Bioequivalence of a Liquid Formulation of Alpha₁-Proteinase Inhibitor Compared with Prolastin®-C (Lyophilized Alpha₁-PI) in Alpha₁-Antitrypsin Deficiency

ABSTRACT

This study evaluated the bioequivalence, safety, and immunogenicity of a new liquid formulation of human plasma–derived alpha₁-proteinase inhibitor, Liquid Alpha₁-PI, compared with the Lyophilized Alpha₁-PI formulation (Prolastin®-C), for augmentation therapy in patients with alpha₁-antitrypsin deficiency (AATD). In this double-blind, randomized, 20-week crossover study, 32 subjects with AATD were randomized to receive 8 weekly infusions of 60 mg/kg of Liquid Alpha₁-PI or Lyophilized Alpha₁-PI. Serial blood samples were drawn for 7 days after the last dose followed by 8 weeks of the alternative treatment. The primary endpoint was bioequivalence at steady state, as measured by area under the concentration versus time curve from 0 to 7 days (AUC_0–7 days) postdose using an antigenic content assay. Bioequivalence was defined as 90% confidence interval (CI) for the ratio of the geometric least squares (LS) mean of AUC_0–7 days for both products within the limits of 0.80 and 1.25. Safety and immunogenicity were assessed. Mean alpha₁-PI concentration versus time curves for both formulations were superimposable. Mean AUC_0–7 days was 20 320 versus 19 838 mg × h/dl for Liquid Alpha₁-PI and Lyophilized Alpha₁-PI, respectively. The LS mean ratio of AUC_0–7 days (90% CI) for Liquid Alpha₁-PI versus Lyophilized Alpha₁-PI was 1.05 (1.03–1.08), indicating bioequivalence. Liquid Alpha₁-PI was well tolerated and adverse events were consistent with Lyophilized Alpha₁-PI. Immunogenicity to either product was not detected. In conclusion, Liquid Alpha₁-PI is bioequivalent to Lyophilized Alpha₁-PI, with a similar safety profile. The liquid formulation would eliminate the need for reconstitution and shorten preparation time for patients receiving augmentation therapy for AATD.

KEYWORDS:

• Antigenic content assay
• augmentation therapy
• clinical trial
• functional activity assay
• pharmacokinetics

Introduction

Alpha₁-antitrypsin deficiency (AATD) is an autosomal codominant genetic disorder characterized by low serum levels of alpha₁-protease inhibitor (alpha₁-PI; historically called alpha₁-antitrypsin). AATD manifests clinically in adults as chronic obstructive pulmonary disease (COPD) and liver cirrhosis (1–3). Evidence-based guidelines recommend augmentation therapy with alpha₁-PI in subjects with AATD (excluding PiMZ phenotype) with evidence of COPD to bolster the protective protease inhibitory shield and slow emphysema progression (4).

Augmentation therapy is available in the United States as an approved therapy for AATD since 1987. The first lyophilized formulation of alpha₁-PI, Prolastin®, was further modified with additional purification, filtration, and production steps with viral clearance capacity and rebranded as Prolastin®-C (Grifols Therapeutics Inc., Research Triangle Park, NC, USA) (5). Liquid Alpha₁-PI is a new alanine-stabilized, liquid formulation of human plasma–derived alpha₁-PI, developed as an alternate dosage form of Lyophilized Alpha₁-PI that does not require reconstitution during its preparation for use; thus, the preparation time is shortened. This study evaluated the bioequivalence, safety, and immunogenicity of Liquid Alpha₁-PI relative to Lyophilized Alpha₁-PI in subjects with AATD.

Material and Methods

Subject selection

Eligible subjects had a diagnosis of congenital AATD and were ≥18 and ≤70 years of age, with documented allelic combination of ZZ, SZ, Z(null), (null)(null), S(null), or other “at-risk” alleles, and a serum alpha₁-PI concentration <11 µM. Postbronchodilator forced expiratory volume in 1 second (FEV₁) was required to be ≥30% and <80% of predicted and FEV₁/forced vital capacity <70% (Global Initiative for Chronic Obstructive Lung Disease stage II or III) (6). Prior or^{Figure 1} current alpha₁-PI augmentation therapy was permitted, provided the treatment was discontinued at the baseline visit (week 1).

Key exclusion criteria included a moderate or severe pulmonary exacerbation during the 4 weeks before baseline; history of lung or liver transplant; lung surgery during the previous 2 years; confirmed liver cirrhosis; severe concomitant disease (e.g., congestive heart failure, clinically significant pulmonary fibrosis, malignant disease [with the exception of skin cancers other than melanoma], history of acute hypersensitivity pneumonitis reaction, or current chronic hypersensitivity pneumonitis); known previous infection or clinical signs and symptoms consistent with current hepatitis A virus, hepatitis B virus, hepatitis C virus, or human immunodeficiency virus infection; smoking within the past 6 months (or positive urine cotinine test); participated in another investigational drug study within 1 month prior to the week 1 (baseline) visit; use of systemic or aerosolized antibiotics for exacerbation within 4 weeks of study start; and use of steroids above a stable dose equivalent to 5 mg/day prednisone within 4 weeks of study start (permissible stable doses could be maintained at the same dose throughout the study); known selective or severe immunoglobulin A deficiency.

The study was conducted in compliance with the principles of the Declaration of Helsinki and the International Conference on Harmonization Good Clinical Practice guidelines. In addition, all local regulatory requirements were followed. The protocol was approved by the Institutional Review Board or Ethics Committee at each center (www.ClinicalTrials.gov NCT02282527). All subjects signed an informed consent statement.

Objectives

The pharmacokinetic primary objective was to demonstrate the bioequivalence of 60 mg/kg of Liquid Alpha₁-PI compared with 60 mg/kg of Lyophilized Alpha₁-PI, both at approximate steady state after eight weekly infusions, as measured by area under the concentration versus time curve from 0 to 7 days (AUC_0–7 days) of alpha₁-PI using an antigenic content assay. In addition, AUC_0–7 days was assessed using a functional activity assay and an adjusted AUC_0–7 days was calculated by subtracting estimated endogenous levels of alpha₁-PI. Safety objectives included evaluation and comparison of adverse events (AE), COPD exacerbations, and immunogenicity of the two investigational treatments when administered for eight weeks.

Study design

This study had a multicenter, randomized, double-blind crossover design (Figure 1). The screening and baseline period included chemistry and hematology labs, inclusion/exclusion criteria check, virus safety testing, physical examination, pulmonary function testing, and reporting of genotyping and phenotyping results for potentially eligible subjects whose AATD genotype had not previously been documented. Eligible subjects were randomized 1:1 by computer-generated schedule to one of two treatment sequences to either receive weekly infusions of 60 mg/kg Liquid Alpha₁-PI (supplied in 20-ml glass vials as a sterile, stable solution containing approximately 1 g of functionally active alpha₁-PI per vial, delivered at a rate of 0.08 ml/kg/min) for 8 weeks or weekly infusions of 60 mg/kg Lyophilized Alpha₁-PI (supplied as a lyophilized powder in 50-ml glass vials that was reconstituted with 20-ml sterile water; infused dose contained approximately 1 g of active alpha₁-PI per vial) for 8 weeks followed by weekly infusions of the alternate treatment for another 8 weeks, according to the crossover study design (Figure 1). The off-treatment follow-up period (weeks 17 to 20) was to provide safety information off treatment, collect pharmacokinetic samples, and provide a measure of endogenous concentration (C_week20) of alpha₁-PI.

Figure 1. Schematic of the study design. Liquid Alpha₁-PI: human plasma–derived alpha₁-proteinase inhibitor; IV: intravenous; PI: proteinase inhibitor; PK: pharmacokinetic.

Following preparation by an unblinded pharmacist, prepared intravenous infusion bags for investigational treatments were indistinguishable to blinded study staff and subjects, and were marked only with sufficient identification for correct administration and were covered with nontransparent blinding covers.

Data collection

Serum concentration of alpha₁-PI was measured in trough (preinfusion) and serial blood samples from all subjects for the purpose of pharmacokinetic analysis (Figure 1). Trough samples during the treatment periods were drawn prior to infusions at weeks 6–9 and weeks 14–17. For AUC determination, a total of 12 serial blood samples were collected, starting at the end of the treatment period (week 8 or 16) and extending over 7 days after the previous treatment period infusion (week 9 or 17). Blood samples were collected immediately prior to the infusion, immediately after a saline flush when the infusion was completed, and at the following times from completion of the infusion: 0.25, 0.5, 1, 2, 4, 8 hours, and then days 1 (±4 hours), 2 (±4 hours), 5 (±1 day), and 7 (±1 day).

A blood sample for alpha₁-PI concentration was also drawn at the week 20 visit, at the end of the 4-week off-treatment follow-up period, for the purpose of calculating the adjusted AUC_0–7 days values.

AEs/treatment-emergent AEs, serious adverse events (SAEs), and adverse drug reactions were recorded. Treatment-emergent AEs were defined as any AE during the study that began on or after the date and time of first dose of investigational product. A suspected adverse drug reaction included any AEs with investigator's causality assessment of “definitive,” “probable,” “possible,” or “doubtful/unlikely”. In addition, vital signs and COPD exacerbations were recorded (7). As COPD exacerbations are part of the natural history of AATD, they were not reported as AEs unless they met SAE criteria. Additionally, immunogenicity assessment and parvovirus B19 safety testing were carried out (at least at baseline and at the end of each treatment period).

Laboratory assessments and statistical analyses

Blood concentration of alpha₁-PI was assessed by two assays: antigenic content assay and functional activity (potency) assay. Antigenic content assay measures both functionally active and inactive forms of the alpha₁-PI protein and was the primary assay. The concentration of alpha₁-PI was measured by immunonephelometry using a standard control that is traceable to the international reference standard CRM 470. Functional activity assay measures only the concentration of protein capable of inhibiting neutrophil elastase and was supportive to the primary assay. To quantify alpha₁-PI inhibitory activity, a chromogenic elastase substrate was exposed to a patient sample and residual elastase activity was measured using a standard control. Both assays were validated according to current regulatory guidelines (5).

The primary pharmacokinetic endpoint was measured by the AUC_0–7 days postdose (i.e., at steady state) using the antigenic content assay. Bioequivalence was defined as 90% confidence interval (CI) for the geometric least squares mean (LSM) ratio of AUC_0–7 days for the two products falling between the limit of 0.80 and 1.25 (8). Natural log-transformed AUC_0–7 days values were analyzed by the mixed-effect model, including treatment, treatment sequence, and study period as fixed effects and subject (nested within sequence) as a random effect.

Exploratory analysis of the geometric LSM ratio of AUC_0–7 days adjusted by subtraction of C_week20 (adjusted AUC_0–7 days) and the geometric LSM ratio of maximum concentration (C_max) were also performed for both antigenic content assay and functional activity assay. Time to reach C_max, time to descend to half of C_max (t_½), rate of clearance, and mean trough concentration at steady state were also descriptively summarized.

Pharmacokinetic data from a previous study comparing Prolastin®-C with Prolastin® indicated that the standard deviation in log scale for AUC_0–7 days is 0.174 (5). Assuming a standard deviation of 0.174, a sample size of 12 subjects (6 subjects per treatment sequence) was needed to demonstrate bioequivalence with 90% power. Considering additional assessment of safety and immunogenicity, a sample size of 30 subjects (15 subjects per treatment sequence) was planned for the study.

Results

Subjects

Subjects were enrolled^{Table 1} at six investigative centers in the United States. Of the 38 subjects screened, six did not meet the entry criteria (four spirometric, one specific allelic combination, and one informed consent) and 32 were randomized and dosed (intent-to-treat and safety populations; Figure 2). One subject in the Liquid Alpha₁-PI/Lyophilized Alpha₁-PI treatment sequence discontinued after week 1 owing to lack of home health care service and was excluded from the pharmacokinetic analysis. One patient in the Lyophilized Alpha₁-PI/Liquid Alpha₁-PI treatment sequence was lost to follow-up after completing both treatment periods and was included in the pharmacokinetic analysis. Thus, 31 subjects were included in the pharmacokinetic analysis.

Figure 2. Subject disposition – CONSORT diagram. Liquid Alpha₁-PI: human plasma–derived alpha₁-proteinase inhibitor; ITT: intent-to-treat; PI: proteinase inhibitor; PK: pharmacokinetic.

Demographics and baseline characteristics were similar across the two treatment sequences (Table 1). All subjects were white (genotype PI^*ZZ, n = 31; PI^*SZ, n = 1). Alpha₁-PI blood levels at time of diagnosis ranged from 2.7 µM to 9.2 µM. Four patients were naïve to alpha₁-PI augmentation therapy, whereas 28 had received prior therapy with at least one drug (Prolastin®-C, 20; Prolastin®, 5; Zemaira®, 3; Glassia®, 1). Baseline postbronchodilator FEV₁ percentage predicted values ranged from 30.0% to 79.0%.

Table 1. Demographics and AATD history.

	Treatment Sequence^*		Total in ITT Population
Characteristic	Liquid Alpha1-PI/ Lyophilized Alpha1-PI (n = 16)	Lyophilized Alpha1-PI/ Liquid Alpha1-PI (n = 16)	(N = 32)
Sex, n (%)
Female	8 (50)	6 (38)	14 (44)
Male	8 (50)	10 (63)	18 (56)
Age, mean (SD), y	60.7 (7.94)	63.1 (5.93)	61.9 (7.00)
Race, white, n (%)	16 (100)	16 (100)	32 (100)
Ethnicity, n (%)
Hispanic or Latino	0	1 (6)	1 (3)
Not Hispanic or Latino	16 (100)	15 (94)	31 (97)
Weight, mean (SD), kg	83.7 (21.06)	78.6 (16.78)	81.2 (18.91)
Time since diagnosis, mean (SD), y	9.9 (5.89)	14.2 (8.50)	12.1 (7.51)
Alpha1-PI level at time of diagnosis, mean (SD), µM	4.6 (0.99)	5.0 (1.58)	4.8 (1.31)
Genotype, n (%)
PI^*ZZ	16 (100)	15 (94)	31 (97)
PI^*SZ	0	1 (6)	1 (3)
Naïve status, n (%)
Naïve	1 (6)	3 (19)	4 (13)
Non-naïve	15 (94)	13 (81)	28 (88)
Baseline alpha1-PI concentration
Naïve
mean (SD), µM,	7.0 (—)	4.8 (0.90)	5.4 (1.33)
[mg/dl (SD)]	[31 (—)]	[21 (4.0)]	[24 (5.9)]
Non-naïve
mean (SD), µM,	16.3 (6.40)	15.6 (4.86)	15.8 (5.61)
[mg/dl (SD)]	[72 (28.3)]^†	[69 (21.5)]	[70 (24.8)]^†
% predicted FEV1, mean (SD)	54.2 (15.26)	53.4 (14.26)	53.8 (14.53)

FEV₁: forced expiratory volume in 1 second; ITT: intent-to-treat; PI: proteinase inhibitor; SD: standard deviation.

^* The order of the sequence of treatments per randomization is indicated in the column headers;

^† Baseline alpha₁-PI concentration was not available for 1 subject owing to sample instability.

Pharmacokinetic parameters

Mean alpha₁-PI concentrations versus time curves at steady state after 8 weeks of weekly 60 mg/kg intravenous doses of Liquid Alpha₁-PI were essentially indistinguishable from those following a weekly 60 mg/kg intravenous dose of Lyophilized Alpha₁-PI using the antigenic content assay (Figure 3A) or the functional activity assay (Figure 3B). Mean AUC_0–7 days was 20 320 versus 19 838 mg × h/dl for Liquid Alpha₁-PI and Lyophilized Alpha₁-PI, respectively (Table 2) and the geometric LSM ratio for AUC_0–7 days with Liquid Alpha₁-PI versus Lyophilized Alpha₁-PI was 1.05 with 90% CI of 1.03 to 1.08 (Figure 4).

Figure 3. Mean alpha₁-PI concentration (± SD) using the (A) antigenic content assay and (B) functional activity assay at week 8 (steady state). Liquid Alpha₁-PI: human plasma–derived alpha₁-proteinase inhibitor; PI = proteinase inhibitor; SD: standard deviation.

Table 2. Summary of pharmacokinetic parameters.

	Antigenic Content Assay		Functional Activity Assay
	Liquid Alpha1-PI (N = 31)	Lyophilized Alpha1-PI (N = 31)	Liquid Alpha1-PI (N = 31)	Lyophilized Alpha1-PI (N = 31)
AUC0–7 days, mean (%CV), mg × h/dl	20 320 (11.3)	19 838 (12.7)	17 116 (16.8)	16 850 (16.3)
Adjusted^* AUC0–7 days, mean (%CV), mg^*h/dL	15 349 (14.2)	14 767 (17.9)	13 910 (17.0)	13 685 (17.9)
Cmax (mg/dl), mean (%CV)	254 (15.3)	249 (20.0)	208 (14.2)	204 (17.0)
tmax, median, h (range)	0.64 (0.28–2.25)	0.69 (0.28–4.33)	0.52 (0.28–2.25)	0.74 (0.25–1.42)
t½, mean (%CV), h	156.39 (18.0)	164.10 (21.1)	126.57 (20.7)	126.82 (26.7)
CL, mean (%CV), dL/kg/h	0.0040 (14.0)	0.0042 (22.1)	0.0045 (20.8)	0.0045 (17.4)

AUC_0–7 days: area under the concentration versus time curve from 0 to 7 days; C_max: maximum concentration; CL: clearance; CV: coefficient of variation; PI: proteinase inhibitor; t_½: half-life; t_max: time to reach C_max.

^* Adjusted AUC_0–7 days is calculated using the concentrations with subtraction by the endogenous concentration measured at week 20.

Figure 4. Geometric least squares mean ratio (90% confidence interval) for Liquid Alpha₁-PI versus Lyophilized Alpha₁-PI (antigenic content assay). AUC_0–7 days: area under the concentration versus time curve from 0 to 7 days; C_max: maximum concentration; Liquid Alpha₁-PI: human plasma–derived alpha₁-proteinase inhibitor.

Mean alpha₁-PI concentration at week 20 (4 weeks after the last study drug infusion) was 29.9 mg/dl using the antigenic content assay (Figure 5). The geometric LSM ratio for the adjusted AUC_0–7 days for Liquid Alpha₁-PI versus Lyophilized Alpha₁-PI was 1.07 with 90% CI of 1.03 to 1.11, also indicating bioequivalence between the two products (Figure 4). The geometric LSM ratio of Liquid Alpha₁-PI/Lyophilized Alpha₁-PI for C_max also falls within the range of bioequivalence (Figure 4).

Figure 5. Mean (± SD) steady-state trough Alpha₁-PI concentrations (antigenic content). Liquid Alpha₁-PI: human plasma–derived alpha₁-proteinase inhibitor; PI: proteinase inhibitor; SD: standard deviation.

All of the alpha₁-PI concentration–based pharmacokinetic parameters were larger with the antigenic content assay compared with the functional assay, but both showed comparable similarity between the two study treatments (Table 2). From exploratory statistical analyses of functional assay data, the 90% CI corresponding to geometric LSM ratios for AUC_0–7 days (1.007–1.072), adjusted AUC_0–7 days (0.998–1.089), and C_max (0.997–1.075) were also bound within the predefined range to indicate bioequivalence. With the antigenic content assay, once C_max was achieved (≈250 mg/dl) after approximately 40 minutes, alpha₁-PI concentrations declined in a multi-exponential manner (mean clearance ≈ 0.0040 dl/kg/h, t_½ ≈ 160 hours). The intersubject variability (% coefficient of variation) in AUC_0–7 days, adjusted AUC_0–7 days, C_max, t_½, and clearance of alpha₁ PI was ≤26.7% for each treatment using either assay, indicating general consistency in the distribution and elimination of alpha₁-PI between subjects and treatments.

Steady state for trough alpha₁-PI concentrations using the antigenic content assay was reached by the start of the sixth weekly infusion at week 6/14 (Figure 5). The mean trough of alpha₁-PI over the 4-week period at steady state was above the historical target of 11 µM in all subjects of this study at each time point, with the exception of one subject that measured 10.62 µM at week 8 (while receiving Lyophilized Alpha₁-PI). Steady-state trough alpha₁-PI concentrations using the functional assay were similar between treatments.

Safety

Overall, 32 subjects received at least one dose of Liquid Alpha₁-PI and 31 subjects received at least one dose of Lyophilized Alpha₁-PI. Mean duration of exposure was 7.92 and 7.90 weeks with Liquid Alpha₁-PI and Lyophilized Alpha₁-PI, respectively. The mean number of infusions per subject was 7.9 for both treatments, which represented overall 252 infusions with Liquid Alpha₁-PI and 245 infusions with Lyophilized Alpha₁-PI.

No deaths occurred during the study and no subject experienced an SAE while receiving Liquid Alpha₁-PI. One subject had an SAE (moderate infective exacerbation of COPD) during the follow-up period after receiving Lyophilized Alpha₁-PI that was not considered to be related to investigational product. No subjects withdrew from the study owing to an AE.

Treatment-emergent AEs were experienced by 59% of subjects when treated with Liquid Alpha₁-PI and 42% of subjects when treated with Lyophilized Alpha₁-PI (Table 3). Overall, the AE profile for both study treatments showed no consistent treatment-related pattern for any particular type of event and none occurred in more than two subjects per treatment (Table 3). All treatment-emergent AEs in the study were considered by the investigator to be of mild or moderate intensity, except one case of severe back pain (worsened back pain due to motorcycle riding) with Lyophilized Alpha₁-PI. Adverse drug reactions suspected by the investigator to be related to treatment included fatigue, diarrhea, contusion, decrease in platelet count, and insomnia during infusions with Liquid Alpha₁-PI and abnormal hepatic function and dizziness with Lyophilized Alpha₁-PI. No AEs with either treatment were considered by the investigator as definitely related to treatment.

Table 3. Summary of safety.

	Liquid Alpha1-PI (N = 32) n (%)	Lyophilized Alpha1-PI (N = 31) n (%)
Subjects with TEAE(s)	19 (59)	13 (42)
Total number of TEAEs	31	25
Rate of TEAEs per infusion	0.123	0.102
TEAEs in ≥5% of subjects/group
Diarrhea	2 (6)	1 (3)
Dermatitis contact	2 (6)	0
Fatigue	2 (6)	0
Pyrexia	2 (6)	0
Nasopharyngitis	1 (3)	2 (6)
Number of SAEs	0	1^*
Subjects with suspected ADRs	5 (16)	2 (6)
Total number of suspected ADRs events	7	3
Subjects with COPD exacerbations	12 (38)	9 (29)
Total number of COPD exacerbations events	13	10

ADR: adverse drug reaction; COPD: chronic obstructive pulmonary disease; PI: proteinase inhibitor; SAE: serious adverse event; TEAE: treatment-emergent adverse event.

No TEAE was reported by >2 subjects within a treatment group.

^* Moderate infective exacerbation of COPD.

Exacerbations of COPD occurred in 18 subjects, with a total of 23 events. The distribution of occurrences was similar across treatments (Table 3). Of these, four COPD exacerbations occurred during the follow-up period: three after Liquid Alpha₁-PI treatment and one after Lyophilized Alpha₁-PI treatment. All COPD exacerbations were graded as mild or moderate, though one was reported as an SAE (discussed above) owing to hospitalization. No treatment-emergent COPD exacerbation resulted in discontinuation or was considered to be related to the study treatment.

No changes from baseline of clinical concern were noted at any time point for any laboratory measurement, vital sign, physical exam, or pulmonary function test parameter.

Discussion

This trial compared pharmacokinetic parameters and the safety profile of Liquid Alpha₁-PI with Lyophilized Alpha₁-PI in subjects with AATD. The trial achieved the primary pharmacokinetic objective by demonstrating bioequivalence of the AUC_0–7 days at steady state for the two treatments following weekly infusions over 8 weeks. Safety outcomes and other pharmacokinetic parameters were also comparable between the two treatments. Importantly, the 90% CI for the geometric LSM ratio, Liquid Alpha₁-PI/Lyophilized Alpha₁-PI, of the AUC_0–7 days fell within the range that demonstrated that a weekly infusion of 60 mg/kg Liquid Alpha₁-PI produces an AUC of serum alpha₁-PI that is bioequivalent to that produced by a weekly infusion of 60 mg/kg Lyophilized Alpha₁-PI. This is supported by the mean alpha₁-PI concentration versus time curves of the two treatments that were essentially superimposable. Bioequivalence was also confirmed by AUC_0–7 days adjusted for the week 20 alpha₁-PI concentrations (representative of endogenous alpha₁-PI) and C_max based on both antigenic content assay and functional activity assay.

The mean C_week20 (29.9 mg/dl) is consistent with the known endogenous alpha₁-PI concentrations in AATD patients with the PI^*ZZ genotype (10–30 mg/dl) (9,10), and with baseline alpha₁-PI levels of four naïve AATD patients in this study (mean concentration of 24 mg/dl) as well as four patients from the SPARK study (mean ≈ 30 mg/dl) (11). Trough concentrations determined over a 4-week period in each treatment period indicate that an approximate steady-state condition was achieved by the start of the sixth weekly infusion for both Liquid Alpha₁-PI and Lyophilized Alpha₁-PI. This confirms that the pharmacokinetic parameters of alpha₁-PI after 8 weeks of treatment in this study were determined under an approximately steady-state condition for both Liquid Alpha₁-PI and Lyophilized Alpha₁-PI. Furthermore, trough values of alpha₁-PI were above the historical “protective” level of 11 µM, as has been previously reported with weekly doses of Lyophilized Alpha₁-PI in the ChAMP and SPARK studies (5,11).

This study used two assays to measure alpha₁-PI levels. Concentrations measured by the functional activity assay were slightly lower than those using the antigenic content assay, and so pharmacokinetic parameter results were slightly different depending on the assay used. However, Liquid Alpha₁-PI and Lyophilized Alpha₁-PI were comparable across pharmacokinetic parameters whether using the functional activity assay or the antigenic content assay.

In addition, the key pharmacokinetic parameters in this study are similar to those observed in the ChAMP study and the SPARK study—two pharmacokinetic studies that also evaluated the pharmacokinetics of Lyophilized Alpha₁-PI (5,11). The mean AUC_0–7 days values (antigenic content) for Liquid Alpha₁-PI and Lyophilized Alpha₁-PI were 20 320 and 19 838 mg × h/dL, respectively, which are consistent with those for Lyophilized Alpha₁-PI observed in ChAMP (19 010 mg × h/dL (5)) and SPARK (20 360 mg × h/dL) (11)). The mean C_max values for Liquid Alpha₁-PI and Lyophilized Alpha₁-PI were 254 and 249 mg/dL, respectively, for the antigenic content assay, which are consistent with those for Lyophilized Alpha₁-PI observed in the two previous studies: 233 mg/dL (5) and 247 mg/dL (11). In addition, the results obtained with Lyophilized Alpha₁-PI in this study based on the functional activity assay are also consistent with those reported in ChAMP (5) for Lyophilized Alpha₁-PI (eg, C_max of 208 mg/dL in this study vs C_max of 180 mg/dL in ChAMP).

The AE profile of Liquid Alpha₁-PI was consistent with Lyophilized Alpha₁-PI and showed no consistent treatment-related pattern for any particular type of event. Most AEs were infrequent and few were reported in more than 1 subject. All AEs were mild or moderate in intensity, with the exception of one SAE considered unrelated to study treatment that occurred during the off-treatment follow-up period after Lyophilized Alpha₁-PI treatment. Overall, the frequencies of AEs and COPD exacerbations were similar between treatments. However, because this study was not powered nor long enough to assess the effect of the treatments on exacerbation rates, no conclusions can be made regarding efficacy.

Immunogenicity was not detected in any subject in the trial. There were no instances of Parvovirus seroconversions or any clinical indication of seroconversions against any other blood-borne agent.

In the clinic, practical differences between Liquid Alpha₁-PI and Lyophilized Alpha₁-PI should be considered. First, the Liquid Alpha₁-PI does not require reconstitution, whereas the Lyophilized Alpha₁-PI is a lyophilized powder that requires sterile water; thus, the use of Liquid Alpha₁-PI may save preparation time over Lyophilized Alpha₁-PI. Second, the two versions provide the same concentrated formula and volume per dose. Third, the Liquid Alpha₁-PI requires refrigeration for long-term storage (up to the expiration date) but can be kept at room temperature for up to 1 month, whereas Lyophilized Alpha₁-PI, in the lyophilized form, can be stored at room temperature for the period indicated by the expiration date. Once reconstituted, Lyophilized Alpha₁-PI should be administered within 3 hours. Similarly, Liquid Alpha1-PI should be used within 3 hours of withdrawing it from the vial and adding it to the empty intravenous bag for administration.

Conclusions

Liquid Alpha₁-PI 60 mg/kg/wk was demonstrated as bioequivalent to Lyophilized Alpha₁-PI 60 mg/kg/wk with a comparable tolerability profile. Liquid Alpha₁-PI treatment was well tolerated with no reported SAEs. Liquid Alpha₁-PI may provide enhanced convenience to patients receiving augmentation therapy for AATD.

Declaration of Interest

AF Barker: Clinical research funding provided by Grifols Therapeutics Inc.

MA Campos: Research grants from the Alpha-1 Foundation and CSL Behring; participated in clinical trials sponsored by Baxalta, CSL Behring, and Grifols Therapeutics Inc.; participated in advisory boards for CSL Behring and Grifols Therapeutics Inc.

ML Brantly: Research grants from Grifols Therapeutics Inc.; owner of GeneAidyx.

JM Stocks: Research grants from Grifols Therapeutics Inc. and Kamada.

RA Sandhaus: Research grants from CSL Behring and Grifols Therapeutics Inc.; trains speakers on their speaker bureau for Grifols Therapeutics Inc.; advisory boards for Shire (Baxalta).

D Lee: Speaker for Boehringer Ingleheim, Genentech, Grifols Therapeutics Inc., and Otsuka; participated in clinical trials sponsored by Grifols Therapeutics Inc.

K Steinmann, J Lin and S Sorrells: employees and stockholders of Grifols Therapeutics Inc.

Authors' Contributions

AFB, MAC, MLB, JMS, RAS, DL, KS, JL, and SS contributed to the study design and implementation and provided meaningful input to this manuscript. JL provided statistical analyses for this study.

Acknowledgments

The authors would like to thank the study coordinators for their dedication and care of the study participants, and the participants of this study for giving of their valuable time. Writing and editorial assistance was provided to the authors by David Macari, PhD, and Susan Sutch, PharmD, CMPP, on behalf of Evidence Scientific Solutions, Philadelphia, Pennsylvania, USA, and funded by Grifols Inc., Research Triangle Park, North Carolina, USA.

Funding

Grifols Therapeutics Inc. (Research Triangle Park, NC, USA) funded this study.