Formulations For Clinical Trials Research Articles

Enhancing breadth and durability of humoral immune responses in non-human primates with an adjuvanted group 1 influenza hemagglutinin stem antigen

Seasonal influenza vaccines must be updated annually and suboptimally protect against strains mismatched to the selected vaccine strains. We previously developed a subunit vaccine antigen consisting of a stabilized trimeric influenza A group 1 hemagglutinin (H1) stem protein that elicits broadly neutralizing antibodies. Here, we further optimized the stability and manufacturability of the H1 stem antigen (H1 stem v2, also known as INFLUENZA G1 mHA) and characterized its formulation and potency with different adjuvants in vitro and in animal models. The recombinant H1 stem antigen (50 µg) was administered to influenza-naïve non-human primates either with aluminum hydroxide [Al(OH)3] + NaCl, AS01B, or SLA-LSQ formulations at week 0, 8 and 34. These SLA-LSQ formulations comprised of varying ratios of the synthetic TLR4 agonist ‘second generation synthetic lipid adjuvant’ (SLA) with liposomal QS-21 (LSQ). A vaccine formulation with aluminum hydroxide or SLA-LSQ (starting at a 10:25 µg ratio) induced HA-specific antibodies and breadth of neutralization against a panel of influenza A group 1 pseudoviruses, comparable with vaccine formulated with AS01B, four weeks after the second immunization. A formulation with SLA-LSQ in a 5:2 μg ratio contained larger fused or aggregated liposomes and induced significantly lower humoral responses. Broadly HA stem-binding antibodies were detectable for the entire period after the second vaccine dose up to week 34, after which they were boosted by a third vaccine dose. These findings inform about potential adjuvant formulations in clinical trials with an H1 stem-based vaccine candidate.

Nanomedicine for the Delivery of RNA in Cancer.

Simple SummaryCancer is a multifaceted, life-threatening, and genomically complex disease. The worldwide prevalence of cancer is so high that one in three people will develop cancer during their lifetime. Although the use of RNA therapy is promising to fight cancer, its efficient and safe delivery is still one of the significant challenges hampering its therapeutic application. Thus, the aim of the present review was to highlight the most recent developments in the field of nanomedicine RNA-associated therapies to fight cancer.The complexity, and the diversity of the different types of cancers allied to the tendency to form metastasis make treatment efficiency so tricky and often impossible due to the advanced stage of the disease in the diagnosis. In recent years, due to tremendous scientific breakthroughs, we have witnessed exponential growth in the elucidation of mechanisms that underlie carcinogenesis and metastasis. The development of more selective therapies made it possible to improve cancer treatment. Although interdisciplinary research leads to encouraging results, scientists still have a long exploration journey. RNA technology represents a promise as a therapeutic intervention for targeted gene silencing in cancer, and there are already some RNA-based formulations in clinical trials. However, the use of RNA as a therapeutic tool presents severe limitations, mainly related to its low stability and poor cellular uptake. Thus, the use of nanomedicine employing nanoparticles to encapsulate RNA may represent a suitable platform to address the major challenges hampering its therapeutic application. In this review, we have revisited the potential of RNA and RNA-associated therapies to fight cancer, also providing, as support, a general overview of nanoplatforms for RNA delivery.

Physiologically based pharmacokinetic modeling and simulations to inform dissolution specifications and clinical relevance of release rates on elagolix exposure

The aim of this analysis was to use a physiologically based pharmacokinetic (PBPK) model to predict the impact of changes in dissolution rates on elagolix exposures and define clinically relevant acceptance criteria for dissolution. Varying in vitro dissolution profiles were utilized in a PBPK model to describe the absorption profiles of elagolix formulations used in Phase 3 clinical trials and for the to be marketed commercial formulations. Single dose studies of 200 mg elagolix formulations were used for model verification under fasted conditions. Additional dissolution scenarios were evaluated to assess the impact of dissolution rates on elagolix exposures. Compared to the Phase 3 clinical trial formulation, sensitivity analysis on dissolution rates suggested that a hypothetical scenario of ∼75% slower dissolution rate would result in 14% lower predicted elagolix plasma exposures, however, the predicted exposures are still within the bioequivalence boundaries of 0.8–1.25 for both Cmax and AUC. A clinically verified PBPK model of elagolix was utilized to evaluate the impact of wider dissolution specifications on elagolix plasma exposures. The simulation results indicated that a slower in vitro dissolution profile, would not have a clinically significant impact on elagolix exposures. These model results informed the setting of wider dissolution specifications without requiring in vivo studies.

Improving Curcumin Bioavailability: Current Strategies and Future Perspectives.

Curcumin possesses a plethora of interesting pharmacological effects. Unfortunately, it is also characterized by problematic drug delivery and scarce bioavailability, representing the main problem related to the use of this compound. Poor absorption, fast metabolism, and rapid systemic clearance are the most important factors contributing to low curcumin levels in plasma and tissues. Accordingly, to overcome these issues, numerous strategies have been proposed and are investigated in this article. Due to advances in the drug delivery field, we describe here the most promising strategies for increasing curcumin bioavailability, including the use of adjuvant, complexed/encapsulated curcumin, specific curcumin formulations, and curcumin nanoparticles. We analyze current strategies, already available in the market, and the most advanced technologies that can offer a future perspective for effective curcumin formulations. We focus the attention on the effectiveness of curcumin-based formulations in clinical trials, providing a comprehensive summary. Clinical trial results, employing various delivery methods for curcumin, showed that improved bioavailability corresponds to increased therapeutic efficacy. Furthermore, advances in the field of nanoparticles hold great promise for developing curcumin-based complexes as effective therapeutic agents. Summarizing, suitable delivery methods for this polyphenol will ensure the possibility of using curcumin-derived formulations in clinical practice as preventive and disease-modifying therapeutics.

Biomaterials-based formulations and surfaces to combat viral infectious diseases.

Rapidly growing viral infections are potent risks to public health worldwide. Accessible virus-specific antiviral vaccines and drugs are therapeutically inert to emerging viruses, such as Zika, Ebola, and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Therefore, discovering ways to prevent and control viral infections is among the foremost medical challenge of our time. Recently, innovative technologies are emerging that involve the development of new biomaterial-based formulations and surfaces endowed with broad-spectrum antiviral properties. Here, we review emerging biomaterials technologies for controlling viral infections. Relevant advances in biomaterials employed with nanotechnology to inactivate viruses or to inhibit virus replication and further their translation in safe and effective antiviral formulations in clinical trials are discussed. We have included antiviral approaches based on both organic and inorganic nanoparticles (NPs), which offer many advantages over molecular medicine. An insight into the development of immunomodulatory scaffolds in designing new platforms for personalized vaccines is also considered. Substantial research on natural products and herbal medicines and their potential in novel antiviral drugs are discussed. Furthermore, to control contagious viral infections, i.e., to reduce the viral load on surfaces, current strategies focusing on biomimetic anti-adhesive surfaces through nanostructured topography and hydrophobic surface modification techniques are introduced. Biomaterial surfaces functionalized with antimicrobial polymers and nanoparticles against viral infections are also discussed. We recognize the importance of research on antiviral biomaterials and present potential strategies for future directions in applying these biomaterial-based approaches to control viral infections and SARS-CoV-2.

A head-to-head Caco-2 assay comparison of the mechanisms of action of the intestinal permeation enhancers: SNAC and sodium caprate (C10)

Salcaprozate sodium (SNAC) and sodium caprate (C10) are the two leading intestinal permeation enhancers (PEs) in oral peptide formulations in clinical trials. There is debate over their mechanism of action on intestinal epithelia. The aims were: (i) to compare their effects on the barrier function by measuring transepithelial electrical resistance (TEER), permeability of FITC-4000 (FD4) across Caco-2 monolayers, and on immunohistochemistry of tight junction (TJ)-associated proteins; and (ii) to compare cellular parameters using conventional end-point cytotoxicity assays and quantitative high content analysis (HCA) of multiple sub-lethal parameters in Caco-2 cells. C10 (8.5 mM) reversibly reduced TEER and increased FD4 permeability across monolayers, whereas SNAC had no effects on either parameter except at cytotoxic concentrations. C10 exposure induced reorganization of three TJ proteins, whereas SNAC only affected claudin-5 localization. High concentrations of C10 and SNAC were required to cause end-point toxicology changes in vitro. SNAC was less potent than C10 at inducing lysosomal and nuclear changes and plasma membrane perturbation. In parallel, HCA revealed that both agents displayed detergent-like features that reflect initial membrane fluidization followed by changes in intracellular parameters. In conclusion, FD4 permeability increases in monolayers in response to C10 were in the range of concentrations that altered end-point cytotoxicity and HCA parameters. For SNAC, while HCA parameters were also altered in a similar overall pattern as C10, they did not lead to increased paracellular flux. These assays show that both agents are primarily surfactants, but C10 has additional TJ-opening effects. While these in vitro assays illucidate their epithelial mechanism of action, clinical experience suggests that they over-estimate their toxicology in the dynamic intestinal environment.

Polymeric micelles for the delivery of poorly soluble drugs: From nanoformulation to clinical approval.

Over the last three decades, polymeric micelles have emerged as a highly promising drug delivery platform for therapeutic compounds. Particularly, poorly soluble small molecules with high potency and significant toxicity were encapsulated in polymeric micelles. Polymeric micelles have shown improved pharmacokinetic profiles in preclinical animal models and enhanced efficacy with a superior safety profile for therapeutic drugs. Several polymeric micelle formulations have reached the clinical stage and are either in clinical trials or are approved for human use. This furthers interest in this field and underscores the need for additional learning of how to best design and apply these micellar carriers to improve the clinical outcomes of many drugs. In this review, we provide detailed information on polymeric micelles for the solubilization of poorly soluble small molecules in topics such as the design of block copolymers, experimental and theoretical analysis of drug encapsulation in polymeric micelles, pharmacokinetics of drugs in polymeric micelles, regulatory approval pathways of nanomedicines, and current outcomes from micelle formulations in clinical trials. We aim to describe the latest information on advanced analytical approaches for elucidating molecular interactions within the core of polymeric micelles for effective solubilization as well as for analyzing nanomedicine's pharmacokinetic profiles. Taking into account the considerations described within, academic and industrial researchers can continue to elucidate novel interactions in polymeric micelles and capitalize on their potential as drug delivery vehicles to help improve therapeutic outcomes in systemic delivery.

Platinum-Based Antitumor Drugs and Their Liposomal Formulations in Clinical Trials

Platinum-based antitumor drugs are the most common drugs in modern oncology practice. They are used for the treatment of a large number of malignant tumors of different etiology. However, their low specificity and high systemic toxicity significantly reduce the efficacy of chemotherapy. With the goal of improving pharmacological properties of platinum-based drugs, an intense search for supramolecular drug delivery systems is being carried out in many laboratories. This review focuses on liposomal drugs, which are currently at different stages of clinical trials. In the first part of the review we described modern concepts on the action of Pt-based drugs. Information about the drugs used in clinical practice as well as data about some new promising compounds is also provided. Also, justification of the rationale for the creation of liposomal drug delivery systems is given, and six Pt-containing liposomal formulations under clinical trials are reviewed, including the published data on the composition, how they act, and the results of preclinical and clinical trials. Several examples of new Pt-based liposomal formulations are given. The prospects of the liposomal platform development for platinum-based drugs are discussed.

Liposomal therapies in oncology: does one size fit all?

Liposomal therapies opened the chapter of nanomedicine, in 1995, with the approval of liposomal doxorubicin (Doxil®) for the treatment of numerous types of cancer. For the first time, liposomes permitted the employment of potent chemotherapeutic agents with improved pharmacokinetic and pharmacodynamic profiles, with less undesired side effects. Liposomal therapies allow the drug encapsulation and more selective delivery in the tumor bed, particularly due to the enhanced permeation and retention effect. These unique characteristics explain why liposomal therapies are being increasingly considered as alternatives in cancer therapy and represent an important research field with the recent approval for clinical use and emergent formulations in clinical trials. Even so, the response rate of liposomal therapies varies between 5 and 71%, and they are still indistinguishably given to every patient. As already well-demonstrated for conventional chemotherapies and targeted therapies, there is also the need for predictive biomarkers that allow a better use of liposomal drugs, with higher patient quality of life. Our aim in this review was to address the approved liposomal therapies and to summarize the information concerning possible predictive markers of response in their various clinical applications, to personalize each patient treatment and maximize its efficacy.

Micelles As Delivery System for Cancer Treatment.

Micelles are nanoparticles formed by the self-assembly of amphiphilic block copolymers in certain solvents above concentrations called critical micelle concentration (CMC). Micelles are used in different fields like food, cosmetics, medicine, etc. These nanosized delivery systems are under spotlight in the recent years with new achievements in terms of their in vivo stability, ability to protect entrapped drug, release kinetics, ease of cellular penetration and thereby increased therapeutic efficacy. Drug loaded micelles can be prepared by dialysis, oil-in-water method, solid dispersion, freezing, spray drying, etc. The aim of this review is to give an overview of the research on micelles (in vitro, in vivo and clinical) as delivery system for cancer treatment. Passive targeting is one route for accumulation of nanosized micellar drug formulations. Many research groups from both academia and industry focus on developing new strategies for improving the therapeutic efficacy of micellar systems (active targeting to the tumor site, designing multidrug delivery systems for overcoming multidrug resistance or micelles formed by prodrug conjugates, etc). There is only one micellar drug formulation in South Korea that has reached clinical practice. However, there are many untargeted anticancer drug loaded micellar formulations in clinical trials, which have potential for use in clinics. Many more products are expected to be on the market in the near future.

Effects of surfactant-based permeation enhancers on mannitol permeability, histology, and electrogenic ion transport responses in excised rat colonic mucosae

Surfactant-based intestinal permeation enhancers (PEs) are constituents of several oral macromolecule formulations in clinical trials. This study examined the interaction of a test panel of surfactant-based PEs with isolated rat colonic mucosae mounted in Ussing chambers in an attempt to determine if increases in transepithelial permeability can be separated from induction of mucosal perturbation. The aim was to assess the effects of PEs on (i) apparent permeability coefficient (Papp) of [14C]-mannitol (ii) histology score and (iii) short-circuit current (ΔIsc) responses to a cholinomimetic (carbachol, CCh). Enhancement ratio increases for Papp values followed the order: C10 > C9 = C11:1 > a bile salt blend > sodium choleate > sucrose laurate > Labrasol® >C12E8 > C12 > Cremophor® A25 > C7 > sucrose stearate > Kolliphor® HS15 > Kolliphor® TPGS. Exposures that increased the Papp by ≥2-fold over 120 min were accompanied by histological damage in 94% of tissues, and by a decreased ΔIsc response to CCh in 83%. A degree of separation between the increased Papp of [14C]-mannitol and histological damage and diminution of the ΔIsc response to CCh was observed at selected concentrations of Labrasol®. Overall, this surfactant-based PE selection caused transcellular perturbation at similar concentrations to those that enhanced permeability.

Microfluidic Production and Application of Lipid Nanoparticles for Nucleic Acid Transfection.

Lipid nanoparticles (LNPs) are established in the biopharmaceutical industry for efficient encapsulation and cytosolic delivery of nucleic acids for potential therapeutics, with several formulations in clinical trials. The advantages of LNPs can also be applied in basic research and discovery with a microfluidic method of preparation now commercially available that allows preparations to be scaled down to quantities appropriate for cell culture. These preparations conserve expensive nucleic acids while maintaining the particle characteristics that have made LNPs successful in later stages of genetic medicine development. Additionally, this method and the resulting LNPs are seamlessly scalable to quantities appropriate for in vivo models and development of nucleic acid therapeutics.The present work describes the methodology for preparing LNPs loaded with siRNA, mRNA or plasmids using a commercially available microfluidic instrument and an accompanying transfection kit. Guidelines for application to cultured cells in a well-plate format are also provided.

Eliminating Schistosomes through Vaccination: What are the Best Immune Weapons?

The successful development of vaccines depends on the knowledge of the immunological mechanisms associated with the elimination of the pathogen. In the case of schistosomes, its complex life cycle and the mechanisms developed to evade host immune system, turns the development of a vaccine against the disease into a very difficult task. Identifying the immunological effector mechanisms involved in parasite attrition and the major targets for its response is a key step to formulate an effective vaccine. Recent studies have described some promising antigens to compose a subunit vaccine and have pointed to some immune factors that play a role in parasite elimination. Here, we review the immune components and effector mechanisms associated with the protective immunity induced by those vaccine candidates and the lessons we have learned from the studies of the acquired resistance to infection in humans. We will also discuss the immune factors that correlate with protection and therefore could help to evaluate those vaccine formulations in clinical trials.

Development of lipid nanoparticle formulations of siRNA for hepatocyte gene silencing following subcutaneous administration

Recently developed lipid nanoparticle (LNP) formulations of siRNA have proven to be effective agents for hepatocyte gene silencing following intravenous administration with at least three LNP–siRNA formulations in clinical trials. The aim of this work was to develop LNP–siRNA systems for hepatocyte gene silencing that can be administered subcutaneously (s.c.). Three parameters were investigated, namely LNP size, residence time of the polyethylene glycol (PEG)-lipid coating and the influence of hepatocyte-specific targeting ligands. LNP sizes were varied over the range of 30 to 115nm in diameter and PEG-lipid that dissociates rapidly (PEG-DMG) and slowly (PEG-DSG) were employed. In mice, results show that large (~80nm) LNP exhibited limited accumulation in the liver and poor Factor VII (FVII) gene silencing at 1mg siRNA/kg body weight. Conversely, small (~30nm) LNP systems showed maximal liver accumulation yet still had minimal activity. Interestingly, intermediate size (~45nm) LNP containing PEG-DSG exhibited nearly equivalent liver accumulation as the smaller systems following s.c. administration but reduced FVII levels by 80% at 1mg siRNA/kg body weight. Smaller systems (~35nm diameter) containing either PEG-DMG or PEG-DSG were less active; however addition of 0.5mol.% of a GalNAc-PEG lipid to these smaller systems improved activity to levels similar to that observed for the ~45nm diameter systems. In summary, this work shows that appropriately designed LNP–siRNA systems can result in effective hepatocyte gene silencing following s.c administration.

ChemInform Abstract: Smart Drug Delivery Systems: From Fundamentals to the Clinic

AbstractReview: 244 refs.

Paclitaxel Nano-Delivery Systems: A Comprehensive Review

Paclitaxel is one of the most effective chemotherapeutic drugs ever developed and is active against a broad range of cancers, such as lung, ovarian, and breast cancers. Due to its low water solubility, paclitaxel is formulated in a mixture of Cremophor EL and dehydrated ethanol (50:50, v/v) a combination known as Taxol. However, Taxol has some severe side effects related to Cremophor EL and ethanol. Therefore, there is an urgent need for the development of alternative Taxol formulations. The encapsulation of paclitaxel in biodegradable and non-toxic nano-delivery systems can protect the drug from degradation during circulation and in-turn protect the body from toxic side effects of the drug thereby lowering its toxicity, increasing its circulation half-life, exhibiting improved pharmacokinetic profiles, and demonstrating better patient compliance. Also, nanoparticle-based delivery systems can take advantage of the enhanced permeability and retention (EPR) effect for passive tumor targeting, therefore, they are promising carriers to improve the therapeutic index and decrease the side effects of paclitaxel. To date, paclitaxel albumin-bound nanoparticles (Abraxane®) have been approved by the FDA for the treatment of metastatic breast cancer and non-small cell lung cancer (NSCLC). In addition, there are a number of novel paclitaxel nanoparticle formulations in clinical trials. In this comprehensive review, several types of developed paclitaxel nano-delivery systems will be covered and discussed, such as polymeric nanoparticles, lipid-based formulations, polymer conjugates, inorganic nanoparticles, carbon nanotubes, nanocrystals, and cyclodextrin nanoparticles.

1574P - Bioequivalence and Absolute Bioavailability of a Single Oral Dose of Two Formulations of 0.75 Mg Palonosetron in Healthy Volunteers (HV)

ABSTRACT Background Palonosetron HCl (PALO) is a potent 5-HT3 receptor antagonist (5HT3-RA) with a stronger receptor binding affinity and unique mechanism of action compared to older 5HT3-RAs. The primary objectives were to investigate the bioequivalence of two oral PALO 0.75 mg formulation(s) (F(s)) b1 and b2 and their absolute bioavailability when compared to b3 PALO 0.75 mg i.v F (see table for: b1, b2 and b3). Secondary objective was to evaluate the safety profile of the three PALO F(s). Methods 36 male and female hv were randomized in this open-label, 3-treatment, cross-over study. 30 subjects received treatments b1, b2 and b3 in a 3- period randomized order (14-day interval set between periods). Treatments b1 and b2 were analyzed for bioequivalence by ANOVA CVs and defined bioequivalent if the 90% confidence intervals (CI) for the AUC0-∞, AUC0-t and Cmax treatment ratios lie within 80% to 125% range CI Absolute Ratios. Absolute bioavailability of the oral F(s) was calculated using the extent of exposure measure AUC0-∞ and AUC0-t. Results The oral 0.75 mg F(s) b1 and b2 showed bioequivalence in terms of rate and extent of absorption: The 90% CI for PALO AUC0-t, AUC0-∞ and Cmax are 97.91% to 109.47%, 96.47% to 107.76% and 93.40% to 103.59%, respectively. All CI's are well within the 80-125% equivalence limits. The absolute bioavailability of both oral and the i.v. F(s), calculated using AUC0-t is essentially not different (97% for b1, and 99% for b2 F(s)). All study F(s) were safe and well tolerated: constipation and headache were the main adverse events. No clinical influence was observed on safety laboratory or vital signs variability. No drop out was due to safety issue. Conclusions The study has shown bioequivalence of two PALO 0.75 mg oral formulations. No significant changes were observed in the absolute bioavailability between the two oral formulations and the i.v. 0.75 mg. All tested formulations showed a good safety profile. Disclosure D. Voisin: The Author is an Helsinn Employee, C. Giuliano: The Author is an Helsinn Employee, All other authors have declared no conflicts of interest. Table: 1574P . Pharmacokinetic parameter palonosetron Ratio Point estimate Test/Ref. ratio 90% Confidence Interval AUC(0-t) [ng h/L] b1 / b3 b2 / b3 96.56 99.88 91.47 - 101-93 94.61 - 105.44 AUC(0-inf) [ng h/L] b1 / b3 b2 / b3 97.26 98.99 92.33 - 102.45 93.98 - 104.27 Cmax [ng/L] b1 / b3 b2 / b3 72.60 71.71 67.01 - 78.67 66.18 - 77.70 b1: oral administration of 0.75 mg palonosetron as a softgel capsule (formulation A, clinical trial formulation). b2: oral administration of 0.75 mg palonosetron as a softgel capsule (formulation B, Proposed commercial formulation). b3: intravenous administration of 0.75 mg palonosetron (3 x 0.25 mg i.v. Aloxi® - reference for the bioavailability analysis). ANOVA and 90% CIs for l0g-transformed.

Aluminum Adjuvant Dose Guidelines in Vaccine Formulation for Preclinical Evaluations

Aluminum (Al) salt-based adjuvants are present in a large variety of licensed vaccines and their use is widely considered for formulations in clinical trials. Although the regulatory agencies have clearly stated the acceptable levels of Al salts in vaccines for human use, there are no general indications for preclinical research. This brief commentary reviews the current status of Al concentrations in licensed vaccines, the related potential toxicity in preclinical species, and proposes a general guideline for selection of suitable Al salt levels in preclinical models, focusing on the formulation development for recombinant protein antigens. A table with conversion factors is included in order to provide a tool for calculation of doses with different Al salts.

Polymeric Nano-Assemblies as Emerging Delivery Carriers for Therapeutic Applications: A Review of Recent Patents

Multi-morphologic polymer nano-assemblies, such as micelles and vesicles, have been intensively studied recently by scientists in the multidisciplinary fields for their promising applications in bioengineering, biomedicine and pharmaceutics. With the success of several micellar formulations in clinical trials, more and more therapeutics based on polymer assemblies are on the pipeline for clinical applications. The current review summarizes some recent patents on the polymer nano-assemblies including micelles and vesicles, with the focus on drug delivery and gene therapy. For the lack of updated patents, the selected progress made most recently in this field has been presented based on the newly published articles. The future development in this active and exciting field has been discussed as well. Because of the huge literature of scientific papers on delivery systems based on polymer assemblies in recent years, this review attempts to limit the specific examples to those that are currently in clinical trials. Accordingly, it is impossible to properly credit all the scientists in this field. The author apologizes in advance for all omissions.

Stability studies with a high-performance liquid chromatographic method for the determination of a new anthracycline analogue, 3′-deamino-3′-[2-( S)-methoxy-4-morpholio]doxorubicin (FCE 23762), in the final drug formulation

A high-performance liquid chromatographic method was studied to optimize the separation of FCE 23762, a new antitumour agent, from both synthetic impurities and degradation products having very similar molecular structures. The main problems faced in the analytical method development using the most common reversed-phase columns available arose from the presence of analytical peaks with poor symmetry, a long analysis time and the separation between FCE 23762 and its R-isomer, which was often unsuitable for the correct determination of the drug substance. The use of a new stationary phase, Zorbax R x-C8, together with a suitable mobile phase resulted in a good separation between the diasteromers, with satisfactory peak symmetry and run time. The method permitted the study of the stability of the drug substance in formulations for clinical trials.