A novel simulated media system for in vitro evaluation of bioequivalent intestinal drug solubility.

Orally administered solid drug must dissolve in the gastrointestinal tract before absorption to provide a systemic response. Intestinal solubility is therefore crucial but difficult to measure since human intestinal fluid (HIF) is challenging to obtain, varies between fasted (Fa) and fed (Fe) states and exhibits inter and intra subject variability. A single simulated intestinal fluid (SIF) cannot reflect HIF variability, therefore current approaches are not optimal. In this study we have compared literature Fa/FeHIF drug solubilities to values measured in a novel in vitro simulated nine media system for either the fasted (Fa9SIF) or fed (Fe9SIF) state. The manuscript contains 129 literature sampled human intestinal fluid equilibrium solubility values and 387 simulated intestinal fluid equilibrium solubility values. Statistical comparison does not detect a difference (Fa/Fe9SIF vs Fa/FeHIF), a novel solubility correlation window enclosed 95% of an additional literature Fa/FeHIF data set and solubility behaviour is consistent with previous physicochemical studies. The Fa/Fe9SIF system therefore represents a novel in vitro methodology for bioequivalent intestinal solubility determination. Combined with intestinal permeability this provides an improved, population based, biopharmaceutical assessment that guides formulation development and indicates the presence of food based solubility effects. This transforms predictive ability during drug discovery and development and may represent a methodology applicable to other multicomponent fluids where no single component is responsible for performance.

Structured solubility behaviour in fed simulated intestinal fluids.

Intestinal drug solubility is a key parameter controlling absorption after the administration of a solid oral dosage form. The ability to measure fed state solubility in vitro is limited and multiple simulated intestinal fluid recipes have been developed but with no consensus which is optimal. This study has utilised nine bioequivalent simulated fed intestinal media recipes that cover over 90% of the compositional variability of sampled fed human intestinal fluid. The solubility of 24 drugs (Acidic; furosemide, ibuprofen, indomethacin, mefenamic acid, naproxen, phenytoin, piroxicam, valsartan, zafirlukast: Basic; aprepitant, atazanavir, bromocriptine, carvedilol, dipyridamole, posaconazole, tadalafil: Neutral; acyclovir, carbamazepine, felodipine, fenofibrate, griseofulvin, itraconazole, paracetamol, probucol) has been assessed to determine if structured solubility behaviour is present. The measured solubility behaviour can be split into four categories and is consistent with drug physicochemical properties and previous solubility studies. For acidic drugs (category 1) solubility is controlled by media pH and the lowest and highest pH media identify the lowest and highest solubility in 90% of cases. For weakly acidic, basic and neutral drugs (category 2) solubility is controlled by media pH and total amphiphile concentration (TAC), a consistent solubility pattern is evident with variation related to individual drug media component interactions. The lowest and highest pH × TAC media identify the lowest and highest solubility in 70% and 90% of cases respectively. Four drugs, which are non-ionised in the media systems (category 3), have been identified with a very narrow solubility range, indicating minimal impact of the simulated media on solubility. Three drugs exhibit solubility behaviour that is not consistent with the remainder (category 4). The results indicate that the use of two bioequivalent fed intestinal media from the original nine will identify in vitro the maximum and minimum solubility values for the majority of drugs and due to the media derivation this is probably applicable in vivo. When combined with a previous fasted study, this introduces interesting possibilities to measure a solubility range in vitro that can provide Quality by Design based decisions to rationalise drug and formulation development. Overall this indicates that the multi-dimensional media system is worthy of further investigation as in vitro tool to assess fed intestinal solubility.

Excipient Impact on Fenofibrate Equilibrium Solubility in Fasted and Fed Simulated Intestinal Fluids Assessed Using a Design of Experiment Protocol.

Solubility is a critical parameter controlling drug absorption after oral administration. For poorly soluble drugs, solubility is influenced by the complex composition of intestinal media and the influence of dosage form excipients, which can cause bioavailability and bioequivalence issues. This study has applied a small scale design of experiment (DoE) equilibrium solubility approach in order to investigate the impact of excipients on fenofibrate solubility in simulated fasted and fed intestinal media. Seven media parameters (bile salt (BS), phospholipid (PL), fatty acid, monoglyceride, cholesterol, pH and BS/PL ratio) were assessed in the DoE and in excipient-free media, and only pH and sodium oleate in the fasted state had a significant impact on fenofibrate solubility. The impact of excipients were studied at two concentrations, and for polyvinylpyrrolidone (PVP, K12 and K29/32) and hydroxypropylmethylcellulose (HPMC, E3 and E50), two grades were studied. Mannitol had no solubility impact in any of the DoE media. PVP significantly increased solubility in a media-, grade- and concentration-dependent manner, with the biggest change in fasted media. HPMC and chitosan significantly reduced solubility in both fasted and fed states in a media-, grade- and concentration-dependent manner. The results indicate that the impact of excipients on fenofibrate solubility is a complex interplay of media composition in combination with their physicochemical properties and concentration. The results indicate that in vitro solubility studies combining the drug of interest, proposed excipients along with suitable simulated intestinal media recipes will provide interesting information with the potential to guide formulation development.

Fed intestinal solubility limits and distributions applied to the Developability classification system.

For solid oral dosage forms drug solubility in intestinal fluid is an important parameter influencing product performance and bioavailability. Solubility along with permeability are the two parameters applied in the Biopharmaceutics and Developability Classification Systems (DCS) to assess a drug's potential for oral administration. Intestinal solubility varies with the intestinal contents and the differences between the fasted and fed states are recognised to influence solubility and bioavailability. In this study a novel fed state simulated media system comprising of nine media has been utilised to measure the solubility of seven drugs (ibuprofen, mefenamic acid, furosemide, dipyridamole, griseofulvin, paracetamol and acyclovir) previously studied in the fasted state DCS. The results demonstrate that the fed nine media system provides a range of solubility values for each drug and solubility behaviour is consistent with published design of experiment studies conducted in either the fed or fasted state. Three drugs (griseofulvin, paracetamol and acyclovir) exhibit very narrow solubility distributions, a result that matches published behaviour in the fasted state, indicating that this property is not influenced by the concentration of simulated media components. The nine solubility values for each drug can be utilised to calculate a dose/solubility volume ratio to visualise the drug's position on the DCS grid. Due to the derivation of the nine media compositions the range and catergorisation could be considered as bioequivalent and can be combined with the data from the original fed intestinal fluid analysis to provide a population based solubility distribution. This provides further information on the drugs solubility behaviour and could be applied to quality by design formulation approaches. Comparison of the fed results in this study with similar published fasted results highlight that some differences detected match in vivo behaviour in food effect studies. This indicates that a combination of the fed and fasted systems may be a useful in vitro biopharmaceutical performance tool. However, it should be noted that the fed media recipes in this study are based on a liquid meal (Ensure Plus) and this may not be representative of alternative fed states achieved through ingestion of a solid meal. Nevertheless, this novel approach provides greater in vitro detail with respect to possible in vivo biopharmaceutical performance, an improved ability to apply risk-based approaches and the potential to investigate solubility based food effects. The system is therefore worthy of further investigation but studies will be required to expand the number of drugs measured and link the in vitro measurements to in vivo results.

High spatial resolution ToF-SIMS imaging and image analysis strategies to monitor and quantify early phase separation in amorphous solid dispersions.

Amorphous solid dispersions (ASDs) are formulations with enhanced drug solubility and dissolution rate compared to their crystalline counterparts, however, they can be inherently thermodynamically unstable. This can lead to amorphous phase separation and drug re-crystallisation, phenomena that are typically faster and more dominant at the product's surfaces. This study investigates the use of high-resolution time of flight-secondary ion mass spectrometry (ToF-SIMS) imaging as a surface analysis technique combined with image-analysis for the early detection, monitoring and quantification of surface amorphous phase separation in ASDs. Its capabilities are demonstrated for two pharmaceutically relevant ASD systems with distinct re-crystallisation behaviours, prepared using hot melt extrusion (HME) followed by pelletisation or grinding: (1) paracetamol-hydroxypropyl methylcellulose (PCM-HPMC) pellets with drug loadings of 10%-50% w/w and (2) indomethacin-polyvinylpyrrolidone (IND-PVP) ground material with drug loadings of 20%-85% w/w. PCM-HPMC pellets showed intense phase separation, reaching 100% PCM surface coverage within 1-5 months. In direct comparison, IND-PVP HME ground material was more stable with only a moderate formation of isolated IND-rich clusters. Image analysis allowed the reliable detection and quantification of local drug-rich clusters. An Avrami model was applied to determine and compare phase separation kinetics. The combination of chemical sensitivity and high spatial resolution afforded by SIMS was crucial to enable the study of early phase separation and re-crystallisation at the surface. Compared with traditional methods used to detect crystalline material, such as XRPD, we show that ToF-SIMS enabled detection of surface physical instability already at early stages of drug cluster formation in the first days of storage.

Microfluidic Manufacture of Lipid-Based Nanomedicines.

Nanoparticulate technologies have revolutionized drug delivery allowing for passive and active targeting, altered biodistribution, controlled drug release (temporospatial or triggered), enhanced stability, improved solubilization capacity, and a reduction in dose and adverse effects. However, their manufacture remains immature, and challenges exist on an industrial scale due to high batch-to-batch variability hindering their clinical translation. Lipid-based nanomedicines remain the most widely approved nanomedicines, and their current manufacturing methods remain discontinuous and face several problems such as high batch-to-batch variability affecting the critical quality attributes (CQAs) of the product, laborious multistep processes, need for an expert workforce, and not being easily amenable to industrial scale-up involving typically a complex process control. Several techniques have emerged in recent years for nanomedicine manufacture, but a paradigm shift occurred when microfluidic strategies able to mix fluids in channels with dimensions of tens of micrometers and small volumes of liquid reagents in a highly controlled manner to form nanoparticles with tunable and reproducible structure were employed. In this review, we summarize the recent advancements in the manufacturing of lipid-based nanomedicines using microfluidics with particular emphasis on the parameters that govern the control of CQAs of final nanomedicines. The impact of microfluidic environments on formation dynamics of nanomaterials, and the application of microdevices as platforms for nanomaterial screening are also discussed.

Linked experimental and modelling approaches for tablet property predictions.

Recent years have seen the advent of Quality-by-Design (QbD) as a philosophy to ensure the quality, safety, and efficiency of pharmaceutical production. The key pharmaceutical processing methodology of Direct Compression to produce tablets is also the focus of some research. The traditional Design-of-Experiments and purely experimental approach to achieve such quality and process development goals can have significant time and resource requirements. The present work evaluates potential for using combined modelling and experimental approach, which may reduce this burden by predicting the properties of multicomponent tablets from pure component compression and compaction model parameters. Additionally, it evaluates the use of extrapolation from binary tablet data to determine theoretical pure component model parameters for materials that cannot be compacted in the pure form. It was found that extrapolation using binary tablet data - where one known component can be compacted in pure form and the other is a challenging material which cannot be - is possible. Various mixing rules have been evaluated to assess which are suitable for multicomponent tablet property prediction, and in the present work linear averaging using pre-compression volume fractions has been found to be the most suitable for compression model parameters, while for compaction it has been found that averaging using a power law equation form produced the best agreement with experimental data. Different approaches for estimating component volume fractions have also been evaluated, and using estimations based on theoretical relative rates of compression of the pure components has been found to perform slightly better than using constant volume fractions (that assume a fully compressed mixture). The approach presented in this work (extrapolation of, where necessary, binary tablet data combined with mixing rules using volume fractions) provides a framework and path for predictions for multicomponent tablets without the need for any additional fitting based on the multicomponent formulation composition. It allows the knowledge space of the tablet to be rapidly evaluated, and key regions of interest to be identified for follow-up, targeted experiments that that could lead to an establishment of a design and control space and forgo a laborious initial Design-of-Experiments.

Crystal Structure and Twisted Aggregates of Oxcarbazepine Form III.

Polymorphism and crystal habit play vital roles in dictating the properties of crystalline materials. Here, the structure and properties of oxcarbazepine (OXCBZ) form III are reported along with the occurrence of twisted crystalline aggregates of this metastable polymorph. OXCBZ III can be produced by crystallization from the vapor phase and by recrystallization from solution. The crystallization process used to obtain OXCBZ III is found to affect the pitch, with the most prominent effect observed from the sublimation-grown OXCBZ III material where the pitch increases as the length of aggregates increases. Sublimation-grown OXCBZ III follows an unconventional mechanism of formation with condensed droplet formation and coalescence preceding nucleation and growth of aggregates. A crystal structure determination of OXCBZ III from powder X-ray diffraction methods, assisted by crystal structure prediction (CSP), reveals that OXCBZ III, similar to carbamazepine form II, contains void channels in its structure with the channels, aligned along the c crystallographic axis, oriented parallel to the twist axis of the aggregates. The likely role of structural misalignment at the lattice or nanoscale is explored by considering the role of molecular and closely related structural impurities informed by crystal structure prediction.

Small scale in vitro method to determine a potential bioequivalent equilibrium solubility range for fed human intestinal fluid.

Intestinal drug solubility is a key parameter controlling oral absorption but varies both intra and inter individuals and between the fasted and fed states, with food intake known to alter the bioavailability of many compounds. Intestinal solubility can be measured in vitro either using sampled fed human intestinal fluid (FeHIF) or simulated fed intestinal fluid (SIF) but neither approach is optimal. FeHIF is difficult to obtain and variable, whilst for fed SIF multiple recipes are available with no consensus on the ideal version. A recent study characterised FeHIF aspirates using a multidimensional approach and calculated nine simulated media recipes that covered over ninety percent of FeHIF compositional variability. In this study the equilibrium solubility of thirteen drugs have been measured using the nine simulated media recipes and compared to multiple previous design of experiment (DoE) studies, which have examined the impact of fed SIF media components on solubility. The measured nine media solubility data set is only statistically equivalent to the large scale 92 media DoE in 4 out of 13 drug comparisons, but has improved equivalence against small scale DoEs (9 or 10 media) with 6 out of 9 or 10 out of 12 (9 and 10 media respectively) equivalent. Selective removal of non-biorelevant compositions from the 92 media DoE improves statistical equivalence to 9 out of 13 comparisons. The results indicate that solubility equivalence is linked to media component concentrations and compositions, the nine media system is measuring a similar solubility space to previous systems, with a narrower solubility range than the 92 point DoE but equivalent to smaller DoE systems. Phenytoin and tadalafil display a narrow solubility range, a behaviour consistent with previous studies in fed and fasted states and only revealed through the multiple media approach. Custom DoE analysis of the nine media results to determine the most statistically significant component influencing solubility does not detect significant components. Indicating that the approach has a low statistical resolution and is not appropriate if determination of media component significance is required. This study demonstrates that it is possible to assess the fed intestinal equilibrium solubility envelope using the nine media recipes obtained from a multi-dimensional analysis of fed HIF. The derivation of the nine media compositions coupled with the results in this study indicate that the solubility results are more likely to reflect the fed intestinal solubility envelope than previous DoE studies and highlight that the system is worthy of further investigation.

Manufacture of tablets with structurally-controlled drug release using rapid tooling injection moulding.

With advancements in the pharmaceutical industry pushing more towards tailored medicines, novel approaches to tablet manufacture are in high demand. One of the main drivers towards micro-scale batch production is the ability to fine-tune drug release. This study demonstrates the use of rapid tooling injection moulding (RTIM) for tablet manufacture. Tablets were manufactured with varying structural features to alter the surface area whilst maintaining the same volume, resulting in differing specific surface area (SSA). The precision of this technique is evaluated based on eleven polymer formulations, with the tablets displaying <2% variability in mass. Further tablets were produced containing paracetamol in three different polymer-based formulations to investigate the impact of SSA on the drug release. Significant differences were observed between the formulations based on the polymers polyvinyl alcohol (PVA) and Klucel ELF. The polymer base of the formulation was found to be critical to the sensitivity of the drug release profile to SSA modification. The drug release profile within each formulation was modified by the addition of structural features to increase the SSA.

Structured solubility behaviour in bioequivalent fasted simulated intestinal fluids.

Drug solubility in intestinal fluid is a key parameter controlling absorption after the administration of a solid oral dosage form. To measure solubility in vitro simulated intestinal fluids have been developed, but there are multiple recipes and the optimum is unknown. This situation creates difficulties during drug discovery and development research. A recent study characterised sampled fasted intestinal fluids using a multidimensional approach to derive nine bioequivalent fasted intestinal media that covered over 90% of the compositional variability. These media have been applied in this study to examine the equilibrium solubility of twenty one exemplar drugs (naproxen, indomethacin, phenytoin, zafirlukast, piroxicam, ibuprofen, mefenamic acid, furosemide, aprepitant, carvedilol, tadalafil, dipyridamole, posaconazole, atazanavir, fenofibrate, felodipine, griseofulvin, probucol, paracetamol, acyclovir and carbamazepine) to determine if consistent solubility behaviour was present. The bioequivalent media provide in the majority of cases structured solubility behaviour that is consistent with physicochemical properties and previous solubility studies. For the acidic drugs (pKa < 6.3) solubility is controlled by media pH, the profile is identical and consistent and the lowest and highest pH media identify the lowest and highest solubility in over 70% of cases. For weakly acidic (pKa > 8), basic and neutral drugs solubility is controlled by a combination of media pH and total amphiphile concentration (TAC), a consistent solubility behaviour is evident but with variation related to individual drug interactions within the media. The lowest and highest pH × TAC media identify the lowest and highest solubility in over 78% of cases. A subset of the latter category consisting of neutral and drugs non-ionised in the media pH range have been identified with a very narrow solubility range, indicating that the impact of the simulated intestinal media on their solubility is minimal. Two drugs probucol and atazanavir exhibit unusual behaviour. The study indicates that the use of two appropriate bioequivalent fasted intestinal media from the nine will identify in vitro the maximum and minimum solubility boundaries for drugs and due to the media derivation this is probably applicable in vivo. These media could be applied during discovery and development activities to provide a solubility range, which would assist placement of the drug within the BCS/DCS and rationalise drug and formulation decisions.

Intermittent PI3Kδ inhibition sustains anti-tumour immunity and curbs irAEs.

Phosphoinositide 3-kinase δ (PI3Kδ) has a key role in lymphocytes, and inhibitors that target this PI3K have been approved for treatment of B cell malignancies1-3. Although studies in mouse models of solid tumours have demonstrated that PI3Kδ inhibitors (PI3Kδi) can induce anti-tumour immunity4,5, its effect on solid tumours in humans remains unclear. Here we assessed the effects of the PI3Kδi AMG319 in human patients with head and neck cancer in a neoadjuvant, double-blind, placebo-controlled randomized phase II trial (EudraCT no. 2014-004388-20). PI3Kδ inhibition decreased the number of tumour-infiltrating regulatory T (Treg) cells and enhanced the cytotoxic potential of tumour-infiltrating T cells. At the tested doses of AMG319, immune-related adverse events (irAEs) required treatment to be discontinued in 12 out of 21 of patients treated with AMG319, suggestive of systemic effects on Treg cells. Accordingly, in mouse models, PI3Kδi decreased the number of Treg cells systemically and caused colitis. Single-cell RNA-sequencing analysis revealed a PI3Kδi-driven loss of tissue-resident colonic ST2 Treg cells, accompanied by expansion of pathogenic T helper 17 (TH17) and type 17 CD8+ T (TC17) cells, which probably contributed to toxicity; this points towards a specific mode of action for the emergence of irAEs. A modified treatment regimen with intermittent dosing of PI3Kδi in mouse models led to a significant decrease in tumour growth without inducing pathogenic T cells in colonic tissue, indicating that alternative dosing regimens might limit toxicity.

Mefenamic acid solid dispersions: Impact of formulation composition on processing parameters, product properties and performance.

The objective of this study was to develop an immediate release (IR), crystalline solid dispersion (CSD) formulation of Mefenamic acid (MFA) by hot-melt-extrusion (HME) and assess the impact of drug loading on process parameters, product physico-chemical properties and product performance. An HME process to produce a range of MFA-Soluplus®-Sorbitol polymer matrix CSD formulations was developed based on rheological screening assays of physical mixtures (PM). The impact of drug loading on process parameters was compared to the impact of drug loading on the physico-chemical properties of formulations. Based on process and product data, three groupings of API drug loading were identified: sub-saturated, saturated, and supersaturated systems. CSD formulations were obtained for 20-50% (w/w) drug loading containing the stable polymorphic form I of MFA. CSD formulations predominantly improved the consistency of the product performance. An Amorphous Solid Dispersion (ASD) was obtained for 10% (w/w) drug loading, exhibiting faster drug release even at physiologically relevant pH. This study illustrates the impact of drug loading on process and product characteristics and how a better understanding of maximum API solubility in a given polymer system can improve targeted formulation development.

Fasted intestinal solubility limits and distributions applied to the biopharmaceutics and developability classification systems.

After oral administration, a drug's solubility in intestinal fluid is an important parameter influencing bioavailability and if the value is known it can be applied to estimate multiple biopharmaceutical parameters including the solubility limited absorbable dose. Current in vitro measurements may utilise fasted human intestinal fluid (HIF) or simulated intestinal fluid (SIF) to provide an intestinal solubility value. This single point value is limited since its position in relation to the fasted intestinal solubility envelope is unknown. In this study we have applied a nine point fasted equilibrium solubility determination in SIF, based on a multi-dimensional analysis of fasted human intestinal fluid composition, to seven drugs that were previously utilised to investigate the developability classification system (ibuprofen, mefenamic acid, furosemide, dipyridamole, griseofulvin, paracetamol and acyclovir). The resulting fasted equilibrium solubility envelope encompasses literature solubility values in both HIF and SIF indicating that it measures the same solubility space as current approaches with solubility behaviour consistent with previous SIF design of experiment studies. In addition, it identifies that three drugs (griseofulvin, paracetamol and acyclovir) have a very narrow solubility range, a feature that single point solubility approaches would miss. The measured mid-point solubility value is statistically equivalent to the value determined with the original fasted simulated intestinal fluid recipe, further indicating similarity and that existing literature results could be utilised as a direct comparison. Since the multi-dimensional approach covered greater than ninety percent of the variability in fasted intestinal fluid composition, the measured maximum and minimum equilibrium solubility values should represent the extremes of fasted intestinal solubility and provide a range. The seven drugs all display different solubility ranges and behaviours, a result also consistent with previous studies. The dose/solubility ratio for each measurement point can be plotted using the developability classification system to highlight individual drug behaviours. The lowest solubility represents a worst-case scenario which may be useful in risk-based quality by design biopharmaceutical calculations than the mid-point value. The method also permits a dose/solubility ratio frequency distribution determination for the solubility envelope which permits further risk-based refinement, especially where the drug crosses a classification boundary. This novel approach therefore provides greater in vitro detail with respect to possible biopharmaceutical performance in vivo and an improved ability to apply risk-based analysis to biopharmaceutical performance. Further studies will be required to expand the number of drugs measured and link the in vitro measurements to in vivo results.

Small scale in vitro method to determine a bioequivalent equilibrium solubility range for fasted human intestinal fluid.

Drug solubility is a key parameter controlling oral absorption, but intestinal solubility is difficult to assess in vitro. Human intestinal fluid (HIF) aspirates can be applied but they are variable, difficult to obtain and expensive. Simulated intestinal fluids (SIF) are a useful surrogate but multiple recipes are available and the optimum is unknown. A recent study characterised fasted HIF aspirates using a multi-dimensional approach and determined nine bioequivalent SIF media recipes that represented over ninety percent of HIF compositional variability. In this study these recipes have been applied to determine the equilibrium solubility of twelve drugs (naproxen, indomethacin, phenytoin, piroxicam, aprepitant, carvedilol, zafirlukast, tadalafil, fenofibrate, griseofulvin, felodipine, probucol) previously investigated using a statistical design of experiment (DoE) approach. The bioequivalent solubility measurements are statistically equivalent to the previous DoE, enclose literature solubility values in both fasted HIF and SIF, and the solubility range is less than the previous DoE. These results indicate that the system is measuring the same solubility space as literature systems with the lower overall range suggesting improved equivalence to in vivo solubility, when compared to DoEs. Three drugs (phenytoin, tadalafil and griseofulvin) display a comparatively narrow solubility range, a behaviour that is consistent with previous studies and related to the drugs' molecular structure and properties. This solubility behaviour would not be evident with single point solubility measurements. The solubility results can be analysed using a custom DoE to determine the most statistically significant factor within the media influencing solubility. This approach has a lower statistical resolution than a formal DoE and is not appropriate if determination of media factor significance for solubilisation is required. This study demonstrates that it is possible to assess the fasted intestinal equilibrium solubility envelope using a small number of bioequivalent media recipes obtained from a multi-dimensional analysis of fasted HIF. The derivation of the nine bioequivalent SIF media coupled with the lower measured solubility range indicate that the solubility results are more likely to reflect the fasted intestinal solubility envelope than previous DoE studies and highlight that intestinal solubility is a range and not a single value.

Manufacturing Considerations for the Development of Lipid Nanoparticles Using Microfluidics.

In the recent of years, the use of lipid nanoparticles (LNPs) for RNA delivery has gained considerable attention, with a large number in the clinical pipeline as vaccine candidates or to treat a wide range of diseases. Microfluidics offers considerable advantages for their manufacture due to its scalability, reproducibility and fast preparation. Thus, in this study, we have evaluated operating and formulation parameters to be considered when developing LNPs. Among them, the flow rate ratio (FRR) and the total flow rate (TFR) have been shown to significantly influence the physicochemical characteristics of the produced particles. In particular, increasing the TFR or increasing the FRR decreased the particle size. The amino lipid choice (cationic-DOTAP and DDAB; ionisable-MC3), buffer choice (citrate buffer pH 6 or TRIS pH 7.4) and type of nucleic acid payload (PolyA, ssDNA or mRNA) have also been shown to have an impact on the characteristics of these LNPs. LNPs were shown to have a high (>90%) loading in all cases and were below 100 nm with a low polydispersity index (≤0.25). The results within this paper could be used as a guide for the development and scalable manufacture of LNP systems using microfluidics.

Improving Consistency for a Mefenamic Acid Immediate Release Formulation.

The objective of this study was to develop an immediate release dose form containing 250 mg Mefenamic acid (MFA) presented as a crystalline solid dispersion in order to achieve improved consistency in drug release through a simplified formulation compared to a commercial product. An MFA-Soluplus®-Sorbitol polymer matrix was developed using an HME process based on rheological screening assays of physical mixtures. The physico-chemical properties of these formulations were assessed by thermal analysis, FTIR, mechanical testing and SEM image analysis, confirming the crystalline character and stable polymorphic form I of the API in the polymer matrix. A faster release and a significant improvement in consistency (±6%) of drug release was observed compared to a commercially available MFA product (±17%) (250 mg capsule). This study illustrates advantages of applying a structured development program aimed at retaining API physical properties in the final dosage form.

First-in-Human Study of AT13148, a Dual ROCK-AKT Inhibitor in Patients with Solid Tumors.

PURPOSE: AT13148 is an oral AGC kinase inhibitor, which potently inhibits ROCK and AKT kinases. In preclinical models, AT13148 has been shown to have antimetastatic and antiproliferative activity. PATIENTS AND METHODS: The trial followed a rolling six design during dose escalation. An intrapatient dose escalation arm to evaluate tolerability and a biopsy cohort to study pharmacodynamic effects were later added. AT13148 was administered orally three days a week (Mon-Wed-Fri) in 28-day cycles. Pharmacokinetic profiles were assessed using mass spectrometry and pharmacodynamic studies included quantifying p-GSK3ß levels in platelet-rich plasma (PRP) and p-cofilin and p-MLC2 levels in tumor biopsies. RESULTS: Fifty-one patients were treated on study. The safety of 5-300 mg of AT13148 was studied. Further, the doses of 120-180-240 mg were studied in an intrapatient dose escalation cohort. The dose-limiting toxicities included hypotension (300 mg), pneumonitis, and elevated liver enzymes (240 mg), and skin rash (180 mg). The most common side effects were fatigue, nausea, headaches, and hypotension. On the basis of tolerability, 180 mg was considered the maximally tolerated dose. At 180 mg, mean C max and AUC were 400 nmol/L and 13,000 nmol/L/hour, respectively. At 180 mg, ≥50% reduction of p-cofilin was observed in 3 of 8 posttreatment biopsies. CONCLUSIONS: AT13148 was the first dual potent ROCK-AKT inhibitor to be investigated for the treatment of solid tumors. The narrow therapeutic index and the pharmacokinetic profile led to recommend not developing this compound further. There are significant lessons learned in designing and testing agents that simultaneously inhibit multiple kinases including AGC kinases in cancer.

Rapid scale-up and production of active-loaded PEGylated liposomes.

Manufacturing of liposomal nanomedicines (e.g. Doxil®/Caelyx®) is a challenging and slow process based on multiple-vessel and batch processing techniques. As a result, the translation of these nanomedicines from bench to bedside has been limited. Microfluidic-based manufacturing offers the opportunity to address this issue, and de-risk the wider adoption of nanomedicines. Here we demonstrate the applicability of microfluidics for continuous manufacturing of PEGylated liposomes encapsulating ammonium sulfate (250 mM). Doxorubicin was subsequently active-loaded into these pre-formed liposomes. Critical process parameters and material considerations demonstrated to influence the liposomal product attributes included solvent selection and lipid concentration, flow rate ratio, and temperature and duration used for drug loading. However, the total flow rate did not affect the liposome product characteristics, allowing high production speeds to be adopted. The final liposomal product comprised of 80-100 nm vesicles (PDI < 0.2) encapsulating ≥ 90% doxorubicin, with matching release profiles to the innovator product and is stable for at least 6 months. Additionally, vincristine and acridine orange were active-loaded into these PEGylated liposomes (≥ 90% and ~100 nm in size) using the same process. These results demonstrate the ability to produce active-loaded PEGylated liposomes with high encapsulation efficiencies and particle sizes which support tumour targeting.

Small scale design of experiment investigation of equilibrium solubility in simulated fasted and fed intestinal fluid.

It is widely recognised that drug solubility within the gastrointestinal tract (GIT) differs from values determined in a simple aqueous buffer and to circumvent this problem measurement in biorelevant fluids is determined. Biorelevant fluids are complex mixtures of components (sodium taurocholate, lecithin, sodium phosphate, sodium chloride, pancreatin and sodium oleate) at various concentrations and pH levels to provide systems simulating fasted (FaSSIF) or fed (FeSSIF) intestinal media. Design of Experiment (DoE) studies have been applied to investigate FaSSIF and FeSSIF and indicate that a drug's equilibrium solubility varies over orders of magnitude, is influenced by the drug type and individual or combinations of media components, with some of these interactions being drug specific. Although providing great detail on the drug media interactions these studies are resource intensive requiring up to ninety individual experiments for FeSSIF. In this paper a low sample number or reduced DoE system has been investigated by restricting components with minimal solubility impact to a single value and only investigating variations in the concentrations of sodium taurocholate, lecithin, sodium oleate, pH and additionally in the case of fed media, monoglyceride. This reduces the experiments required to ten (FaSSIF) and nine (FeSSIF). Twelve poorly soluble drugs (Ibuprofen, Valsartan, Zafirlukast, Indomethacin, Fenofibrate, Felodipine, Probucol, Tadalafil, Carvedilol, Aprepitant, Bromocriptine and Itraconazole) were investigated and the results compared to published DoE studies and literature solubility values in human intestinal fluid (HIF), FaSSIF or FeSSIF. The solubility range determined by the reduced DoE is statistically equivalent to the larger scale published DoE results in over eighty five percent of the cases. The reduced DoE range also covers HIF, FaSSIF or FeSSIF literature solubility values. In addition the reduced DoE provides lowest measured solubility values that agree with the published DoE values in ninety percent of the cases. However, the reduced DoE only identified single and in some cases none of the major components influencing solubility in contrast to the larger published DoE studies which identified multiple individual components and component interactions. The identification of significant components within the reduced DoE was also dependent upon the drug and system under investigation. The study demonstrates that the lower experimental number reduces statistical power of the DoE to resolve the impact of media components on solubility. However, in a situation where only the solubility range is required the reduced DoE can provide the desired information, which will be of benefit during in vitro development studies. Further refinements are possible to extend the reduced DoE protocol to improve biorelevance and application into areas such as PBPK modelling.