 ml/h; p=0.238), but a significantly faster R clearance in elderly males compared to females (10.50 vs 8.25 ml/h; p=0.006). In the RICOVER-60 trial, elderly females had a higher 3-year PFS (68% vs. 61%) and OS (74% vs. 68%) than male pts. due to a greater outcome improvement by the addition of R in females. In a multivariable analysis adjusting for IPI, the hazard for progression in male compared to female pts. was not significantly increased after CHOP (HR=1.1; p=0.348), but was significantly higher after R-CHOP (OR=1.6; p=0.004). In contrast, young males treated in the MInT and Mega-CHOEP trials benefitted as much as females from the addition of rituximab, with a similar hazard for male pts. after CHOP and R-CHOP (HR=1.2) with no significant difference to female patients (HRPFS=1.2, p=0.552; HROS=1.0; p=0.898). Conclusions: While no differences in R clearance and benefit from rituximab were found in young female compared to male patients, the reduced benefit of adding R to CHOP in elderly male DLBCL pts. who have a shorter rituximab serum half life and hence lower serum levels suggests that this subpopulation is suboptimally dosed when R is given based on body surface area at 375 mg/m2. Ongoing studies of the DSHNHL investigate whether higher R doses for pts. with a shorter R serum half life can improve the outcome of the respective patients. Supported by Deutsche Krebshilfe and Roche.

Pharmacokinetic Exploration of Potential for Bendamustine to Affect Rituximab Concentrations and of Cytochrome P450 1A2 Inhibitors to Affect Bendamustine Concentrations in Patients with Non-Hodgkin and Mantle Cell Lymphoma

AbstractAbstract 4919 BackgroundBendamustine, a unique alkylating agent that leads to cancer cell death via several pathways, and rituximab, an anti-CD20 monoclonal antibody, are effective as combination therapy for non-Hodgkin's lymphoma (NHL) and mantle cell lymphoma (MCL). Because bendamustine and rituximab are molecularly unique, drug-drug interactions may be unlikely. Bendamustine is primarily metabolized via hydrolysis, also reducing the likelihood of drug-drug interactions. A recently reported analysis confirmed that rituximab does not affect systemic exposure to bendamustine, but the potential impact of bendamustine on rituximab has not been reported. Moreover, because bendamustine is a CYP1A2 substrate, the potential may exist for drug-drug interactions with CYP1A2 inhibitors and inducers. This analysis graphically explored the potential for drug-drug interactions by examining (1) the effect of bendamustine on rituximab pharmacokinetics (PK) and (2) bendamustine metabolism in the presence of cytochrome P450 (CYP) 1A2 inhibitors. MethodsData from a phase 3 study of combination bendamustine-rituximab in patients with advanced indolent NHL or MCL were assessed to evaluate the potential of bendamustine to affect systemic rituximab exposure. Rituximab 375 mg/m2 was administered on day 1 followed by bendamustine 90 mg/m2 30-min infusion on days 1 and 2 of 6 to 8 consecutive 28-day cycles. Serum rituximab concentrations at end of rituximab infusion (ie, rituximab monotherapy), 24 hours and 7 days postinfusion (ie, after bendamustine exposure) were measured. Data were compared with published literature on PK of rituximab administered without bendamustine (data on methods used in the published reports were limited). To evaluate effect of CYP1A2 inhibitors on bendamustine PK, graphical displays of observed bendamustine concentrations versus time since previous dose stratified by presence or absence of CYP1A2 inhibitors were generated. ResultsThe full analysis population included 49 patients (median age, 63. 1 years; range 37–84). To analyze the effect of bendamustine on rituximab PK, 77 rituximab PK sample records from a subset of 19 patients were available. Minimum, median, and maximum observed serum rituximab concentrations in the absence (at end of rituximab infusion) and presence (at 24 hours and 7 days after rituximab infusion) of bendamustine are shown in Figure 1. The slope of rituximab concentrations from end of infusion to 24 hours and 7 days was similar to that seen in rituximab studies without bendamustine. In this study, rituximab concentrations with and without bendamustine were consistently lower than those in the literature (24% at end of infusion, 30% at 24 hours, and 53% at 7 days).To analyze the effect of CYP1A2 inhibitors on bendamustine PK, 141 samples from 29/49 (59%) patients were taken in the presence of CYP1A2 inhibitors, and 102 samples from the remaining 20/49 patients were CYP1A2 inhibitor-free. Bendamustine concentrations were similar in observed time plots stratified by presence or absence of CYP1A2 inhibitors (Figure 2).Serious adverse events considered related to bendamustine plus rituximab were neutropenia, transient acantholytic dermatosis, bacteremia, pyrexia, and febrile neutropenia, which is consistent with the serious adverse event profile for this combination in larger studies. ConclusionsThis graphical exploration suggests that no drug-drug interactions were demonstrated between bendamustine and rituximab or CYP1A2 inhibitors. Just as rituximab does not affect bendamustine PK, bendamustine did not appear to affect rituximab PK over time. Differences in observed rituximab concentration could be due to potential differences in assay methods/sensitivity or the length of rituximab infusion. Analysis of graphical displays of bendamustine concentrations in the presence and absence of CYP1A2 inhibitors suggests that CYP1A2 inhibitors do not increase systemic exposure to bendamustine, a result that is consistent with the near instantaneous hydrolytic metabolism of bendamustine. These results suggest that CYP1A2 is not a substantial metabolic pathway for bendamustine; however, whether CYP1A2 inducers have clinically relevant drug-drug interactions was not analyzed in this study. Based on results from this study, the likelihood of drug-drug interactions with bendamustine appears to be low.Support: Teva Pharmaceutical Industries Ltd. [Display omitted] [Display omitted] Disclosures:Darwish:Teva Pharmaceuticals: Employment. Bond:Teva Pharmaceuticals: Employment. Burke:Spectrum Pharmaceuticals: Consultancy. Hellriegel:Teva Pharmaceutical Industries Ltd: Employment. Robertson:Teva Pharmaceutical Industries Ltd: Employment. Phillips:Cognigen Corporation, which received research funding from Teva Pharmaceuticals: Employment, Research Funding. Grasela:Cognigen Corporation, which received research funding from Teva Pharmaceuticals: Employment, Research Funding.

Appraisal of IgM Kappa/IgM Lambda Variations Using HevyLite® After Rituximab As Consolidation Therapy in Patients with Waldenström’s Macroglobulinemia

AbstractAbstract 4879 Background:In Waldenström’s Macroglobulinemia (WM), response rates to rituximab (RTX) range from 30% to 75%. RTX is also known to generate a transient IgM flare in some patients. Our study aimed to characterize this phenomenon using the newly Hevylite® assay developed by The Binding Site that specifically quantifies IgM-kappa(K) and IgM-lambda(L). Methods:Between 2007 and 2009, 33 WM patients received a 6-course fludarabine-based regimen, 32 as first and 1 as second line. Partial response (PR), defined as a reduction ≥50% but <90% in baseline monoclonal peak (BMP) on serum protein electrophoresis (SPE), was the minimal goal. Two of the 32 patients treated at first line needed further treatment with CHOP regimen to reach the minimal PR. All received 4 consecutive weekly IV injections of 375 mg/m2 of RTX as consolidation. All were tested using the SPAplus analyzer for residual IgM-K and IgM-L at first and fourth injection. Monoclonal IgM-K or IgM-L level variations and IgM-K/IgM-L ratios were analyzed. SPE and immunofixation (IF) were performed 3 months after the fourth RTX injection to evaluate the impact of consolidation on response. Results:Of the 33 tested, at a variation threshold ≥20% relative to baseline IgM levels, 3 subgroups were defined: 15 or 45.5%, the no-flare patients, showed an increase or a decrease <20%, 11 or 33.3%, the flare patients, showed an increase ≥20% while 7 or 21.2%, the anti-flare patients, showed a decrease ≥20%. In the no-flare subgroup, there was no significant difference in monoclonal K (9) and L (6) light chains while in the 18 but 1 patients of the merged flare and anti-flare subgroups, monoclonal K light chain was overrepresented as there were 17 K vs 1 L (p<0.05). A highest initial IgM-K/L ratio (p<0.005) characterized the flare subgroup. No association was seen with demographic and hematologic characteristics and RTX pharmacokinetics. Regarding Fc gamma receptor (FcγR) IIIa-158 polymorphism, 10 patients were defined as VF and 5 as FF in the no-flare subgroup, 3 as VF, 3 as FF and 1 as VV in the anti-flare subgroup, 3 as VF and 8 as FF in the flare subgroup, with no statistically significant difference between the last 2 subgroups (p=0.22).We also analyzed the data without defining a critical threshold in monoclonal IgM level variations. Patients were, then, separated in 2 subgroups: 17 flare patients and 16 anti-flare patients. Monoclonal K light chain was overrepresented in the 2 subgroups, with 13 K vs 4 L in flare patients and 13 K vs 3 L in anti-flare patients; no statistically significant difference was found between them regarding monoclonal K and L light chains (p=0.32). No association was seen with demographic and hematologic characteristics and RTX pharmacokinetics. Regarding FcgR IIIa-158 polymorphism, there was a statistically significant difference (p=0.03) as 12 patients were of the FF subtype and 5 of the VF one in the flare subgroup while 4 were of the FF subtype, 11 of the VF one and 1 of the VV one in the anti-flare subgroup.At response evaluation 3 months after the fourth injection of RTX, all patients were screened for SPE and 22 for IF. When we compared the 17 patients of the flare subgroup to the 16 ones of the anti-flare subgroup, no patient vs 1 reached complete remission defined as disappearance of BMP on SPE and negativity of IF, 1 vs 6 advanced to very good partial response defined as a decrease ≥90% in BMP on SPE and positivity of IF and the remaining 16 vs 9 improved their partial response by a mean additional reduction of 20% (range: 0–30%) vs 18.75% (range: 0–50%) in BMP, but there was no statistically significant difference in this regard (p=0.79). Conclusions:Using Hevylite® assay, monoclonal IgM up and down variations of at least 20% following RTX occurred in up to 54.5% of our patients and this raised to 100% when no threshold value was considered. Such assay provides a reliable and highly sensitive quantitative assessment, especially in patients whose monoclonal IgM was at low concentration.Patients of the flare subgroup were predominantly of the FF subtype of FcgR IIIa-158 polymorphism while patients of the anti-flare subgroup were predominantly of VF subtype, which was statistically significant. It is tempting to hypothesize a difference in RTX mechanism of action, with ADCC predominating in the anti-flare subgroup and CDC in the flare subgroup. It would be of interest to assess such an hypothesis on a larger cohort and to analyze its impact on response profile. Disclosures:Pietrantuono:The Binding Site: Employment.

Rituximab pharmacokinetics in ascites and serum in the treatment of follicular lymphoma with massive ascites

Dear Editor, Rituximab is an effective and safe monoclonal antibody in the treatment of CD20+ malignant lymphoma [1]. In patients with massive pleural effusions or ascites associated with lymphoma, it remains unclear whether rituximab should be administered not only by intravenous infusion but also via the intraperitoneal route [2–4]. We studied the association of rituximab concentration and disappearance of lymphoma cells in ascites after intravenous application. The concentration of rituximab was low 3 h after infusion, but increased 24 h later. Flow cytometry (FCM) analyses revealed several CD20+ lymphoma cells at 3 h which completely disappeared 24 h later. These observations indicated that rituximab antibody may be resorbed from the circulation to the peritoneal cavity and may have an anti-tumor role in ascites. We report a patient who developed massive ascites due to a follicular lymphoma who was treated with rituximab. A 40-year-old male presented with generalized lymphadenopathy in 1999. Pathology of the lymph node revealed a grade-2 follicular B cell lymphoma with the chromosomal abnormality t(14;18). He received combination chemotherapy with cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP). After three cycles of CHOP, his symptoms promptly resolved. He discontinued hospital visits from 2001 onwards. This patient presented again to our hospital with generalized lymphadenopathy, abdominal discomfort, and massive ascites in 2008 (Fig. 1a). Biopsy of the left inguinal lymph node showed regrowth of a CD20+ grade-2 follicular lymphoma. A bone marrow specimen exhibited several atypical lymphocytes with t(14;18) by fluorescence in situ hybridization (FISH). Aspiration of ascites demonstrated CD20+ B cells and CD3+ T cells by FCM. FISH analyses showed t(14;18) cells (16%) in ascites. His symptoms (in particular, abdominal discomfort due to ascites) persisted despite the first cycle of rituximab plus CHOP (R-CHOP). We supposed that the concentration of rituximab in ascites by intravenous infusion was not sufficient to elicit an effect. Therefore, upon secondary therapy, we administered only rituximab without CHOP by intravenous infusion to examine the relationship between rituximab concentration and the disappearance of lymphoma cells in ascites. Concentrations of rituximab in ascites and serum were 0.8 and 98.1 μg/mL, respectively, 3 h after infusion, and 3.3 and 21.7 μg/mL 24 h later. Massive ascites was resolved after 3 days of infusion (Fig. 1b). FCM analyses demonstrated that CD20+ lymphoma cells (22.4%) in ascites at 3 h (Fig. 1c) completely disappeared 24 h later (Fig. 1d). The patient received six cycles of R-CHOP and was doing well. Fig. 1 CT of the abdomen before and after intravenous instillation of rituximab. a Massive ascites before infusion. b Three days after infusion, ascites was resolved. Changes in lymphoma cells double-positive for CD19+/20+ cells in ascites by FCM analyses. ... A few reports have suggested that lymphoma with massive effusions or ascites demands local therapies such as intraperitoneal administration of rituximab as well as systemic chemotherapy [2–4]. Unfortunately, those studies did not measure the level of rituximab in ascites, effusions, or serum before and after local instillation. Thus, whether local (not systemic) administration is required for the management of ascites or effusions is not known. The mechanism of action of rituximab is incompletely understood. However, rituximab is thought to induce cell destruction, including apoptosis, complement-dependent cytotoxicity (CDCC), and antibody-dependent cellular cytotoxicity (ADCC) [5]. In in vitro studies, rituximab induced direct cytotoxicity against the RAJI CD20+-expressing lymphoma cell line at >0.1 μg/mL [5]. That report showed that, after 4-h incubation in human serum, direct cytotoxicity was not evident, but after 3 days of culture, remarkable combined direct cytotoxicity and CDCC was induced [5]. In our case, we identified a rituximab concentration in ascites >0.1 μg/mL 3 and 24 h later only by intravenous administration. The remaining CD20+ lymphoma cells (22.4%) in ascites at 3 h completely disappeared 24 h later. These findings suggested that rituximab in ascites may mediate not only direct cytotoxicity against lymphoma cells, but also CDCC or ADCC induced by serum and several CD3+ cells in ascites as effector cells. The efficacy of rituximab may be dependent upon the relative ratio of effector cells (CD3+ lymphocytes) to target cells (CD20+ lymphoma cells) in ascites. However, the exact mechanism of cytotoxicity remains unclear. The pharmacokinetics and efficacy of rituximab should be investigated in a larger study population.

Identification of patients with indolent B cell lymphoma sensitive to rituximab monotherapy

The potential predictive value of tumor bulk, genetic, and immunological variants in patients with low-grade non-Hodgkin's lymphoma to respond to treatment with rituximab (RTX) monotherapy was evaluated. Thus, the value of assessing the effect of 18-fluoro-desoxy-D-glucose (FDG) uptake on PET scan, polymorphisms in Fc gamma receptor (FcγR) IIIa-158, FcγRIIa-131, and C1qA-276 genes in predicting the response to treatment were evaluated in 50 low-grade non-Hodgkin's lymphoma patients. The influence of RTX pharmacokinetics, plasma levels of the B cell-activating factor (BAFF), and human antichimeric antibodies was also investigated. The therapeutic response was evaluated 10 weeks after treatment using revised Cheson's criteria. Lower maximal standardized uptake values (SUV(max)) at baseline were predictive of complete response. FcγRIIIa-158 polymorphism was also associated with complete response to RTX confirming previous findings, whereas polymorphisms in the FcγRIIa-131 and C1qA-276 genes were not. Lower blood levels of RTX were observed in males, but the effectiveness of RTX in males and females was the same. BAFF was not detectable in plasma before or after treatment, and no patients developed human antichimeric antibodies. Low-grade non-Hodgkin's lymphoma patients with a low SUV(max) at baseline and an FcγRIIIa-158 V/V genotype generally had a complete response to RTX.

Subcutaneous Absorption of Monoclonal Antibodies: Role of Dose, Site of Injection, and Injection Volume on Rituximab Pharmacokinetics in Rats

To determine the effect of dose, the anatomical site of injection, and the injection volume on subcutaneous absorption of rituximab in rats and to explore absorption mechanisms using pharmacokinetic modeling. Rituximab serum concentrations were measured following intravenous and subcutaneous administration at the back, abdomen, and foot of rats. Several pharmacokinetic models were developed that included linear and saturable absorption, and degradation and/or protective binding at the injection site. Rituximab exhibited linear kinetics following intravenous administration; however, bioavailability following subcutaneous injection was inversely related to the dose level. For the 1mg/kg dose, bioavailability was approximately 70% at all tested injection sites, with faster absorption from the foot (T(max) = 12h for foot vs. 4.6days for back). Bioavailability for the 10mg/kg dose was 44 and 31% for the abdomen and back sites and 18% for 40mg/kg injected at the back. A pharmacokinetic model that included binding as part of the absorption mechanism successfully captured the nonlinearities in rituximab absorption. The anatomical site of subcutaneous injection influences the rate of absorption and bioavailability of rituximab in rats. Saturable binding may be a major determinant of the nonlinear absorptive transport of monoclonal antibodies.

Rituximab‐induced late onset neutropenia in newly‐diagnosed B‐cell lymphoma correlates with Fc receptor FcγRIIIa 158(V/F) polymorphism

Rituximab is a commonly utilized treatment agent for B-cell lymphoma. Late onset neutropenia (LON) has been identified as a complication associated with rituximab, primarily in conjunction with hematopoietic stem cell transplantation (HSCT). Scant data exists regarding rituximab-related LON outside the spectrum of HSCT, including newly-diagnosed lymphoma. We examined a large cohort of newly-diagnosed B-cell lymphoma patients treated with rituximab-based therapy. We identified patients with LON and analyzed the characteristics and outcomes. Furthermore, we utilized multiplex PCR for the detection of the FcgRIIIa 158 V/F polymorphism and correlated this with LON. Eighty consecutive B-cell lymphoma patients were examined. Nine of 80 (11.3%) patients developed LON. The clinical course of LON was generally self-limiting without adverse events. The onset of LON occurred at a mean of 66 days after the last course of treatment, while the mean duration of LON was 97 days. Moreover, the V/V and V/F polymorphisms were significantly associated with the occurrence of LON (P 5 0.046) yielding an odds ratio for the development of LON of1.47 (95% CI 1.21-1.78). We identified an incidence of LON following frontline rituximab-based treatment of 11.3%. The FcgRIIIa polymorphism was highly associated with development of LON.

Validation of an ELISA for the determination of rituximab pharmacokinetics in clinical trials subjects

Rituximab is a chimeric anti-CD20 monoclonal antibody that has revolutionised the treatment of many B-cell malignancies, and is now increasingly being used in non-malignant conditions such as auto-immune disorders. Serum rituximab levels are highly variable in patients receiving similar ‘standard’ approved doses. Little is known regarding the factors that affect serum rituximab concentration and that in turn may influence clinical outcome. In order to provide a tool that may ultimately enable patient specific dosing of rituximab therapy, we have validated a reliable, robust ELISA for the quantitation of serum rituximab levels to provide accurate pharmacokinetic (PK) data that will guide the optimisation of rituximab dosing regimes. Extensive validation of the assay was performed in order to utilise the assay for clinical applications. The within and between day plate coating reproducibility was tested and proved a robust starting platform for the assay. The within day precision for the assay was determined using spiked serum samples and was shown to have a coefficient of variation (CV) of < 10% with an accuracy between 91 and 125%. The between day precision (CV) was < 25% with an accuracy between 95 and 109%. Dilution linearity and parallelism were demonstrated. Spike recovery for all concentrations and donors was shown to be within +/−15% on average, with a CV below 10%. This assay is highly accurate and reproducible in determining the levels of rituximab in spiked serum samples. It meets stringent acceptance criteria, is fit for purpose, and is currently being applied to several clinical trials incorporating rituximab in the treatment of lymphoma. This assay represents a useful tool for clinical application of this widely used therapeutic.

Pharmacokinetics of rituximab in patients with CD20 positive B-cell malignancies

In this study, we investigated the pharmacokinetics of rituximab in patients with CD20 positive non-Hodgkin lymphoma, to get more insight into the factors that influence the pharmacokinetics of rituximab. This may aid to understand variability of treatment outcome, in patients with a CD20 positive malignancy treated with rituximab. In this study, patients with a CD20 positive B-cell malignancy who were treated with rituximab containing regimens were included. Induction treatment schedules consisted of a combination of rituximab with chemotherapy for 4-8 cycles. Maintenance treatment consisted of a 2 or 3-monthly dose of 375 mg/m2 rituximab intravenously for 2 years. On the day of the treatment with rituximab, preinfusion blood samples were taken. Also, after the end of treatment, selected blood samples were taken. Rituximab levels were measured with a validated enzyme-linked immunosorbent assay (ELISA). An antigen binding assay was applied for determination of human-antibodies against chimeric-antibodies (HACAs). Eight patients were on induction therapy. Rituximab levels of one patient on induction therapy remained very low after the first course. This patient had a chronic lymphoid leukemia with circulating tumor cells and a high tumor burden. Apart from one patient with mantle cell lymphoma, all patients on induction therapy had a complete response. Five patients were on maintenance therapy. Trough levels of 4 patients on three-monthly schedule maintenance therapy remained constant, with a median concentration of 6 mu g/mL (range 0.5-11.7 microg/mL). One patient had a relapse during his maintenance treatment. The elimination half-life at steady state of rituximab in all patients was estimated to be 19.2 (+/- 15.2%) days with a between-subject variability of 54%, indicating wide variability. Possible pharmacokinetic-pharmacodynamic relationship was observed as rituximab levels of the non-responders remained low compared to the rituximab levels of the responders. For all patients, concentration of HACAs remained below the quantification limit. Considerable inter-individual variability of rituximab levels was observed. Although the patient population was small, the results support the need for more research into the pharmacokinetics and factors that might influence the pharmacokinetic-pharmacodynamic relationships of rituximab in patients with non-Hodgkin lymphoma.

Rituximab pharmacokinetics during the management of acute idiopathic thrombotic thrombocytopenic purpura.

Increasingly, patients with acute, idiopathic, antibody mediated thrombotic thrombocytopenic purpura (TTP) are being treated with rituximab to achieve a durable remission, however, there is the potential that it is removed by plasma exchange (PEX). To look at the pharmacokinetics and pharmacodynamics of rituximab in patients with acute idiopathic TTP undergoing PEX. Patients who received rituximab for acute idiopathic TTP (group 1, n = 30) and a control group (group 2, n = 3) of TTP patients in remission receiving rituximab electively as maintenance were included. Rituximab levels were measured before/after each infusion, before/after PEX and in follow-up. ADAMTS-13 activity, anti-ADAMTS-13 IgG and CD19% were measured to assess response. The median number of PEX to remission after rituximab was 10 (range 4-25). In group 1 there was no significant incremental rise in the peak serum rituximab level until dose 4. Trough levels were lower in patients who had had PEX since their last rituximab infusion. In the control group, there was an incremental rise in the peak serum rituximab level and all patients had detectable trough levels. The median fall in rituximab per PEX was 65%. All patients achieved CD19 < 1%. In group 1, the median time to undetectable rituximab was 5 months (range 0-12 months) and to B cell return was 7 months (range 3-24 months). ADAMTS-13 increased and anti-ADAMTS-13 fell after therapy. There were three deaths and two relapses in group 1. Relapse was not temporally related to B cell return.

Mechanistic exposure‐response model of combination rituximab chemotherapy in murine lymphoma xenografts

ObjectivesTo develop a model for describing rituximab effects (CD20 antibody), alone and in combination with fenretinide or rhApo2L, in murine Ramos cell xenografts.MethodsPharmacokinetic (PK) and tumor growth data were extracted from the literature. Drug exposure driving functions were derived, and rituximab dynamics were set proportional to antibody‐dependent cell‐mediated cytotoxicity (ADCC) and complement‐dependent cytotoxicity (CDC) driven by CD20 occupancy. Combination regimen effects reflected changes in Bcl‐xL (rhApo2L) and NFκB (fenretinide) expression, and a series of transit compartments were evaluated for capturing signal transduction. All model parameter estimation and simulations were performed in MATLAB.ResultsCompartmental models well described fenretinide, rhApo2L, and rituximab PK. Efficacy and potency parameters of CDC and ADCC signals were estimated with good precision. Fenretinide and rhApo2L combination regimens required modified cell kill terms modulated by Fas induction and decreased Bcl‐xL expression. For validation, the model based on single agents was able to predict combination therapy synergism.ConclusionA mechanism‐based model of rituximab combination chemotherapy has been developed which is capable of predicting observed synergistic responses.This research was funded by the Center for Protein Therapeutics at the University at Buffalo

Abstract C184: Validation of an ELISA for the determination of Rituximab pharmacokinetics in clinical trials subjects

Abstract Rituximab is a chimeric anti-CD20 monocolonal antibody directed against the CD20 antigen on the B cell surface. Rituximab was originally developed by IDEC Pharmaceuticals from a high affinity murine monoclonal Antibody (mAb). CD20 is expressed on mature B cells, present on 95% of B cell lymphomas and is thought to act as a calcium channel that regulates progression through the cell cycle. Cross-linking of CD20 by anti-CD20 mAb results in apoptosis through a caspase dependent pathway and may trigger complement dependent cytotoxicity (CDC) or antibody directed cell cytotoxicity. Rituximab has revolutionised the treatment of many B-cell malignancies and is now increasingly being used in non-malignant conditions such as auto-immune disorders. Serum Rituximab levels are highly variable in patients receiving similar ‘standard’ approved doses, and little is known regarding the factors that affect serum Rituximab concentration and that in turn may influence clinical outcome. In order to provide a pharmacokinetic (PK) tool that may ultimately enable patient specific dosing of Rituximab therapy; we have generated a reliable, robust ELISA to quantitate serum Rituximab levels and to provide accurate PK data that will guide the optimisation of rituximab dosing regimes. An extensive validation of the assay was performed in order to use the assay in the clinical setting. The plate coating efficiency (within and between days) proved reliable resulting in a robust starting platform for the assay. The within day precision for the assay determined using spiked serum samples and was shown to be &amp;lt; 25% with accuracy between 96–109%. The between day precision was &amp;lt;10% with accuracy between 91–125%. Dilution linearity and parallelism were demonstrated with spike recovery for all concentrations and donors within +/− 15% with a coefficient of variation below 10%. The assay has been validated for use on clinical serum samples and consequently stability is a critical factor affecting analysis both in terms of freeze thaw cycles as well as length of storage. To establish sample stability criteria a retrospective study was undertaken using clinical samples collected four years previously as part of a previously published clinical trial. Reanalysed samples showed no statistically significant decrease in measured Rituximab levels compared to the first analysis. These same samples can be freeze thawed and analysed up to five times if necessary. This assay is highly accurate and reproducible in determining the pharmacokinetic levels of Rituximab in patient serum samples. This assay represents a powerful new tool for clinical evaluation of this widely used therapeutic, it meets stringent acceptance criteria, is fit for purpose, and is currently being applied to several clinical trials incorporating Rituximab in the treatment of Lymphoma. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C184.

Predictors of Early Response to Single Agent Rituximab in Patients with Indolent Lymphoma.

Abstract 1946Poster Board I-969 STATEMENT OF PURPOSE:Rituximab (RTX) is a chimeric antibody targeting CD20, a membranous B-cell specific protein, currently used in the treatment of B-cell malignancies and various autoimmune diseases. Despite its clear efficacy in non-Hodgkin's lymphoma, clinical and biological responses are highly variable between patients. Therefore, predictive factors are necessary to adapt therapy to each individual as early as possible. MATERIAL AND METHODS:We studied a set of biomarkers in a cohort of 39 patients with indolent non-Hodgkin's B-cell lymphoma (low grade follicular lymphoma or marginal zone lymphoma) and a low tumour burden, treated with single agent RTX as follows: 4 weekly 375 mg/m2 infusions, associated with short-course oral steroids. All patients benefited from FDG-PET analysis before treatment and 10 weeks later, allowing their classification into 3 early response groups: complete isotopic remission, partial response and no response. Dosage of rituximabemia, BAFF and Human Anti-Chimeric Antibodies (HACA) was performed with in-house ELISAs before each infusion and 2 months later. Polymorphisms of FcγRIIIa[158], FcγRIIa[131] and C1qA[276] genes were determined using allele-specific PCR or direct sequencing. Results were compared using Student's t-Test or two-tailed Fischer's exact test when appropriated. RESULTS:Referring to PET analysis, 18 patients were classified as complete isotopic remission, and 21 as partial or no response. Mean age was significantly lower in the first group (54 y+/-19 vs 66+/-13, p=0.022), as well as initial SUVmax (6.3+/-3.4 vs 10.8+/-6.8, p=0.024). Serum levels of RTX were highly variable between patients at each timepoint, but were not statistically different between the different response groups. Basal levels of BAFF were not statistically different between lymphoma patients and a control population, and did not vary after RTX-induced B cell depletion. No patient developed a HACA response during the study. Homozygous FcγRIIIa158V/V genotype was significantly associated with a complete response (p=0.037). Conversely, FcγRIIa[131] and C1qA[276] polymorphisms were not associated with the response. Genetic polymorphisms seemed to modify RTX pharmacokinetics, since homozygous FcγRIIa[131]H/H, FcγRIIIa[158]V/V and C1qA[276]G/G subjects displayed lower levels of RTX than others, without reaching statistical significance. CONCLUSIONS:Younger age, lower initial SUVmax and FcγRIIIa[158]V/V genotype are associated with a better early response to RTX in indolent lymphoma patients, as assessed by PET analysis. FcγR polymorphisms affect RTX kinetics probably through a modification of the efficacy of antibody dependant cell cytotoxicity. Determination of these factors could help in adapting individually therapeutic protocols. Disclosures:No relevant conflicts of interest to declare.

Pharmacokinetics of rituximab associated with CHOP chemotherapy in B‐cell non‐Hodgkin lymphoma

The pharmacokinetics of rituximab administered alone at a dose of 375 mg/m(2) once a week have been widely studied but few data are available for the administration of rituximab in combination with other agents. We carried out a pilot pharmacokinetic study in 10 patients with B-cell non-Hodgkin lymphoma treated with rituximab associated with chemotherapy on a three-weekly basis. Patients received four courses of rituximab (375 mg/m(2)) associated with CHOP (cyclophosphamide, adriamycin, vincristine and prednisone) at 21-day intervals. Blood samples were collected from each patient at selected times throughout the treatment period and analysed using a validated enzyme-linked immunosorbent assay. The NONMEM software was used to obtain population and post hoc estimates of rituximab pharmacokinetic parameters and to study possible time-dependent variation during treatment. Pharmacokinetic parameters obtained from 10 patients were similar to those described for studies in the absence of chemotherapy. We did not observe any intra-subject variation in pharmacokinetic parameters over the treatment period. Our results suggest that rituximab pharmacokinetics is not affected by combination with CHOP chemotherapy. Nevertheless, the substantial intersubject variability observed in this small sample size highlights the need for further study of the determinants of rituximab pharmacokinetic variability.

Two-Monthly Rituximab Infusions Maintain Target Concentrations: Pharmacokinetic Data of the AGMT-NHL9 Maintenance Study in Patients with Follicular Lymphoma in Complete or Partial Remission.

Maintenance therapy with the monoclonal anti-CD20 antibody Rituximab (R) improves clinical outcome in patients with follicular lymphoma in complete or partial remission (CR, PR) after induction therapy. However, optimal dosing schedule and duration of maintenance therapy are still under investigation. Commonly applied regimen include four doses of 375mg/m2 Rituximab once a week every 6 months, one dose every 3 months, or one dose every 2 months. Maintenance is usually given for 2 years. In addition to clinical trials, pharmacokinetic (PK) studies could help determining optimal dosing of Rituximab. PK values have been obtained during induction therapy in several trials and an association between drug trough levels and clinical effectiveness has been established (25μg/ml in responding patients). However, data on Rituximab PK during maintenance therapy are lacking. The Austrian Cooperative Study Group for Cancer Drug Therapy (AGMT) has conducted a phase II trial in 33 patients with follicular lymphoma in CS III/IV with an induction therapy with 6 cycles of Rituximab (375mg/m2 i.v., day 1), Fludarabine (30mg/m2 p.o., day 2–4)) and Mitoxantron (10mg/m2 i.v., day 1) every 4 weeks (R-FM) (NHL9). Thirty two patients obtaining a CR, CRu or PR received maintenance treatment with Rituximab 375mg/m2 every 2 months for 1 year. Pharmacokinetic data for Rituximab were obtained during induction and maintenance treatment in 16 of these patients. All patients converted from BCL2/IgH PCR positivity to negativity in peripheral blood after R-FM induction. PK serum concentrations during induction corresponded well to known data: A steady increase in pre- and post-dose levels was observed with increasing cycle number. Median, minimum and maximum Rituximab serum concentrations are given in the upper panel of Table 1:Table 1. Serum concentrations (μg/ml) during R-FM induction and two-monthly maintenance treatment with 375mg/m2 of Rituximab.R-FM InductionCycle 1246(Q 4 weeks)min.med.max.min.med.max.min.med.max.min.med.max.Pre-doseN.A.<0.5N.A.<0.516.844.825.356.693.243.266.5208.3Post-dose44.8161.4236.7146.4204.6300.5176.6218.4346.2196.3249.5370.2Interval 2 monthsR-MaintenanceCycle 1246(Q 2 months)min.med.max.min.med.max.min.med.max.min.med.max.Pre-dose13.841.5113.76.231.072.814.835.6261.213.726.756.0Post-dose136.6237.9364.6198.0235.9350.7214.2235.0280.9185.1249.2271.1The median interval between day 1 of the last R-FM induction cycle and the initial dose of R maintenance treatment was 2 months (range 1 – 3). PK values were obtained during maintenance cycles 1, 2, 4, and 6. The median interval from the first Rituximab maintenance infusion to the sixth infusion was 10.4 months (range 9.7 – 10.5). Table 1 (lower panel) shows the median, minimum and maximum drug levels before and after R maintenance infusions. These results indicate that (i) Median Rituximab levels 2 months after the last infusion of induction treatment are still higher than the supposedly active concentration of 25μg/ml; (ii) Rituximab target levels of over 25μg/ml are constantly maintained throughout one year of maintenance. (iii) Rituximab trough levels may still be low in some patients on a two monthly schedule. There was no difference in maintenance trough levels between CR and PR patients. So far, no relapses were observed. We conclude that infusion of Rituximab 375mg/m2 once every two month results in constant active serum levels in patients with follicular lymphoma in complete or partial remission. These data should help designing rational maintenance schedules for future clinical trials.

Rituximab, Bevacizumab and CHOP (RA-CHOP) in untreated diffuse large B-cell lymphoma: Safety, biomarker and pharmacokinetic analysis

Bevacizumab is a humanized monoclonal antibody directed against vascular endothelial growth factor (VEGF-A). Non-Hodgkin's lymphoma patients with high serum VEGF levels have an inferior survival compared to patients with low VEGF levels. Bevacizumab was administered through a central line at 15 mg kg−1 IV on day 1 followed by rituximab (R) and CHOP on day 2 for cycle 1 and day 1 for cycles 2 – 8. Serum levels of bevacizumab and R were measured at specified time points to assess pharmacokinetics (PK). Plasma and urine samples were also analysed for VEGF. Tumor samples were stained for VEGF, CD31 and factor VIII by immunohistochemistry. Thirteen patients with newly-diagnosed DLBCL received a total of 88 cycles (range 2 – 8, median 7). Best response included five CR, six PR, one SD and one PD with an overall response rate of 85% and complete response rate of 38%. The 12-month PFS is 77% and a median follow-up of 16.9 months for the surviving patients. All tumor samples stained strongly positive for VEGF and there was a marginal association between baseline plasma VEGF and response (p = 0.04). Patients with higher plasma VEGF levels were generally younger and had bulky disease. Micro-vessel density did not correlate with presenting disease characteristics, VEGF expression or response. The PK of bevacizumab and rituximab were not influenced by combined treatment. In this patient population, treatment with RA-CHOP did not result in any episodes of grade 3 or 4 proteinuria, heart failure or hemorrhage. The RA-CHOP combination was generally well tolerated and safe.

The pharmacokinetics of rituximab following an intravitreal injection

Rituximab is a monoclonal antibody directed against the CD20 B-cell antigen and is approved for the treatment of B-cell lymphoma. We investigated the pharmacokinetics of rituximab following intravitreal administration to assess the feasibility of treating primary intraocular lymphoma. Intravitreal injections of rituximab 0.1 ml (1 mg) were performed in rabbits. Drug concentrations in the aqueous and vitreous humor were measured at intervals from 2 to 17 days after administration. The half-life of the total amount of rituximab in the two compartments was calculated to be 4.7 days. The aqueous and vitreous humor drug levels decayed in parallel maintaining an average ratio of approximately seven. Fitting the data to a two-compartment model yielded a clearance from the aqueous humor of 1.2 μl/min. The clearance was less than the reported rate of aqueous humor outflow indicating that elimination by this route could have been sufficient to account for the disappearance of the drug from the eye. The duration of time over which sustained levels of rituximab were achieved suggest that intravitreal administration warrants further investigation as an approach to treating vitreous and anterior chamber infiltrates in patients with primary intraocular lymphoma.

Blood Pharmacokinetics of Various Monoclonal Antibodies Labeled with a New Trifunctional Chelating Reagent for Simultaneous Conjugation with 1,4,7,10-Tetraazacyclododecane-N,N′,N″,N‴-Tetraacetic Acid and Biotin before Radiolabeling

Knowledge of the blood pharmacokinetics of monoclonal antibodies is crucial in deciding the optimal time for starting the administration of a "clearing agent" or using a "clearing device." The primary purpose was to investigate whether the pharmacokinetics of various antibodies labeled with the same chelator and (111)In differed significantly after i.v. injection in immunocompetent rats. A new trifunctional chelator called "1033" containing a biotin and a radiometal chelation moiety is introduced, making it possible to use only one conjugation procedure for the antibody. Sixty-five non-tumor-bearing rats were included and divided into four groups (I-IV). The blood pharmacokinetics was investigated for rituximab, BR96, and trastuzumab labeled with 1033 and (111)In (I-III). The whole-body activity and activity uptake in muscle, liver, and kidney, which might explain differences in the early pharmacokinetics in blood, were also measured. hMN14 labeled with another chelator [1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA)], but with the same radionuclide ((111)In-biotin-DOTA-hMN14), was studied (IV). The blood pharmacokinetics from another 15 tumor-bearing rats was compared with those of non-tumor-bearing rats (III) by injection of (111)In-1033-BR96. No statistical difference was detected between the groups regarding the blood pharmacokinetics of rituximab, BR96, or trastuzumab. The pharmacokinetics and biodistribution of (111)In-biotin-DOTA-hMN14 exhibited a clear difference compared with others. There were no significant differences in the blood pharmacokinetics of (111)In-1033-BR96 between tumor-bearing rats and non-tumor-bearing rats. Different antibodies labeled with the trifunctional chelator 1033 and (111)In did not exhibit different blood pharmacokinetics, which means that the pharmacokinetics could be predicted irrespective of the IgG1 antibody chosen. A small tumor burden did not change the pharmacokinetics of the radioimmunoconjugates.

Population Pharmacokinetics of Rituximab (Anti‐CD20 Monoclonal Antibody) in Rheumatoid Arthritis Patients During a Phase II Clinical Trial

Rituximab is a B cell-depleting anti-CD20 chimeric IgGkappa monoclonal antibody being investigated for the treatment of rheumatoid arthritis. The purpose of this study was to develop a population pharmacokinetic model in rheumatoid arthritis patients. In addition, the final pharmacokinetic model was used to assess the variability in drug exposure (AUC0-infinity) for fixed versus body surface area-based dosing. A total of 102 patients were included in this population pharmacokinetic analysis. A 2-compartment pharmacokinetic model described the data reasonably well. Body surface area and gender were the most significant covariates for both CL and Vc. Body surface area alone only explained about 19.7% of the total interindividual variability of CL. In a simulation study, body surface area-based dosing normalized drug exposure over a wide range of body surface area but did not seem to improve the predictability of rituximab AUC0-infinity in rheumatoid arthritis patients. Therefore, no rationale for body surface area-based dosing for rituximab in rheumatoid arthritis patients was found.

Rituximab Plus CHOP Followed by Maintenance Rituximab as Initial Therapy for Advanced Stage Indolent Non-Hodgkin's Lymphoma (NHL); Initial Results of Induction Therapy in a Phase II Study.

Background: This study was designed to investigate the activity and safety of rituximab (Rituxan®, R) plus CHOP induction therapy followed by maintenance R in patients with previously untreated advanced stage indolent NHL. The initial results of R-CHOP induction treatment are reported here.Methods: 102 patients with Ann Arbor Stage III or IV indolent NHL (follicle center/follicular grade I/II or nodal marginal zone B-cell by REAL; Type B or C by IWF) were enrolled into this open-label, multi-center, community-based, Phase II, single-arm trial. Patients received 6 cycles of standard R-CHOP induction therapy (cyclophosphamide 750 mg/m2, vincristine 1.4 mg/m2, doxorubicin 50 mg/m2 all IV on Day 1; prednisone 100 mg/day Days 1–5). R 375 mg/m2 was given on Day 1 of each cycle except for cycle 1, when it was administered 2–3 days prior to CHOP chemotherapy. Subjects with an ongoing response (CR/CRu or PR) after induction receive maintenance R (4 weekly infusions) within 28 days after the completion of induction and repeated every 6 months for 2 years for a total of 16 maintenance R doses. The primary endpoints were CR/CRu after 6 cycles of induction and the incidence of R infusion-related toxicity during induction and maintenance therapy. Secondary endpoints included overall response rates (ORR), infusion times, serious adverse events (SAE) in induction and maintenance, and rituximab pharmacokinetics in maintenance therapy.Results: Baseline characteristics were: median age 57y (33.3% ≥60y); 40.2% female; ECOG performance status 0: 70.6%; Ann Arbor stage III: 28.4% and IV: 71.6%. Serum β2-microglobulin and LDH at baseline were above normal in 66.3% and 20.6%, respectively. CR/CRu at the end of induction in 102 patients was 51.0% (ORR 86.3%). 1% had progressive disease (PD) during induction. 2 subjects died during the induction period, both due to SAEs (probable pulmonary embolus, acute respiratory distress). SAEs occurred in 22.5% of subjects during induction with the most common event being febrile neutropenia (7.8%). Grade 3–4 R infusion-related toxicity occurred in 4.9% of patients in cycle 1; this decreased to 1% by cycle 2 of induction. Two subjects discontinued R for infusion-related toxicity. 44% of patients were able to receive their R dose within 3 hrs at cycle 2 and 69.2% of patients at cycle 6.Conclusion: Patients with advanced stage indolent NHL treated in the community setting with R-CHOP induction tolerate therapy well and have an excellent response rate. The outcomes of maintenance therapy and rituximab pharmacokinetics in maintenance await further follow-up.