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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5400_Библиотеки_им_академика_М_И_Перельмана
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This review will focus on the advances in knowledge and frontiers for dry
powder pulmonary delivery of monoclonal antibodies (mAbs). Some
background required to support the review of this ield will include tenets of
pulmonary delivery, processing of pharmaceutical powders for inhalation,
and general formulation knowledge for biologics.
Pulmonary delivery of aerosolized active pharmaceutical ingredients
(APIs) has been explored in various forms for centuries. Despite its
widespread adoption, challenges remain for pulmonary delivery in the areas
of ease of device use and reliable dosing, both impacting patient compliance
[1]. The development of metered dose inhalers (MDIs) in the 1950s for
delivery of atomized solutions created a paradigm shift in the ield since they
were small, inexpensive, and could be used at home [2]. However, incorrect
MDI operations resulting in improper dosing by patients led to the rise of dry
powder inhalers (DPIs) as an alternative technology in the 1980s [3]. Unlike
MDIs, DPIs utilize aerosolized powders and rely on patient inhalation for
delivery, eliminating the complex synchronization of patient breath and
device actuation associated with MDIs. An additional advantage of DPIs
compared with MDIs is that they are propellant-free.
Pulmonary delivery of proteins and nebulizer delivery of mAb are
discussed extensively in other chapters of this book. Nebulizers have been
primarily used as the standard pulmonary delivery device for mAb
therapeutics. In recent years, delivery by DPI has been studied as a way to
reduce the treatment burden on the patient while enabling product storage at
ambient temperatures instead of frozen solutions. This review will discuss
the advantages and challenges of dry powder pulmonary delivery of mAbs.
Regulatoryrequirementsforinhaledproducts
Any DPI product iling (small molecule or biologic) involves meeting
various regulatory quality and eficacy requirements, which differ somewhat
by region. For all DPI products, the drug product and its device are treated as
a combination product. Additional regulations and/or critical quality
attributes (CQAs) apply to inhaled biologic products, including future
pulmonary mAb products. These guidances must be met across agencies such
as the Food & Drug Administration (FDA) and European Medicines Agency
(EMA) to enable worldwide sales of a product [4, 5]. The speciications
generally accepted as the union of these guidances are represented in Table 1.
Note that “Additional tests likely for inhaled mAb” is speculative, as no
inhaled mAb products have yet achieved approved status.
Table1 Quality attributes for inhaled pharmaceutical products [5, 6]
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Quality
attribute
Small
molecule
inhaled
produ cts,US
(FDA)
Small
molecule
inhaled
produ cts,EU
(EMA)
Additional
testslikely
forinhaled
mAb
Commonmethodsfor
testing
Description X X
Visual
Identiication X
Fourier-transform infrared
spectroscopy (FTIR), UV-vis
spectroscopy, retention
time (by liquid
chromatography or gel
electrophoresis)
Assay X X
UV-vis spectroscopy,
retention time (liquid
chromatography or gel
electrophoresis)
Biological
activity
X API speciic (surface
plasmon resonance, ELISA
or cell-based assay)
Impurities and
degradation
products
X X
Liquid chromatography or
gel electrophoresis
Aggregation
X Size-exclusion
chromatography (SEC)
and/or dynamic light
scattering
Aerodynamic
particle size
distribution
X X
Next Generation Impactor,
Fast-Screening Impactor,
breath actuators
Delivered Dose
Uniformity
(DDU)
X X
DDU apparatus USP<601>
Mean delivered
dose
X
Number of
actuations per
container
X
Foreign
particulate
matter
X
Optical, laser diffraction
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Quality
attribute
Small
molecule
inhaled
produ cts,US
(FDA)
Small
molecule
inhaled
produ cts,EU
(EMA)
Additional
testslikely
forinhaled
mAb
Commonmethodsfor
testing
Microbial
limits
X X
Moisture
content
X X
Karl Fischer titration
Net content
weight
X
In the preclinical studies referenced later in this chapter, there is a strong
focus on evaluating the biologic activity, degradation, aggregation and
aerodynamic particle size distribution (APSD) quality attributes. Tests that
are most relevant for later-stage clinical products, such as delivered dose
uniformity, microbial limits, and moisture content, will not be expanded upon
in this chapter.
A primary focus for any inhaled pulmonary product is the eficient
delivery of the API to the target region of the airway. For inhaled powders, the
main critical quality attribute is APSD. The aerodynamic diameter of a
particle is equal to the diameter of a unit-density sphere whose inertial
settling velocity is equivalent [6]. It is measured by cascade impaction (e.g.
Next Generation Impactor (NGI), Anderson Cascade Impactor, or FastScreening Impactor), and provides an assessment of how much active is
delivered to the lung (assuming a 5 micron upper limit for lung delivery).
Delivered dose uniformity, mean dose delivered, and number of actuations
are also key measures to ensure an eficacious dose will reach patients.
Another primary consideration when evaluating an inhalation product
involves the product’s potential immunogenicity. One test toward this risk is
the foreign particulate matter measurement. These particles can come from
excipients or the manufacturing process, but with mAb products there is an
added complication of stability issues resulting in protein aggregation and
potential for increased immunogenicity. The challenges and considerations
associated with protein aggregates in inhaled biologics were recently
highlighted by Ibrahim et al. [7]. Additionally, the microbial testing includes
speciic microbes of interest to inhaled delivery, and these speciications are
an order of magnitude below what is common for oral products as described
in USP <1111>.
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2 ManufacturingProces sOverview
Given the strict requirements for pulmonary delivery of powders, precise
control over the manufacturing process is critical to achieving the desired
aerosolization properties [8, 9]. In this section, we discuss the strategies for
manufacturing any inhalation dry powders with a focus on biologics DPIs and
speciically mAbs.
A common method to make aerosolizable powders of small molecules is
mechanical milling. While this strategy is simple and cost-effective, it can be
challenging due to the generation of electrostatic charges, inconsistent
morphologies, and high surface energies that promote instability [10]. For
proteins, milling is not a feasible particle engineering technique: mechanical
and thermal forces from milling can denature proteins and impair their
activity, and require API supply in the solid state [11]. Instead, particle
engineering to deliver biotherapeutics to the lung is better accomplished via
drying processes that start from a liquid feedstock. This section will
introduce many drying techniques while focusing on spray drying, sprayfreeze drying, and thin ilm freezing, which were also addressed in more
detail in a recent review article [12].
2.1 SprayDrying
Developed originally in the early twentieth century for the production of
powdered milk products in the dairy industry, spray drying is a scalable
manufacturing technique that has become a common unit operation in many
industries [13]. A schematic of the spray drying process is shown in Fig. 1. In a
pharmaceutical spray drying process, API and excipients are co-dissolved in a
volatile solvent. The liquid feed is pumped into a drying chamber through an
atomizer, where small liquid droplets are formed. The droplets come into
contact with heated drying gas, which rapidly removes solvent from the
droplets. In this way, solid particles are formed and subsequently collected
from the outlet gas stream using cyclonic separators, ilter banks, or a
combination thereof. Notably, spray drying has been employed in
manufacturing of at least 24 commercially-approved products [14].
Speciically for particle engineering toward inhalable therapies, the use of
spray drying as a particle engineering tool has been extensively reviewed by
Vehring [6].
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Fig.1 Schematic of the spray drying process
For the manufacture of dry powder inhalation dosage forms, spray drying
is particularly valuable as it accomplishes both the particle engineering and
formulation steps in a single unit operation. The use of spray drying to
manufacture dry powders for pulmonary drug delivery has been reviewed
multiple times over the past 15 years [15–17]. Spray-dried products for
pulmonary delivery include: Tobi Podhaler (tobramycin, DPI), Inbrija
(levadopa, DPI), Exubera (insulin, DPI), Afrezza (insulin, DPI), Cayston
(aztreonam, reconstituted and nebulized), Bronchitol (mannitol, DPI),
Aerovanc (vancomycin, DPI), and Inavir (laninamivir, DPI) [14, 16, 18].
Most of the discussion in this chapter will focus on spray drying as a
manufacturing technique, as it represents the majority of the work in the ield
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of inhaled biologic development. Detailed process and formulation
considerations for spray drying of mAbs are discussed later in this chapter.
2.2 AlternateManufacturingProcesses
Rather than relying on forced, heated drying gas to remove water from the
solution, spray-freeze drying and thin ilm freezing rely on cooled
environments, similar to a lyophilization-based drying step. The spray-freeze
drying process begins with atomization of a solution stream directly into a
cryogenic chamber to freeze the particles. These particles are then
lyophilized to remove the solvent and isolate solid particles [19]. Spray-freeze
drying has the ability to create formulations with particle sizes and densities
amenable to inhaled delivery. A recent review covered the application of this
process to inhaled biotherapeutics in detail [20]. In an early study, Maa et al.
compared spray drying and spray-freeze drying technologies for an anti-IgE
mAb, inding that spray-freeze drying produced larger, low-density particles
with high ine particle fraction [21]. Scale-up to clinical and commercial-scale
equipment is underway for spray-freeze drying of pharmaceutical products
(including an aseptic system, Lyninity, now commercially available), but it is
not yet as readily scalable or energy-eficient as spray drying at this time.
Thin ilm freezing (TFF) is a recently-developed platform to achieve small
respirable particles of delicate molecules such as monoclonal antibodies [22].
In TFF, a liquid protein solution is applied to a cryogenic drum and rapidly
frozen. Next, lyophilization is performed to remove the solvent, forming
brittle matrix particles that can aerosolize upon inhalation. TFF has shown
promise, particularly for particle engineering of proteins that are sensitive to
air-water interfaces during atomization such as lactoferrin [23].
In a variation on the traditional spray drying process, electrostatic spray
drying applies a charge to the liquid droplets as they form within the
atomizer. The charge aids in driving water out of the atomized droplets,
reducing the need for heated atomization gas. Thus, electrostatic atomization
could help reduce water content in a powdered product while potentially
reducing thermal exposure in the droplet. Mutukuri et al. employed
electrostatic spray drying to manufacture inhalable dry powders of
trastuzumab formulated with trehalose, phosphate buffer, and polysorbate 20
[24].
Since there are multiple processes/mechanisms to make inhalable solid
particles, the most appropriate manufacturing process should be selected
with the patient and product/process scalability in mind. Formulation
selection and the resultant particle morphology from commercially-scalable
equipment are likely the strongest driving factors for product performance. In
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the current state, spray drying meets these requirements and will therefore
be the focus of the remainder of this chapter.
3 FormulationofSpray-DriedInhaledM Abs
A challenge common to all dry powder manufacturing strategies is the
structural denaturation of proteins during manufacturing and/or upon
product storage. Many of the challenges associated with dry powder mAbs for
pulmonary delivery also apply to spray-dried mAbs for other applications
such as reconstitution and subsequent injection, so numerous studies on this
topic are instructive and included herein. A recent review covers the role of
stabilizing excipients in biomacromolecule dry powder formulations in great
detail [25]. Excipients for dry powder formulation of biotherapeutics can be
divided into two general categories: stabilizers and surface-active excipients.
Both stabilizers and surface-active excipients have the added constraint that
they must be acceptable for use in the sensitive respiratory tract. A summary
of the current status of common excipients and their functions is provided in
Table 2.
Table2 Common excipients investigated for dry powder antibody formulations
Excipient Suggestedfunctionindry
powdermAbfo rmulation
PrecedenceinFDA-approved
pulmonaryproduct?
Trehalose Stabilizer No
Mannitol Stabilizer Yes
Lactose Stabilizer Yes
Glucose Stabilizer No
Sorbitol Stabilizer No
Cyclodextrins Stabilizer No
Rafinose Stabilizer No
L-leucine Surface active No
Tri-leucine Surface active No
Polysorbate 80 Surface active Yes
Glycine Stabilizer Yes
Sodium lauryl
Sulfate
Surface active Yes
Cysteine Stabilizer No
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Excipient Suggestedfunctionindry
powdermAbfo rmulation
PrecedenceinFDA-approved
pulmonaryproduct?
Phenylalanine Stabilizer No
Arginine Stabilizer No
Polysorbate 20 Surface active No
3.1 StabilizingExcipients
As discussed in the process sections below, mAbs in dry powder inhaled
formulations undergo various stresses depending on their manufacturing
technique, including dehydration, shear from atomization, and air-liquid
interfacial stress for spray drying [26]. Freezing stress is encountered
additionally for spray-freeze drying. On the other hand, thin ilm freezing
eliminates the risk of shear from atomization. In these cases, excipients help
protect antibodies from these processing stresses.
3.1.1 StabilizingSugars
Literature studies of intravenous and nebulized pulmonary formulations of
mAbs focus on dissolving the active in a buffer, typically with a sugar-based
excipient and a surfactant to reduce solution-state aggregation. As with
intravenous (IV) formulations, sugars and sugar alcohols most often serve as
stabilizing excipients for dry powder formulations, where they help in
maintaining the protein’s native conformation via hydrogen bonding/water
replacement and reducing mobility in the solid [27].
Mannitol, lactose, sorbitol, trehalose, rafinose, glucose, and cyclodextrin
are common sugar matrix excipients employed in clinical and preclinical trial
formulations [25, 28–38]. Of these excipients, trehalose has emerged as the
most prevalent stabilizer for dry powder inhalation delivery of
biotherapeutics [39–42]. Trehalose is a non-reducing sugar (i.e., a sugar with
no free ketone or aldehyde group) that is not subject to the Maillard reaction
mechanism of degradation [43]. Trehalose also has a high glass transition
temperature (~120 °C) under dry conditions, leading to good chemical and
physical stability [25]. Materials with glass transition temperatures far above
their storage temperatures will have low molecular mobility, effectively
“locking in” the formulation’s molecular structure, and reducing changes over
time.
Sane et al. compared the processes of lyophilization and spray drying
using a model monoclonal antibody (rhuMAb) with or without sucrose. For
spray-dried samples, increasing the molar ratio of sucrose:rhuMAb from 55:1
to 320:1 decreased the aggregation rate ivefold during storage at 30 °C [36].
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Assuming a mass of 150 kDa, the 320:1 ratio would correspond to roughly
55% antibody in the dry powder by mass. A high active loading such as this
could enable a wide range of dosing strengths, up to approximately 15–18 mg
active in a DPI.
Andya et al. spray-dried rhuMAbE25, an anti-IgE antibody, in formulations
composed of trehalose, lactose, or mannitol from sugar:rhuMAbE25 molar
ratios of 100:1 to 900:1 [44]. These formulations were then assayed for
aggregation following reconstitution by size-exclusion chromatography
(SEC) and the ine particle fraction (FPF, fraction of particles with mass
median aerodynamic diameters <6.4 μm) by using a four-stage inertial
impactor. The powders were also stored at 5, 30, and 40 °C for 1 year. All
trehalose:rhuMAbE25 molar ratios above 100:1 minimized aggregation (<1%
aggregate content vs. 5% without sugar), but ratios above 200:1 decreased
FPF from 30–35% to 10–15%. In contrast, all lactose:rhuMAbE25 molar ratios
from 100:1 to 600:1 exhibited an FPF of ~30% and had aggregation <1%.
However, storage of lactose particles for 9 months at 30 °C led to irreversible
glycation through Maillard reactions that were not observed during storage at
5 °C. Mannitol formulations all had higher aggregate content (2–3%) and
further increasing mannitol steadily decreased FPF. All formulations
decreased aggregation rate constants following storage of powders for 1 year
at 30 °C. The beneit of mannitol plateaued at sugar:Ab ratios >100:1, while
increasing trehalose and lactose content beyond 100:1 continued to decrease
the aggregation rate constant. This observation was corroborated by powder
X-ray diffraction (PXRD). After 1 year at 30 °C, trehalose at 900:1 and lactose
at 600:1 showed no signs of crystallinity. In contrast, the 500:1 mannitol
formulation showed a clear crystalline ingerprint. Although the 200:1 and
100:1 formulations did not exhibit crystallinity by PXRD, the extent of
crystallization may have been at levels below the detection limit of the assay.
These results emphasize that there may be compromises implicit in
simultaneously optimizing both stability and powder properties, especially
depending on the identity and/or amounts of the excipient.
Schu le et al. evaluated solutions containing 0–80% mannitol for spray
drying a model IgG antibody [45]. Higher mannitol content (50–80%) led to
an increase in aggregates as measured by SEC and turbidity, while 20–30%
mannitol formulations were identical to 100% IgG solution. They compared
aggregation in solutions comprising 0%, 30%, and 70% mannitol before and
after one week at 40 °C. The 0% and 30% solutions initially had ~0.5%
aggregate content while the 70% formulation started at 3%, indicating higher
mannitol concentrations destabilize protein structure upon formulation.
After 1 week in solution, the 30% mannitol mixture had unchanged aggregate
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content, while the 0% and 70% mixtures increased to 1.5% and 6%,
respectively. After spray drying and powder dissolution, the powders
contained 0.7–6.4% aggregates; lower mannitol content (20–40%)
maximized stability during spray drying, while higher mannitol content
promoted accelerated aggregation, echoing the results from the Andya et al.
studies. FTIR analysis of the spray-dried powders showed a different amide I
band ingerprint compared to the IgG feed solution. Interestingly, after
reconstitution of the SDI powders, the solutions reverted to the original IR
proile, suggesting reversible denaturation occurs during drying [45]. The
reversibility and magnitude of denaturation during drying may vary
according to the antibody structure and sequence. This could have important
implications for clinical development as denaturation could disrupt antigen
binding and lead to aggregation, which may lead to diminished eficacy and
subsequent immunogenicity in vivo [7].
It is the authors’ opinion, based on available evidence, that to maximize
the stabilizing impact of sugar, the protein and sugar molecules must be in
intimate contact. Thus, manufacturing a single-phase amorphous dispersion
of sugar and protein should be prioritized. Trehalose is the foremost example
of a nonreducing sugar that is often physically stable in an amorphous
dispersion with a protein. Studies have also shown mannitol to be a useful
stabilizing excipient, though it is prone to crystallization during spray drying
under many conditions and formulation compositions. An amorphous
dispersion of mannitol and protein would be preferable to maximize
molecular interaction, but challenging to manufacture.
3.1.2 StabilizingAminoAci ds
Amino acids have also been assessed as DPI mAb excipients, likely serving a
variety of functions dependent on the side chain functionality. In
collaboration with UCB Pharma, the Massant et al. spray-dried two model IgG
antibodies formulated with combinations of sucrose, trehalose, and an amino
acid (13 total screened) at varying concentrations [46]. They found that
stability was almost entirely dictated by the sugar; increasing sucrose or
trehalose from 5 to 75 mg/mL decreased weekly aggregation rates from ~2%
to 0.1%. These results were corroborated by Sane et al., who found that
sugars provided the greatest beneit to long-term stability and histidine only
provided a minor protective effect during drying [36]. It is worthwhile to note
that in both studies, only positively charged amino acid excipients provided
stabilizing effects. The authors suggest that the charged guanidinium group
of arginine can form strong hydrogen and π-bonds, while its aliphatic chain
can favorably interact with hydrophobic surfaces. While histidine can also
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