Implementation of a fully integrated continuous manufacturing line for direct compression and coating at a commercial pharmaceutical facility - Part 1: Operational considerations and control strategy.

We implement a fully integrated continuous manufacturing (CM) line for direct compression and coating of a pharmaceutical oral solid dosage form in a commercial production facility. In this first paper of a two-part series, we describe process design and operational choices made to introduce CM using infrastructure originally intended for batch operations. Consistent with lean manufacturing principles, we select equipment, facilities, and novel process analytical technologies that meet production agility goals alongside an existing batch process. Choices address process risks, are aligned with existing quality systems, yet allow exploration of CM agility benefits in commercial operations. We outline how operating procedures, control schemes, and release criteria from the historical batch process are adapted for CM with modified lot and yield definitions based on patient demand. We devise a hierarchy of complementary controls including real-time process interrogation, predictive residence time distribution models of tablet concentration, real-time product release testing using automated tablet NIR spectroscopy, active rejection and diversion, and throughput-based sampling. Results from lots produced under normal operational conditions confirm our CM process provides assurance of product quality. Qualification strategies to achieve lot size flexibility aims are also described. Finally, we consider CM extensions to formulations with differing risk profiles. Further analysis of results for lots produced under normal operational conditions is provided in part 2 (Rosas et al., 2023).

Application of in vitro transmucosal permeability, dose number, and maximum absorbable dose for biopharmaceutics assessment during early drug development for intraoral delivery.

Intraoral (IO) administration is a unique route that takes advantage of transmucosal absorption in the oral cavity to deliver a drug substance locally or systemically. IO delivery can also enhance or enable oral administration, providing a better therapeutic benefit/safety risk profile for patient compliance. However, there are relatively few systematic biopharmaceutics assessments for IO delivery to date. Therefore, the goals of this study were to i) identify the most relevant in vitro permeability models as alternatives to porcine oral tissues (gold standard) for predicting human IO absorption and ii) establish guidelines for biopharmaceutics assessment during early drug development for IO delivery. Porcine kidney LLC-PK1 cells provided the strongest correlation of transmucosal permeability with porcine oral tissues followed by human Caco-2 cells. Furthermore, cultured human buccal tissues predicted high/low permeability classification and correlated well with porcine oral tissues, which are used for predicting clinical IO absorption. In the meantime, we introduced maximum absorbable dose and dose number in the oral cavity for IO delivery assessment as well as a decision tree to provide guidance for biopharmaceutics assessment during early drug development for IO delivery.

Decoupling the role of image size and calorie intake on gastric retention of swelling-based gastric retentive formulations: pre-screening in the dog model.

Gastric retention is postulated as an approach to improve bioavailability of compounds with narrow absorption windows. To elucidate the role of image size on gastric retention and pharmacokinetics, formulations with different image sizes and swelling kinetics but similar dissolution rates were designed and imaged in dogs. Diet had a clear effect, with increasing calorific intake prolonging retention in the dog model. In contrast to clinical observations, no obvious effect of image size on gastric retention was observed in the dog, with the larger gastric retentive (GR) and smaller controlled release (CR) formulations both demonstrating similar gastric emptying. Comparable pharmacokinetic profiles were observed for the two formulations, corroborating the imaging data and providing evidence of similar in vivo dissolution rates and dosage form integrity in the dog. Food, specifically meal composition, resulted in comparable enhancements in exposure in the dog and clinic due to prolonged gastric retention. However, differentiating retention based on image size in the dog was not feasible due to the smaller pyloric aperture compared to humans. This work illustrates that the dog is capable of determining the pharmacokinetic advantage of gastric retention relative to immediate release (IR) or CR formulations, however, has limited value in differentiating between CR and GR formulations.

Electrostatic interactions of monoclonal antibodies with subcutaneous tissue.

AIM: The majority of the subcutaneously injected monoclonal antibodies already on the market achieve 50-65% bioavailability, yet the fate of the portion that is lost remains unknown. This consistently incomplete systemic absorption affects the efficacy, safety and overall cost of the drug product. There are many potential factors that might influence the absorption, such as charge, hydrophobicity, formulation variables and the depth and volume of the injection. MATERIALS & METHODS: To explore the possibility that the charge of the injected protein and/or formulation components is partially responsible for drug retention at the subcutaneous site, an ex vivo study, where the monoclonal antibodies were exposed to homogenized rat subcutaneous tissue, was performed. RESULTS & CONCLUSION: It was found that positively charged monoclonal antibodies bind to subcutaneous tissue in a manner that is dependent on ionic strength and pH, suggesting the electrostatic nature of the interaction. As expected, saturation of both nonspecific and electrostatic subcutaneous binding sites was observed after incubation with highly concentrated monoclonal antibody solutions. Additionally, it was demonstrated using model proteins that electrostatic effects of buffer components depend on ionic strength of ions bearing opposite charge rather than total ionic strength of the solution. These results suggest that electrostatic interactions may play a role in absorption processes of positively charged therapeutic proteins after subcutaneous administration.

Pharmacokinetic and pharmacodynamic evaluation of a novel in situ forming poly(ethylene glycol)-based hydrogel for the controlled delivery of the camptothecins.

Inadequate drug delivery, due to problems associated with achieving constant therapeutic blood levels, has hampered the use of anticancer agents of the camptothecin (CPT) class. The objective of the current studies was to develop a depot delivery system for the water-soluble analog of CPT, topotecan (TPT). In this study, a 2-phase drug depot consisting of TPT-loaded liposomes entrapped in a poly(ethylene glycol) hydrogel was designed. Physically entrapped unaltered TPT displayed a rapid release rate from the hydrogel. Controlled release was demonstrated in vitro and in vivo from the 2-phase system with constant blood levels being achieved for several days in rats. Cytotoxicity and antitumor activity were also evaluated in rats inoculated with syngeneic MAT B III breast cancer cells. Rats treated with the liposome-loaded hydrogel displayed significantly longer tumor growth suppression and did not exhibit body weight loss compared to those treated with other delivery modes. These experiments constitute a proof-of-principle of the 2-phase depot concept and its potential value for enhancing safety and efficacy in chemotherapy.

Membrane transport of camptothecin: facilitation by human P-glycoprotein (ABCB1) and multidrug resistance protein 2 (ABCC2).

BACKGROUND: The purpose of the present study was to continue the investigation of the membrane transport mechanisms of 20-(S)-camptothecin (CPT) in order to understand the possible role of membrane transporters on its oral bioavailability and disposition. METHODS: The intestinal transport kinetics of CPT were characterized using Caco-2 cells, MDCKII wild-type cells and MDCKII cells transfected with human P-glycoprotein (PGP) (ABCB1) or human multidrug resistance protein 2 (MRP2) (ABCC2). The effects of drug concentration, inhibitors and temperature on CPT directional permeability were determined. RESULTS: The absorptive (apical to basolateral) and secretory (basolateral to apical) permeabilities of CPT were found to be saturable. Reduced secretory CPT permeabilities with decreasing temperatures suggests the involvement of an active, transporter-mediated secretory pathway. In the presence of etoposide, the CPT secretory permeability decreased 25.6%. However, inhibition was greater in the presence of PGP and of the breast cancer resistant protein inhibitor, GF120918 (52.5%). The involvement of additional secretory transporters was suggested since the basolateral to apical permeability of CPT was not further reduced in the presence of increasing concentrations of GF120918. To investigate the involvement of specific apically-located secretory membrane transporters, CPT transport studies were conducted using MDCKII/PGP cells and MDCKII/MRP2 cells. CPT carrier-mediated permeability was approximately twofold greater in MDCKII/PGP cells and MDCKII/MRP2 cells than in MDCKII/wild-type cells, while the apparent Km values were comparable in all three cell lines. The efflux ratio of CPT in MDCKII/PGP in the presence of 0.2 microM GF120918 was not completely reversed (3.36 to 1.49). However, the decrease in the efflux ratio of CPT in MDCKII/MRP2 cells (2.31 to 1.03) suggests that CPT efflux was completely inhibited by MK571, a potent inhibitor of the Multidrug Resistance Protein transporter family. CONCLUSIONS: The current results provide evidence that PGP and MRP2 mediate the secretory transport of CPT in vitro. However, the involvement of other transporters cannot be ruled out based on these studies. Since these transporters are expressed in the intestine, liver and kidney variations in their expression levels and/or regulation may be responsible for the erratic oral absorption and biliary excretion of CPT observed in human subjects.

A hydrogel prepared by in situ cross-linking of a thiol-containing poly(ethylene glycol)-based copolymer: a new biomaterial for protein drug delivery.

A new poly(ethylene glycol)-based copolymer containing multiple thiol (-SH) groups was cross-linked in situ to form a polymer hydrogel under mild conditions. No organic solvent, elevated temperature, or harsh pH is required in the formulation or patient administration processes, making it particularly useful for delivery of fragile therapeutics, such as proteins. The in vitro release of fluorescein-labeled bovine serum albumin and the in vivo release of the model proteins, erythropoietin, RANTES and three PEG-conjugated RANTES derivatives showed sustained release for 2-4 weeks and demonstrated prolonged biological activity of the released proteins in animals.

Enhancing the anticancer efficacy of camptothecin using biotinylated poly(ethylene glycol) conjugates in sensitive and multidrug-resistant human ovarian carcinoma cells.

BACKGROUND: Camptothecin (CPT) is an anticancer agent that kills cells by converting DNA topoisomerase I into a DNA-damaging agent. Although CPT and its derivatives are now being used to treat tumors in a variety of clinical protocols, the low water solubility of the drug and its unique pharmacodynamics and reactivity in vivo limit its delivery to cancer cells. To increase the anticancer efficacy of CPT a special drug delivery system is needed. PURPOSE: To synthesize a novel camptothecin-poly(ethylene glycol) conjugate (CPT-PEG) which includes biotin as a moiety to enhance nonspecific and/or targeted uptake via the sodium-dependent multivitamin transporter (SMVT) and to evaluate its anticancer activity and apoptosis induction. METHODS: CPT-PEG and CPT-PEG-biotin conjugates were synthesized and studied in vitro in A2780 sensitive and A2780/AD multidrug-resistant human ovarian carcinoma cells. Cytotoxicity, apoptosis induction, expression of genes encoding BCL-2 and apoptotic protease-activating factor 1 (APAF-1) proteins and caspases 3 and 9 as well as caspase activity were measured.RESULTS. We found that the conjugation of CPT with a simple linear PEG polymer led to a more than 12-fold enhancement of CPT toxicity in both sensitive and multidrug-resistant cells. Biotinylation of the PEG led to a further increase in CPT toxicity (5.2 times in sensitive and 2.1 times in multidrug-resistant cells) compared to the nonbiotinylated CPT-PEG conjugate. As a result, the cytotoxicity of the CPT-PEG-biotin conjugate increased more than 60 times in sensitive and almost 30 times in resistant cells, probably by enhancing nonspecific passive and/or SMVT-mediated uptake. In contrast, the same amounts of PEG and PEG-biotin conjugates without CPT did not induce cell death in either sensitive or resistant cells. Further analysis showed that the biotinylated CPT-PEG conjugate induced apoptosis more significantly than the same equivalent concentrations of free CPT or nonbiotinylated CPT-PEG. The enhancement of proapoptotic activity was achieved by the overexpression of genes encoding the APAF-1, and caspases 3 and 9, increasing caspase activity and simultaneously downregulating the BCL-2 gene. CONCLUSIONS: The results obtained demonstrate that the binding of CPT to PEG/PEG-biotin polymers increases its cytotoxicity, ability to induce apoptosis by the activation of caspase-dependent cell death signaling pathway and simultaneous suppression of antiapoptotic cellular defense. This suggests that the targeting approach utilizing transporters such as SMVT may substantially improve the delivery of CPT and its anticancer activity by enhancing cellular permeability and possibly retention of CPT.