MXPA00011795A - Recombinant human interferon beta-1a (ifn-beta-1a) formulation - Google Patents

Abstract

Liquid compositions comprising a buffer of pH of about 7.2, recombinant interferon beta and 15 mg/ml of human serum albumin, and kits for parenteral administration comprising said compositions.

Description

FORMULATION OF BETA INTERFERON lA (IFN-ß-lA) RECOMBINANT HUMAN John Alam1, Susan Golez2 and Mark Rogge3. 1 Medical Affairs, Biogen, Inc., Fourteen Cambridge Center, Cambridge MA, 02142. 2 Cellular Biochemistry, Biogen, Inc., Fourteen Cambridge Center, Cambridge MA, 02142. 3 Preclinical Development, Biogen, Inc., Fourteen Cambridge Center, Cambridge MA, 02142 Biogen, Inc. 14 Cambridge Center Cambridge, MA 01921 Phone: 1-617-679-3126 Fax: 1-617-679-3170 INTRODUCTION Interferons are a family of proteins that have antiviral activity, inhibit cell proliferation and modulate the natural immune response (I). The beta human interferon (IFN-β), a member of this family, is a glycoprotein of 166 amino acids produced by fibroblasts, as well as by other cells, after induction by viral infection or by double-stranded RNA. During the previous 20 years, the following three forms of IFN-β have been clinically tested in a wide variety of human disorders (2): natural human IFN-β produced from human foreskin fibroblasts (n-IFN-β), Recombinant human IFN-β produced in E. coli (IFN-β-lb, which contains a genetically substituted serine by cystine in position 17) and recombinant human IFN-β produced in ovarian cells of Chinese hamsters (IFN-β- la, which contains the natural human amino acid sequence). Both n-IFN-β and IFN-β-la are glycosylated with a single complex carbohydrate unit bound to N, while IFN-β-lb is not glycosylated. AVONEX ™, a product of IFN-ß-la, demonstrated a treatment effect delaying the development of physical disability and decreasing the frequency of clinical exacerbations in patients with relapsing multiple sclerosis, when administered intramuscularly (IM) a dose of 6 million units (MU) once a week (3). A second IFN-β-la product, Rebif®, is administered subcutaneously (SC) during the course of phase III trials in the treatment of multiple sclerosis. Previous studies with AVONEX ™ and Rebif® have. The concern was raised that while the two products are designated IFN-β-la, they may not have similar pharmacokinetic and subsequent pharmacodynamic profiles.

In order to determine whether AVONEX ™ and Rebif® can be used interchangeably, an intrasubject cross-comparison of the pharmacokinetics and pharmacodynamics of the two products was performed after IM injection in healthy male and female volunteers. The results reported here demonstrated the surprising finding that AVONEX ™ and Rebif® are not equivalent when administered intramuscularly. AVONEX "is formulated at higher albumin concentrations (15 mg / mL after reconstitution against 9 mg / mL for Rebif®), at a different pH (7.2 against 5.5) and in a different buffer (phosphate against acetate) (5,7,15,16). In addition, Rebif® contains mannitol in its formulation and AVONEX ™, no. These differences in formulation can contribute to the unexpected altered absorption of IFN-β after IM injection, by affecting the extracellular matrix binding of the muscle and / or inactivation by pH-dependent proteases (17). To further evaluate the effect of IM injection on these properties, an additional objective of this study was to evaluate the framacokinetics and pharmacodynamics of a specific formulation of IFN-β-la, AVONEX ™, after intravenous (IV), subcutaneous (SC) administration. ) or intramuscular (IM) in healthy volunteers. Three groups, each with 8 healthy male and female subjects, received a single dose of 60 μg (12 million units) of IFN-β-la, either by intravenous infusion (IV) of 30 minutes or by subcutaneous injection ( SC) or intramuscular (IM). We showed that the levels of serum interferon activity showed a maximum immediately after the end of the IV infusion and then adjusted to a biexponential decrease model. Interferon activity was consistently detectable in serum after IM injection but not after SC injection. The pharmacodynamic response, as determined by changes in serum concentrations of neopterin and ß2-microglobulin, was also maximal after IM injection, followed by SC and then IV administration. These results indicated that the optimal pharmacodynamic and pharmacokinetic response is produced by the administration of IFN-β-la by the IM route, rather than by the SC or IV. Preferred embodiments of the invention for IM administration, include a packaged system or equipment for parenteral administration of the liquid formulations herein. The package may include syringes pre-filled with the liquid formulations of the invention, various swabs with alcohol, at least one needle, one or more adhesive bandages and instructions for use. It will also be understood that the present liquid formulations of the invention can be used with conventional needleless injection systems.

METHODS Study design A. Avonex versus Rebif This was a cross-sectional, randomized, open-label, single-center, two-treatment study designed to compare the pharmacodynamic and pharmacokinetic profiles of AVONEX ™ and Rebif® after IM injection in healthy volunteers. Male and female subjects, aged between 18 and 45 years, who were within 15% of normal body weight for height, with a minimum body weight of 50 kg and who gave written informed consent were eligible for the study. Fifteen male subjects and 15 female subjects were enrolled. The subjects received by means of IM injection a dose of 6 MU of AVONEX ™ and a dose of 6 MU of Rebif® administered with 2 weeks of separation. All subjects except one completed the study. A woman who received Rebif® in the first study period developed headaches and pharyngitis; she withdrew before the AVONEX ™ injection. This subject was not included in any of the pharmacodynamic or pharmacokinetic analyzes. The intramuscular injection was applied to the anterolateral part of the thigh. Doses were administered at equivalent contralateral injection sites, ie, AVONEX ™ and Rebif® were not applied at the same injection site. The subjects entered the clinical pharmacological unit 24 hours before each injection and remained in the unit until the end of the evaluation 24 hours after dosing. The subjects were kept in bed for one hour after each injection; then they were allowed normal activity. Subjects received paracetamol (acetaminophen) after each injection to reduce the likelihood of cold-like symptoms associated with interferon administration. They were instructed not to take any other medication during participation in the study. Blood samples were collected for pharmacokinetic determinations, immediately before each dose and at 6, 9, 12, 18, 24, 30, 36 and 48 hours post-dosing. Pharmacodynamics was estimated by evaluation of serum neopterin concentration profiles. Neopterin is a product of GTP cyclohydrolase induced by interferon; post-injection serum neopterin elevations reflect a slow biological response to receptor activation by IFN-β P1194 (8.9). Blood for determination of serum neopterin concentration was collected immediately before each dose and at 6, 12, 18, 24, 30, 36, 48, 72, 96 and 144 hours after each dose. Adverse events were recorded throughout the study. Hematologic and blood chemistry routine tests were performed 144 hours after each injection. This study was conducted in the United Kingdom and adhered to the principles of the Declaration of Helsinki.

B. Simple dose study This was a single dose, parallel group study to evaluate the comparative pharmacokinetics and pharmacodynamics of IFN-β-la by three different routes of administration: SC, IM and IV infusion for 30 minutes. Were eligible for this study, male and female subjects, aged between 18 and 45 years, who were within 15% of normal body weight for height, with a minimum body weight of 50 kg and who gave informed consent. The subjects were considered healthy at the end of the following: a physical examination; an electrocardiogram (ECG); and urine and blood laboratory tests, including tests for Hepatitis B, Hepatitis C and HIV antigen. Female subjects in pregnancy or lactation period were excluded.

P1194 Twenty-four subjects (12 men and 12 women) were enrolled and assigned to groups of eight to receive 60 μg of IFN-β-la, for each SC or IM injection or as an IV infusion for 30 minutes. They entered the same number of men and women in each group. Blood was taken for pharmacokinetic tests at 1, 3, 6, 9, 12, 15, 18, 24 and 48 hours after each SC or IM injection and at 10, 20 and 30 minutes after the start of the IV infusion and at the 5, 10, 20, 30, 45, 60 and 90 minutes and at 2, 3, 4, 5, 6, 9, 12, 15, 18, 24 and 48 hours after the end of the IV infusion. Measurements of two biological response markers induced by interferon, neopterin and serum β2-microglobulin (9) were made at 6, 12, 24 and 48 hours after the administration of the study drug. Urinalysis, blood chemistry and hematology tests were performed 48 hours after the application of the study drug. Adverse events were monitored throughout the study period. The study was carried out in the clinical pharmacology unit of the United Kingdom after the approval of the local ethics committee.

Study drug A. Comparison of Avonex against Rebif AVONEX ™ was supplied as a sterile lyophilized powder containing IFN-β-la, HSA (human serum albumin), sodium phosphate and sodium chloride; prior to injection, the contents of the vials were reconstituted with sterile injectable water. Rebif® was available as a sterile lyophilized powder containing IFN-β-la, mannitol, HSA (human serum albumin) and sodium acetate. The Rebif® was reconstituted with a solution of sodium chloride (0.9% NaCl) for injection. Each product was stored as a lyophilized powder at a temperature between 2 ° C and 8 ° C before reconstitution. The AVONEX ™ was packaged in 6 MU vials and the Rebif® in 3 MU vials. The activity of each one was accepted as established in the label. Each vial of Rebif® was reconstituted according to the instructions using 1 mL of the supplied vehicle. Two reconstituted vials were combined to prepare a dose of 6 MU in a volume of approximately 2 mL. Each vial of AVONEX ™ was also reconstituted according to the instructions to prepare 1 mL of solution. However, in order to inject equal volumes of the two test drugs, placebo vials containing only excipient, to equalize were also reconstituted with 1 mL of sterile water. Each of the reconstituted vials of AVONEX ™ and placebo were then combined to prepare a 6 MU injection in a volume of 2 mL.

B, Single dose study IFN-β-la (Avonex ™) was supplied as a lyophilized powder in vials containing 60 μg of IFN-β-la, as well as human serum albumin, sodium chloride and sodium phosphate. Each vial was reconstituted with 1 mL of sterile water before injection. The specific activity of IFN-β-la used in this study was 200 million units (MU) of antiviral activity (interferon) per milligram of IFN-β-la protein. Thus, each vial contained 12 MU of interferon activity.

Test methods A. Comparison of Avonex against Rebif The serum levels of IFN-β were quantified in Biogen using a cytopathic effect bioassay (CPE). The levels of interferon-mediated antiviral activity, determined in the CPE assay, were proportional to the concentration of active interferon in the serum sample. Historically, this assay has been the standard method for evaluating the pharmacokinetics of IFN-β (10,11). The CPE assay detected the ability of interferon beta (IFN-β) to protect human lung carcinoma cells (A549, # CCL-185, ATCC, Rockville, MD) from cytotoxicity due to the encephalomyelocarditis virus (EMC). The cells were preincubated between 15 and 20 hours with serum samples that allow the induction and synthesis of interferon-inducible proteins that produce an antiviral response. After preincubation, EMC virus was added in each well and incubated for 30 more hours; Cytotoxicity was determined using a methyl violet stain. An internal Biogen IFN-ß standard was tested in parallel with the samples in each assay plate. This pattern was calibrated against a reference standard of natural human fibroblast interferon (HO Second International Standard for Interferon, Human Fibroblast, Second International Standard for Interferon, Human Fibroblast, Gb-23-902-531) (12). The samples and serum standards were analyzed in duplicate in each of the two repeated test plates, giving four data points per sample. The geometric mean concentration of the four duplicates was reported. The inter-assay variability was determined by calculating the 95% confidence interval with respect to the average concentration of the internal IFN-β pattern for 323 test plates. As defined, the variability was P1194 less than 10% of the average. The limit of quantification was generally 10 U / mL. Serum neopterin concentrations were determined using a commercially available 125IRIA kit (Immuno Biological Laboratories, Hamburg, Germany). The study personnel who carried out pharmacokinetic and pharmacodynamic trials were blinded to the treatment assignment.

B. Simple dose study The levels of IFN-β in serum were quantified in Biogen using a cytopathic effect bioassay (CPE). The CPE assay measures levels of antiviral activity mediated by interferon. The level of antiviral activity in a sample reflects the number of active interferon molecules contained in that sample at the time the blood was drawn. This approach has been the standard method to evaluate the pharmacokinetics of IFN-β (11). The CPE assay used in the current study detects the ability of beta interferon (IFN-ß) to protect human lung carcinoma cells (A549, # CCL-185, ATCC, Rockville, MD) from cytotoxicity due to encephalomyelocarditis virus (EMC). The cells were preincubated between 15 and 20 hours with serum samples that allow the induction and synthesis of interferon-inducible proteins that then produced an antiviral response. Later, EMC virus was added and incubated for an additional 30 hours before carrying out the cytotoxicity evaluation using a methyl violet stain. An internal Biogen IFN-ß standard was tested in parallel with the samples in each assay plate. This pattern had been calibrated against a reference standard of natural human fibroblast interferon (WHO Second International Standard for Inferior, Human Fibroblast, Second International Standard for Interferon, Human Fibroblast, Gb-23-902-531) (12). Each assay plate also includes cell growth control wells that contained neither interferon nor EMC and virus control wells containing cells and EMC, but not beta interferon. Control plates containing the standard and samples were also prepared to determine the effect, if any, of the samples on cell growth. These plates were stained without the addition of viruses. Samples and standards were analyzed in duplicate on each of the two repeated test plates, giving four data points per sample. The geometric mean concentration of the four duplicates was reported. The limit of detection in this assay is 10 units (U) / mL. The serum concentrations of neopterin and β2-microglobulin were determined in the pharmacology unit P1194 clinical using commercially available assays. The neopterin determinations were not performed for the IV dosing group, due to the lack of availability of the assay by the manufacturer, at the time this group was enrolled.

Pharmacokinetic and statistical methods A. Comparison of Avonex against Rebif The standard descriptive analysis of the serum interferon activity data for the two IFN-β-la products was performed. The following pharmacokinetic parameters were calculated: (i) area under the curve, AUC (Uxh / mL), from 0 to 48 hours post-dosing, using the trapezoidal algorithm; (ii) maximum observed serum activity, Cmay (U / mL), by inspection; and (iii) time for maximum serum activity, tma? (h), per inspection. In the calculation of AUC and Cmax, the baseline serum interferon activity was subtracted from all post-dose values; all non-detectable values were adjusted to 0 U / mL. However, in the calculation of the relative bioavailability, the undetectable post-dose values were adjusted to 5 U / mL (representing the P1194 half of the lower limit of quantification). This was done because the serum interferon activity levels were 0 U / mL throughout the period after the administration of Rebif® in five subjects and after the administration of AVONEX ™ in a subject. In the case of the five subjects with Rebif®, adjusting the non-detectable concentrations to 5 U / mL, allowed the calculation of a finite relative bioavailability. The AUC and Cmax were analyzed using a two-way cross-variance analysis (ANOVA). Terms in the analysis included sequence, subject, sex, period and treatment (13). A term for the interaction of the genus with the treatment was included at the beginning, but later it was withdrawn because the interaction was not significant. The AUC and Cmay were transformed logarithmically before analysis. The following pharmacodynamic parameters were calculated: (i) area under the curve normalized to the baseline, from 0 to 144 hours, EAuc; (ii) maximum increase from baseline, (III) time, tmax, at which the maximum effect was observed; and (iv) concentration difference between 0 and P1194 144 hours after dosing. All EAUC and Emax values were corrected with respect to the baseline. The statistical analysis was performed in EAUC loge and Ema * loge using the two-way crossed ANOVA described above. Estimates were also made of the EAUC and Emax ratios of AVONEX ™ and Rebif® with 90% confidence limits. To determine if the concentrations of serum neopterin at 144 post-dosing hours were still high after the administration of each product, the differences between the baseline value and that of time 144 were compared to zero by a paired t-test.

B. Simple dose study Rstrip® software was used (MicroMath, Inc., Salt Lake City, UT), to adjust the data to pharmacokinetic models. The geometric mean concentrations were plotted by time for each group. Since the test results are expressed in dilutions, the geometric means are considered more appropriate than the arithmetic means. Serum interferon levels were adjusted for baseline values and non-detectable serum concentrations were set at 5 U / mL, which represents half of the lower limit of detection. For the IV infusion data, a P1194 model of IV infusion of two compartments to the detectable serum concentrations for each subject and the data of SC and IM were adapted to a model of injection of two compartments. Pharmacokinetic parameters were calculated (20) following: (i) maximum observed concentration, Cmax (U / mL); (ii) area under the curve from 0 to 48 hours, AUC (U.H / mL) using the trapezoidal rule; (iii) elimination half-life; and additionally from the IV infusion data: (iv) distribution half-life (h); (v) depuration (mL / h) (vi) apparent distribution volume, Vd (L). The inNonlin software (Version 1.0, Scientific Consulting Inc., Apex, NC) to calculate the elimination half-lives after SC and IM injection. For ß2-microglobulin and neopterin, arithmetic means were presented per time for each group. Ema, the maximum change from the baseline, was calculated. Cmax, AUC and Emax were subjected to a one-way analysis of variance to compare dosage groups. Cax and AUC were transformed logarithmically before analysis; the geometric means were reported.

P1194 RESULTS AND ANALYSIS A. Comparison of Avonex against Rebif Figure 1 shows the average serum interferon activity per time, for each product. At each post-dosing time point, the mean serum interferon activity subsequent to the administration of AVONEX ™ was greater than that following the administration of Rebif®. Table I summarizes the pharmacokinetic parameters for each product and the results of the cross-variance analysis. The minimum mean squared values of AUC for AVONEX ™ and Rebif® were 824 and 403 Uxh / mL, respectively. The average minimum squared ratio of AUC for AVONEX ™ and Rebif® was 204% with 90% confidence limits of 172 to 243% (p <0.001). The minimum mean squared values of Cmax were 33.8 U / mL after the administration of AVONEX ™ and 15.2 U / mL after the administration of Rebif®. The average minimum squared ratio of Cma? for AVONEX ™ and Rebif® it was 222% with 90% confidence limits of 172 to 285% (p <0.001). The average time for the maximum concentration was between 12 and 16 hours for AVONEX ™ and Rebif®. The pharmacodynamic effect of AVONEX ™ was comparable with the pharmacokinetic findings. Figure 2 illustrates the geometric mean neopterin concentrations related to the drug, with respect to the time after each treatment. After each product, neopterin concentrations rose during the initial 36 hours; the concentrations remained at a plateau of 36 to 72 hours post-dosing and then declined gradually. However, the induction of neopterin was greater for AVONEX ™ compared to Rebif®. The mean concentrations during the time period from 36 to 72 hours post-dosing were approximately 12.0 nmoles / L for AVONEX ™ and 9.3 nmoles / L for Rebif®. The concentrations of neopterin at 144 hours post-dosing were significantly higher than those of pre-dosing for each treatment (p <; 0.001). Table II summarizes the pharmacodynamic parameters, EAUc and E ^ x. The minimum average EAUC values for AVONEX ™ and Rebif® were 693 and 481 nmoles x h / L, respectively (p <0.001). The average EAUC ratio for AVONEX ™ and Rebif® was 144% with 90% confidence limits of 131 to 159%. The minimum average squared values of Emax for AVONEX ™ and Rebif® were 9.5 and 6.9 nmoles / L, respectively (p <0.001). The average ratio of Emax for AVONEX ™ and Rebif® was 138% with 90% confidence limits of 123 to 156%. All subjects were included in the safety analysis. After AVONEX ™, 21 of 29 subjects (72%) P1194 experienced an adverse event; after Rebif®, 21 of 30 subjects (70%) experienced an event. Adverse events related to interferon-mediated catarrhal syndrome were observed after treatment with both products. The most commonly reported adverse event was headache: 40% of subjects after Rebif®; 38% of subjects after AVONEX ™. Nausea, back pain and muscle pain were reported in 24%, 28% and 31% of subjects, respectively, after injection with AVONEX ™ and in 10% or less subjects after dosing with Rebif®. The higher incidence rates of these last three symptoms may be related to the relatively greater bioavailability associated with AVONEX ™. No reactions were reported at the injection site with any of the treatments. At the same time, these results indicate that two proteins that may be similar in their biochemical properties do not possess identical absorption profiles or pharmacodynamic effects after IM administration. Potential explanations for this discrepancy include: (i) unidentified structural differences between the two molecules; (ii) differences in the amount of drug administered or (iii) differences in formulation. With respect to the first possibility, since the two molecules are produced by inserting the natural human gene for IFN-β in Chinese hamster ovarian cells, it is unlikely that there are significant structural differences in the portions of active drug contained in AVONEX ™ and Rebif ®. Instead, the amounts of drug with less mass in the respective vials could partially explain the lower dose absorbed after the Rebif® injection. Based on the product label, the study subjects received identical doses of each product. However, it has been reported that the specific activity of IFN-β-la in Rebif® is 3 x 108 units / mg (ie, 300 MU of antiviral activity per milligram of IFN-β-la protein), while AVONEX ™ has a specific activity of 2 x 108 units / mg (14). This would indicate that a 6 MU vial of Rebif® contains 20 μg of IFN-β-la, while a 6 MU vial of AVONEX ™ contains 30 μg of IFN-β-la. Direct confirmation of this difference could not be obtained because both products are formulated with a 300-fold excess of HSA that interferes with the precise determination of the IFN-β-la concentration. However, this apparent difference in the mass of the dose does not fully justify the difference two times greater in pharmacokinetic measurements. As will be discussed later, another known difference that could explain the results is the formulation.

P1194 Significantly, the differences in formulation and the differences observed in pharmacokinetic and pharmacodynamic parameters suggest that precise claims regarding the clinical efficacy of Rebif® administered IM to patients with multiple sclerosis require clinical studies with Rebif® administered by that route. Until such data become available, the findings provided here indicate that the simple replacement of Rebif® IM by AVONEX ™ at similar doses according to the label is unlikely to replicate the therapeutic effects observed with AVONEX ™.

B. Demographic Simple Dose Study Table III summarizes the baseline demography for each group. Age, height and weight were similar in the groups.

Pharmacokinetics Figure 3 shows average levels of interferon activity per time, starting from the beginning of the IV infusion. The maximum levels Cmay were detected either at 20 minutes (in a subject) or at 30 minutes, ie when the infusion was stopped (in the other subjects). The CMA? ranged from 160 to 640 U / mL. The data was very well described by the two compartment model. From this model, the average distribution and elimination half-lives were 4 minutes and 4 hours, respectively. The mean distribution volume was 61.6 L and the mean total clearance was 334 mL / h / kg. Figure 4 shows average serum interferon activity levels per time, after SC and IM injection. Maximal interferon activity levels after IM injection were 40 U / mL in all but one subject, in whom the maximum level was 80 U / mL. Mean Cmax after SC administration was 20 U / mL. However, serum interferon activity levels remained below the limit of detection at all time points after injection in 4 of 8 subjects in the SC dosing group. After IM administration, AUC and Cmax were significantly higher than after SC administration (p <; 0.001 and p = 0.033, respectively). The mean apparent elimination half-life was 10.0 hours after the IM injection and 8.6 hours of the SC injection, both greater than those observed in the IV infusion group. Since the depuration mechanisms are likely to be the same as after the IV infusion, these elimination half-life estimates indicate prolonged absorption from the SC or IM injection site.

P1194 A C? M greater than an AUC? V is an unexpected result, but can be justified by the prolonged absorption phase, which can lead to an increase in serum exposure (see analysis). Bioavailability was not calculated, since the available data were insufficient to reliably adjust the AUC ratio for the prolonged absorption time. Table II summarizes the pharmacokinetic results.

Pharmacodynamics The mean serum concentrations of ß2-microglobulin and neopterin per time are shown in Figures 5 and 6, respectively. The maximum levels of the two markers were observed either at 24 or at 48 hours post-dosing. The mean Emav for serum β2-microglobulin was 796 μg / L after IM injection, 628 μg / L after SC injection and 559 μg / L after IV infusion (p = 0.050 for IM versus SC, p = 0.008 for IM against IV). Emax for serum neopterin was 16.0 nmol / L after IM injection and 12.4 nmol / L after SC injection (p = 0.18). The values are summarized in Table IV. This simple study defined the pharmacodynamic and pharmacokinetic response profiles of the single dose of this product IFN-ß-the particular, AVONEX ™, P1194 after IV, SC or IM administration. After a 30-minute IV infusion, IFN-β-la is rapidly redistributed in a few minutes, then cleanses more slowly for several hours. The elimination half-life observed in this study was approximately 4 hours compared to the 1.5 to 5 hours previously reported for other beta interferon products. Similarly, serum interferon activity levels raised to a minimum after SC administration are consistent with other beta interferon products. However, detectable interferon serum levels had not been observed after IM administration of these other beta interferon products. [See, for example (15)]. In the present study, detectable interferon levels were consistently observed after the IM injection. In addition, the AUC of the serum interferon activity after the IM injection was 2 to 3 times higher than after the SC administration. The finding of increased absorption after the IM injection of the IFN-β-product used in the present study, compared to the SC injection, is unlikely to be due to an intrinsic property of the active drug molecule. In a separate study (7), no differences in pharmacokinetics or pharmacodynamics were reported between the IM and SC injection of Rebif®. The P1194 IFN-β-la molecules used both in the present study and in the other are produced by inserting the natural human beta interferon gene into Chinese hamster ovarian cells. Consequently, it is unlikely that there are significant structural differences in the active drug units of the two products. In addition, after IV administration, the pharmacokinetic properties of IFN-β-Avonex used in the present study are similar to those reported previously. Since the impact of the inactive components of the product (ie excipients) would be expected to be minimal after IV administration, similar pharmacokinetic parameters in IV administration for the various beta-interferon molecules suggest that the intrinsic pharmacokinetic behavior of these molecules is not substantially different. The most likely explanation with respect to the ability to deliver interferon to the bloodstream after IM injection of the present product is related to the inactive components of the product (ie, the excipients). Paulesu [17] et al., Suggested that malabsorption of many beta interferon molecules after IM injection may be due to the binding of beta interferon to the extracellular matrix in the muscle or to inactivation by proteases. They also demonstrated that muscle homogenate can inactivate IFN-β and not IFN-α. This inactivation was partially blocked by albumin, suggesting that manipulation of the formulation in this form could increase the bioavailability of beta interferon after IM injection. The various beta interferon products differ in their formulations and these differences can also affect the interaction with local factors in the muscle. For example, both the IFN-β-lb and the IFN-β-la used in the present study are formulated at physiological pH and contain 15 mg / mL of human albumin as a stabilizer (18). However, IFN-β also contains dextrose, and IFN-β-la, no. The second product IFN-β-la has a pH of 5 to 5.5 after reconstitution and contains mannitol and 9 mg / mL of human albumin as stabilizers (15). The absorption of SC and IM injection sites was prolonged. Consequently, the bioavailability for the two administration routes could not be calculated. The standard method for determining bioavailability is by calculating the fraction absorbed (for example, AUC? M / AUCIV). For this single relationship, the apparent bioavailability was greater than 100%. However, this argument may lead to an overestimation of bioavailability P1194 when the absorption process is considerably less than the elimination process ("flip-flop" switching kinetics). (19) Under these circumstances the AUC? M values need to be adjusted by means of their appropriate velocity constant (ie, the apparent elimination rate constant ke? Should be replaced by the apparent absorption rate constant ka). In the present study, said adjusted bioavailability could not be calculated due to the inter-subject variability in absorption and elimination rates.

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P1194 8. Fuchs D, Weiss G, Reibnegger G, Wachter H, Crit J? Ev Clin Lab Sci 29, 307-341 (1992). 9. Witt PL, Storer BE, Bryan GT, Brown RR, Flashner M, Larocca AT, Colby CB, Borden EC, J "Iiunouno therapy, 13, 191-200 (1993) 10. Chiang J, Gloff C, Yoshizawa C , Williams G, Pharmaceutics Res 10, 567-57 (1993) 11. Chiang J, Gloff CA, Soike K, Williams G, J Interferon Res, 13, 111-120 (1993) 12. WHO Expert Committee of Biological Standarization , 38th World Health Organization Technical Report Series 771, p. 36-38 (1988). 13. Kenward M, Jones B: Design and Analysis of Cross-Over Triáis. Chapman and Hill, 1989. 14. Canosi U, Macia M, Gazza L, Serlupi-Crescenzi O, Donini S, Antonetti F, Galli G. J Immunol Methods, 199, 69-76 (nineteen ninety six) . 15. Liberati A, Horisberger M, Palmisano L, Astolfi S, Nastari A, Mechati S, Villa A, Mancini S, Arzano S, Grignani F, J Interferon Res, 12, 329-336 (1992). 16. Liberati AM, Garofani P, De Angelis V, Clemente FD, Horisberger M, Cecchini M, Betti AR, Palmisano L, Astolfi S, Nastari A, Villa A, Villa A, Arzano S, J Interferon Res, 14, 61- 69 (1994). 17. Paulesu L, Pessina GP, Bocci V, Proc Soc Exp Bio Med, P1194 200, 414-417 (1992). 18. 1997 Physician's Desk Reference. 19. Welling, P.G., Pharmacokinetics, American Chemical Society, Washington, DC 1986. 20. Gibaldi, M., Perrier, D., Pharmacokinetics, Marcel Dekker, New York, 1982.

Table I Summary of the Analysis of Pharmacokinetic Parameters Study Drug AUC1 C '-max2 t'-max2 (U x h / mL) (U / mL) (h) AVONEX ™ 824 33.8 15 Rebif® 403 15.2 12 Relationship: AVONEX ™ and Rebif® 204% 222% N.A.

I.C. 90% 172-243% 172-285% N.A.

Value p < 0.001 < 0.001 > 0.05 1 The values are minimum square means transformed in the inverse of the logarithmic scale, based on the analysis of variance with adjustments for subjects and periods. 2 Values are arithmetic means.

P1194 Table II Summary of Pharmacodynamic Parameters: Serum Neopterin Corrected with Baseline EAuc Emax t '-max 2 (nmoles x (nmoles / L) (h) h / L) AVONEX ™ 693 9.5 44.9 Rebif® 481 6.9 50.3 Relation: AVONEX ™ and Rebif13 144% 138% I.C. 90% 131-159% 123-156% Value p < 0.001 < 0.001 1 The values are means of least squares transformed in the inverse of the logarithmic scale, each one based on the analysis of variance with adjustments for subjects and periods. The p values are based on the comparison of the two products estimated from this model. 2 Values are arithmetic means.

Table III. Summary of demographic data Route of Administration Subcutaneous intramuscular intramuscular parameter Age (years) 27.9 (± 6.7) 27.6 (+ 3.7) 31.5 (± 8.4) Height (m) 171.5 '(+ 10.3) 168.6 (± 8.6) 166.1 (± 4.8) Weight (kg) 68.4 (± 10.6) 63.1 (± 8.9) 65.9 (± 6.9) The values are means (± d.s.) P1194 Table IV. Summary of pharmacokinetic and pharmacodynamic parameters Route of Administration intravenous intramuscular parameter subcutaneous ax (U / mL) 262 (± 45) 44 (± 4) 20 (± 7) AUC (h.U / mL) 537 (± 40) 1352 (± 111) 478 (± 119) Elimination Tl / 2 4.0 (± 1.6) 10.0 (± 0.8) 8.6 (± 1.0) (h) Distribution Tl / 2 0.07 (± 0.02) (h) Depuration 334 (± 18) (mL / h / kg) Vd (L) 61. 6 (± 16 .7) ß2-microglobulin 559 (± 70) 796 (± 58) 628 (± 39) : Emax (μg / L) Neopterin: E, ^ - 16.0 (± 2.0) 12.4 (± 1.5) (nmol / L) The values are means (± s.e.m.).

P1194

Claims (9)

1. CLAIMS: 1. A liquid composition comprising a buffer with pH of about 7.2, recombinant beta-interferon and 15 mg / ml of human serum albumin, wherein the composition is characterized by having a reinforced pharmacokinetic profile compared to the pharmacokinetic profile of a liquid beta-interferon composition with a pH of about 5.0 containing 9 mg / ml of human serum albumin, the reinforced pharmacokinetic profile defined in Table I, and wherein the composition is further characterized by having the reinforced pharmacodynamic profile of a composition beta interferon liquid with pH of about 5.0 containing 9 mg / ml human serum, the reinforced pharmacodynamic profile defined in Table II.
2. 2. The composition according to claim 1, wherein the composition is contained within a container.
3. 3. The composition according to claim 2, wherein the container is a syringe.
4. 4. A liquid pharmaceutical composition contained in a storage container, the liquid suitable for parenteral administration to mammals and which in essence consists of an effective amount of beta-interferon, a buffer that maintains pH at P1194 approximately 7.2 and human serum albumin, wherein the beta-interferon has been subjected to prior lyophilization and wherein the composition is characterized by having a reinforced pharmacokinetic profile compared to the pharmacokinetic profile of a liquid beta-interferon composition with pH 5.0 containing 9 mg / ml of human serum albumin, the reinforced pharmacokinetic profile defined in Table I, and wherein in addition the composition is characterized by having a reinforced pharmacodynamic profile compared to a pharmacodynamic profile of a liquid beta-interferon composition of about 5.0 containing 9 mg / ml of human serum, the reinforced pharmacodynamic profile defined in Table II.
5. 5. The liquid composition according to claim 4, wherein the storage container is a syringe.
6. 6. The liquid composition according to claim 4, which is sterile.
7. 7. The liquid composition according to claim 4, wherein the beta-interferon is human recombinant beta-interferon.
8. 8. A device for parenteral administration of a liquid interferon formulation, wherein the equipment comprises: a) a container containing the liquid formulation according to claim 4 and b) instructions for P1194 the use of it. The equipment according to claim 8, further comprising: c) a swab with alcohol; d) a needle; and at least one adhesive bandage. P1194