formulation design
HOW TO ADMINISTER AMORPHOUS SOLID DISPERSIONS TO SMALL LABORATORY ANIMALS ?
Amorphous solid dispersions (ASDs) are frequently used to enhance the solubility and bioavailability of active pharmaceutical ingredients (APIs) in discovery and development programs. However, ASDs present unique challenges when it comes to administration to small laboratory animals such as mice and rats.
Unlike many other solubility-enhancing formulations that are liquid in their native form (cyclodextrin solutions, many lipid-based formulations, nanosuspensions), ASDs are solid products. Dedicated capsules for dosing solids to small laboratory animals exist (for example size 9 capsules with a fill volume of about 0.025 mL), but the mg-quantities of formulation they hold usually fall well short of the doses targeted in early pharmacology and toxicology studies, which often range from 100 to 1000 mg/kg.
High API doses are much more readily achieved with liquid formulations. A dose volume of 10 mL per kg body weight is widely regarded as good practice for oral gavage in rodents, corresponding to 2.5 mL for a 250 g rat or 0.25 mL for a 25 g mouse; published guidance allows up to 20 mL/kg in rats where justified. Dispersing the ASD powder in a liquid vehicle and administering it as a suspension therefore makes far higher doses attainable than with a capsule.
The challenge of dispersing ASDs in a gavage vehicle
Dispersing an ASD powder in a liquid vehicle creates a new set of problems. Practitioners generally prefer mild aqueous buffers as gavage vehicles for reasons of tolerability, but exposure to aqueous media can greatly reduce the in vivo performance of ASDs. First, water may migrate into the ASD particles, plasticizing the amorphous matrix and inducing API crystallization. Second, API may be released into the vehicle, where it is susceptible to precipitation into a less soluble form. Both effects can cause substantial crystallization before the dose is even administered, largely negating the ASD's potential to improve oral absorption.
A further practical problem at high ASD concentrations is the viscosity increase caused by polymer hydration. Even low-viscosity grades such as HPMC E5, a common ASD carrier, can reach viscosities incompatible with administration through a narrow gavage needle when the polymer concentration in the vehicle is high, as it is at high ASD loadings. Typical needles are around 16–18 G for rats and 20–22 G for mice.
Two main tactics reduce the risk of API crystallization during dispersion: designing the ASD around a polymer with pH-dependent solubility, and using a non-aqueous vehicle. Both also help keep viscosity under control, so that the dispersion remains syringeable.
Designing ASDs based on polymers with pH-dependent solubility
A wide range of pharmaceutical-grade polymers with pH-dependent solubility is commercially available. Most were originally developed as enteric coatings, but several have proven very effective as ASD carriers, the best known being hydroxypropyl methylcellulose acetate succinate (HPMCAS). These polymers remain undissolved at low pH and dissolve above a grade-specific threshold, typically between pH 5 and 7.
Dispersed in an acidic gavage vehicle, ASDs based on these polymers are considerably more robust than those based on water-soluble carriers. Because the polymer does not dissolve, API release into the vehicle is limited and the matrix is less prone to plasticization. The protection is not absolute, however: undissolved polymers such as HPMCAS still take up water when immersed in liquid, lowering the glass transition temperature of the dispersion. Physical stability in the vehicle therefore still needs to be verified.
Account for rodent gastrointestinal pH
Polymer selection should reflect the physiology of the species being dosed, not human values. Rodent GI pH differs markedly from human pH. Gastric pH in mice and rats is typically higher than in humans (roughly pH 3–4.5), while small-intestinal pH is lower: approximately pH 4.7–5.2 in mice and about pH 5–6.5 in rats (McConnell et al., J Pharm Pharmacol, 2008; approximate values). As a result, polymers with higher dissolution thresholds, such as HPMCAS-H or Eudragit S100, may dissolve slowly or incompletely in the rodent small intestine, and even M grades may be borderline in mice. For rodent studies, low-threshold grades such as HPMCAS-L or Eudragit L100-55 are usually the safer choice, and in vitro dissolution testing in media that reflect rodent intestinal pH is advisable.
Setting the vehicle pH
A practical starting point is a vehicle pH about one unit below the polymer's dissolution threshold, with buffer capacity kept as low as possible so as not to disturb pH in the small intestine, where rapid polymer dissolution is wanted. Treat this as a rule of thumb rather than a guaranteed safety margin. For HPMCAS-L, for instance, one unit below threshold is around pH 4.5, close to the pKa of the succinoyl groups (about 5), where the polymer is already partly ionized and may swell. For such grades, a lower vehicle pH may be needed.
Consider the ionization behavior of the API
The approach works best for neutral and weakly acidic APIs. Weakly basic APIs can be highly soluble at low pH, so an acidic vehicle may leach the API out of the ASD before dosing, even while the polymer itself stays intact. The dissolved API may then precipitate in the vehicle, form a salt, or precipitate in the intestine as pH rises. For weak bases, the vehicle pH should be chosen with the API's pKa and pH-solubility profile in mind, and API release into the vehicle should be measured directly.
Benefits and verification
Where the conditions above are met, an additional benefit is that API release is targeted to the small intestine, the main site of absorption. This reduces potential performance losses from supersaturation and precipitation in the stomach, where absorption capacity is small relative to the small intestine.
To verify that an ASD powder resists vehicle-induced crystallization, disperse it in the vehicle of interest, sample over the intended holding time, and examine the samples with X-ray powder diffraction or polarized light microscopy.
Using non-aqueous gavage vehicles
If enteric polymers are not an option, for instance because early formulation work showed that none of them adequately stabilizes the amorphous API, or because the API is a weak base that leaches readily into acidic media, non-aqueous vehicles can be evaluated. Tolerability is the key selection criterion, as relatively large volumes of vehicle are co-administered to deliver the target ASD dose. Triglyceride-rich pharmaceutical-grade oils such as corn oil and soybean oil are frequently used. Dispersion in such vehicles carries a much lower risk of matrix plasticization and premature crystallization, but absence of crystallization should still be verified.
When interpreting pharmacokinetic data, keep in mind that the vehicle itself may have promoted intestinal solubilization and absorption, sometimes dramatically. Lipids and lipophilic excipients can alter the gastrointestinal environment, for example by stimulating bile secretion, as we discussed in one of our earlier articles. While this may be welcome in discovery and preclinical studies, the "vehicle effect" may disappear later in development, when ASDs are administered without a gavage vehicle (for example in dog studies or early clinical studies).
If you would like to learn more about our approach to amorphous solid dispersion design, you can reach out to one of our experts directly via the button below.
Unlike many other solubility-enhancing formulations that are liquid in their native form (cyclodextrin solutions, many lipid-based formulations, nanosuspensions), ASDs are solid products. Dedicated capsules for dosing solids to small laboratory animals exist (for example size 9 capsules with a fill volume of about 0.025 mL), but the mg-quantities of formulation they hold usually fall well short of the doses targeted in early pharmacology and toxicology studies, which often range from 100 to 1000 mg/kg.
High API doses are much more readily achieved with liquid formulations. A dose volume of 10 mL per kg body weight is widely regarded as good practice for oral gavage in rodents, corresponding to 2.5 mL for a 250 g rat or 0.25 mL for a 25 g mouse; published guidance allows up to 20 mL/kg in rats where justified. Dispersing the ASD powder in a liquid vehicle and administering it as a suspension therefore makes far higher doses attainable than with a capsule.
The challenge of dispersing ASDs in a gavage vehicle
Dispersing an ASD powder in a liquid vehicle creates a new set of problems. Practitioners generally prefer mild aqueous buffers as gavage vehicles for reasons of tolerability, but exposure to aqueous media can greatly reduce the in vivo performance of ASDs. First, water may migrate into the ASD particles, plasticizing the amorphous matrix and inducing API crystallization. Second, API may be released into the vehicle, where it is susceptible to precipitation into a less soluble form. Both effects can cause substantial crystallization before the dose is even administered, largely negating the ASD's potential to improve oral absorption.
A further practical problem at high ASD concentrations is the viscosity increase caused by polymer hydration. Even low-viscosity grades such as HPMC E5, a common ASD carrier, can reach viscosities incompatible with administration through a narrow gavage needle when the polymer concentration in the vehicle is high, as it is at high ASD loadings. Typical needles are around 16–18 G for rats and 20–22 G for mice.
Two main tactics reduce the risk of API crystallization during dispersion: designing the ASD around a polymer with pH-dependent solubility, and using a non-aqueous vehicle. Both also help keep viscosity under control, so that the dispersion remains syringeable.
Designing ASDs based on polymers with pH-dependent solubility
A wide range of pharmaceutical-grade polymers with pH-dependent solubility is commercially available. Most were originally developed as enteric coatings, but several have proven very effective as ASD carriers, the best known being hydroxypropyl methylcellulose acetate succinate (HPMCAS). These polymers remain undissolved at low pH and dissolve above a grade-specific threshold, typically between pH 5 and 7.
Dispersed in an acidic gavage vehicle, ASDs based on these polymers are considerably more robust than those based on water-soluble carriers. Because the polymer does not dissolve, API release into the vehicle is limited and the matrix is less prone to plasticization. The protection is not absolute, however: undissolved polymers such as HPMCAS still take up water when immersed in liquid, lowering the glass transition temperature of the dispersion. Physical stability in the vehicle therefore still needs to be verified.
Account for rodent gastrointestinal pH
Polymer selection should reflect the physiology of the species being dosed, not human values. Rodent GI pH differs markedly from human pH. Gastric pH in mice and rats is typically higher than in humans (roughly pH 3–4.5), while small-intestinal pH is lower: approximately pH 4.7–5.2 in mice and about pH 5–6.5 in rats (McConnell et al., J Pharm Pharmacol, 2008; approximate values). As a result, polymers with higher dissolution thresholds, such as HPMCAS-H or Eudragit S100, may dissolve slowly or incompletely in the rodent small intestine, and even M grades may be borderline in mice. For rodent studies, low-threshold grades such as HPMCAS-L or Eudragit L100-55 are usually the safer choice, and in vitro dissolution testing in media that reflect rodent intestinal pH is advisable.
Setting the vehicle pH
A practical starting point is a vehicle pH about one unit below the polymer's dissolution threshold, with buffer capacity kept as low as possible so as not to disturb pH in the small intestine, where rapid polymer dissolution is wanted. Treat this as a rule of thumb rather than a guaranteed safety margin. For HPMCAS-L, for instance, one unit below threshold is around pH 4.5, close to the pKa of the succinoyl groups (about 5), where the polymer is already partly ionized and may swell. For such grades, a lower vehicle pH may be needed.
Consider the ionization behavior of the API
The approach works best for neutral and weakly acidic APIs. Weakly basic APIs can be highly soluble at low pH, so an acidic vehicle may leach the API out of the ASD before dosing, even while the polymer itself stays intact. The dissolved API may then precipitate in the vehicle, form a salt, or precipitate in the intestine as pH rises. For weak bases, the vehicle pH should be chosen with the API's pKa and pH-solubility profile in mind, and API release into the vehicle should be measured directly.
Benefits and verification
Where the conditions above are met, an additional benefit is that API release is targeted to the small intestine, the main site of absorption. This reduces potential performance losses from supersaturation and precipitation in the stomach, where absorption capacity is small relative to the small intestine.
To verify that an ASD powder resists vehicle-induced crystallization, disperse it in the vehicle of interest, sample over the intended holding time, and examine the samples with X-ray powder diffraction or polarized light microscopy.
Using non-aqueous gavage vehicles
If enteric polymers are not an option, for instance because early formulation work showed that none of them adequately stabilizes the amorphous API, or because the API is a weak base that leaches readily into acidic media, non-aqueous vehicles can be evaluated. Tolerability is the key selection criterion, as relatively large volumes of vehicle are co-administered to deliver the target ASD dose. Triglyceride-rich pharmaceutical-grade oils such as corn oil and soybean oil are frequently used. Dispersion in such vehicles carries a much lower risk of matrix plasticization and premature crystallization, but absence of crystallization should still be verified.
When interpreting pharmacokinetic data, keep in mind that the vehicle itself may have promoted intestinal solubilization and absorption, sometimes dramatically. Lipids and lipophilic excipients can alter the gastrointestinal environment, for example by stimulating bile secretion, as we discussed in one of our earlier articles. While this may be welcome in discovery and preclinical studies, the "vehicle effect" may disappear later in development, when ASDs are administered without a gavage vehicle (for example in dog studies or early clinical studies).
If you would like to learn more about our approach to amorphous solid dispersion design, you can reach out to one of our experts directly via the button below.
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