Development of a Sustained Antiplaque and Antimicrobial Chewing Gum of a Decapeptide.

The objective of this paper was to design a chewing gum formulation delivery system in situations where typical dental hygiene practice is not practical. Thus, an analog of decapeptide KSL (KSL-W), known to possess antimicrobial and antiplaque activity, was incorporated into a chewing gum formulation containing cetylpyridinium chloride (CPC). The effect of the excipients, xylitol, and peppermint oil on active ingredients in vitro release was also assessed. Gum formulations were prepared with different excipient parameters, including heating xylitol and gum base at 65 or 85°C, using ground and unground xylitol, and the addition of 1.5, 3, and 7% peppermint oil, to determine the effect of these changes on the in vitro release of KSL-W and CPC using a chewing machine. The antimicrobial and antiplaque activities of solutions released from chewed gum formulation as well as prepared standard solutions with different concentrations were tested against placebo. The optimal temperature to avoid crystallization of xylitol during preparation was 65°C. Grinding xylitol to 104.5 µm improved release of active ingredients as compared to commercially unground xylitol. Peppermint oil had opposite effects on release of KSL-W and CPC. Peppermint oil at 1.5% was determined to be suitable (91 and 88% of KSL-W and CPC released, respectively, after 40 min). The gum formulation illustrated good sustained release of KSL-W and CPC with antibacterial and antiplaque activities after chewing. An effective antimicrobial and antiplaque chewing gum formulation was developed. This formulation has the potential to overcome oral hygiene issues in those unable to follow normal dental protocols.

A short term quality control tool for biodegradable microspheres.

Accelerated in vitro release testing methodology has been developed as an indicator of product performance to be used as a discriminatory quality control (QC) technique for the release of clinical and commercial batches of biodegradable microspheres. While product performance of biodegradable microspheres can be verified by in vivo and/or in vitro experiments, such evaluation can be particularly challenging because of slow polymer degradation, resulting in extended study times, labor, and expense. Three batches of Leuprolide poly(lactic-co-glycolic acid) (PLGA) microspheres having varying morphology (process variants having different particle size and specific surface area) were manufactured by the solvent extraction/evaporation technique. Tests involving in vitro release, polymer degradation and hydration of the microspheres were performed on the three batches at 55°C. In vitro peptide release at 55°C was analyzed using a previously derived modification of the Weibull function termed the modified Weibull equation (MWE). Experimental observations and data analysis confirm excellent reproducibility studies within and between batches of the microsphere formulations demonstrating the predictability of the accelerated experiments at 55°C. The accelerated test method was also successfully able to distinguish the in vitro product performance between the three batches having varying morphology (process variants), indicating that it is a suitable QC tool to discriminate product or process variants in clinical or commercial batches of microspheres. Additionally, data analysis utilized the MWE to further quantify the differences obtained from the accelerated in vitro product performance test between process variants, thereby enhancing the discriminatory power of the accelerated methodology at 55°C.

Reprogramming the tumor microenvironment to improve the efficacy of cancer immunotherapies.

The immunotherapeutic approaches based on checkpoint inhibitors, tumor vaccination, immune cell-based therapy, and cytokines were developed to engage the patient's immune system against cancer and better survival of them. While potent, however, preclinical and clinical data have identified that abnormalities in the tumor microenvironment (TME) can affect the efficacy of immunotherapies in some cancers. It is therefore imperative to develop new therapeutic interventions that will enable to overcome tumor-supportive TME and restrain anti-tumor immunity in patients that acquire resistance to current immunotherapies. Therefore, recognition of the essential nature of the tolerogenic TME may lead to a shift from the immune-suppressive TME to an immune-stimulating phenotype. Here, we review the composition of the TME and its effect on tumor immunoediting and then present how targeted monotherapy or combination therapies can be employed for reprogramming educated TME to improve current immunotherapies outcomes or elucidate potential therapeutic targets.

Evaluation of the Wheat Germ Oil Topical Formulations for Wound Healing Activity in Rats.

Wheat Germ Oil (WGO), the flour-milling by-product of wheat has essential constituents for skin health care as vitamin E, B-complex, squalene and unsaturated fatty acids. Incorporate WGO into polymers of the cream and ointment bases and evaluate the wound healing potential of these WGO formulations in the rat-animal model. <b>Materials and Methods:</b> WGO creams and ointments were prepared in two concentrations, 10 and 20% and evaluated for storage stability, homogeneity and compatibility using Fourier Transform Infrared (FT-IR) spectrometry. An amount of 0.5 g of the WGO formulations was applied daily to the injured area of the rats back. Wounds were observed for any clinical changes and healing compared to the control animal group. <b>Results:</b> The WGO was compatible with the cream and ointment bases and physically stables over 60 days of storage. The formulations of WGO have induced dose-dependent wound healing properties however the ointment formulations were demonstrating wound healing activity significantly better than the creams at all the intervals of the treatment. Within three weeks, 20% WGO ointment has induced a 90%reduction in the wound size diameter. Also, wounds recovered by 50% in 10 and 14 days of treatment with 20% WGO ointment and cream, respectively. <b>Conclusion:</b> The results revealed that WGO is a potential wound-healing agent from the scope that WGO is a common cosmetic ingredient and available at affordable prices.

Development of a peptide-containing chewing gum as a sustained release antiplaque antimicrobial delivery system.

The objective of this study was to characterize the stability of KSL-W, an antimicrobial decapeptide shown to inhibit the growth of oral bacterial strains associated with caries development and plaque formation, and its potential as an antiplaque agent in a chewing gum formulation. KSL-W formulations with or without the commercial antibacterial agent cetylpyridinium chloride (CPC) were prepared. The release of KSL-W from the gums was assessed in vitro using a chewing gum apparatus and in vivo by a chew-out method. A reverse-phase high-performance liquid chromatography method was developed for assaying KSL-W. Raw material stability and temperature and pH effects on the stability of KSL-W solutions and interactions of KSL-W with tooth-like material, hydroxyapatite discs, were investigated. KSL-W was most stable in acidic aqueous solutions and underwent rapid hydrolysis in base. It was stable to enzymatic degradation in human saliva for 1 hour but was degraded by pancreatic serine proteases. KSL-W readily adsorbed to hydroxyapatite, suggesting that it will also adsorb to the teeth when delivered to the oral cavity. The inclusion of CPC caused a large increase in the rate and extent of KSL-W released from the gums. The gum formulations displayed promising in vitro/in vivo release profiles, wherein as much as 90% of the KSL-W was released in a sustained manner within 30 minutes in vivo. These results suggest that KSL-W possesses the stability, adsorption, and release characteristics necessary for local delivery to the oral cavity in a chewing gum formulation, thereby serving as a novel antiplaque agent.

Chewing gum of antimicrobial decapeptide (KSL) as a sustained antiplaque agent: preformulation study.

The purpose of this study was to investigate the potential of KSL, an antimicrobial decapeptide, which has been shown to inhibit the growth of oral bacterial strains associated with caries development and plaque formation, to act as an antiplaque agent in a chewing gum formulation. A reversed-phase high-performance liquid chromatography method was developed for KSL and found to be stability-indicating. KSL was stable in acetate buffer at pH 4 and artificial saliva. On the affinity of KSL to tooth-like materials, the KSL showed favorable interaction with hydroxyapatite discs pretreated with human saliva. A chewing gum formulation of KSL was prepared based on conventional procedures and the release of KSL from the gum was studied in vitro using the chewing apparatus and in vivo by a chew-out method. The gum formulations showed promising in vitro/in vivo release profiles, in which 70-80% KSL was released in a sustained manner over 20 min of chewing time. This study suggests that KSL in a gum formulation is suitable for the delivery in the oral cavity, thereby serving as a novel antiplaque agent.

A model-dependent approach to correlate accelerated with real-time release from biodegradable microspheres.

The purpose of this study was to determine the feasibility of applying accelerated in vitro release testing to correlate or predict long-term in vitro release of leuprolide poly(lactide-co-glycolide) microspheres. Peptide release was studied using a dialysis technique at 37 degrees C and at elevated temperatures (50 degrees C-60 degrees C) in 0.1M phosphate buffered saline (PBS) pH 7.4 and 0.1M acetate buffer pH 4.0. The data were analyzed using a modification of the Weibull equation. Peptide release was temperature dependent and complete within 30 days at 37 degrees C and 3 to 5 days at the elevated temperatures. In vitro release profiles at the elevated temperatures correlated well with release at 37 degrees C. The shapes of the release profiles at all temperatures were similar. Using the modified Weibull equation, an increase in temperature was characterized by an increase in the model parameter, alpha, a scaling factor for the apparent rate constant. Complete release at 37 degrees C was shortened from approximately 30 days to 5 days at 50 degrees C, 3.5 days at 55 degrees C, 2.25 days at 60 degrees C in PBS pH 7.4, and 3 days at 50 degrees C in acetate buffer pH 4.0. Values for the model parameter beta indicated that the shape of the release profiles at 55 degrees C in PBS pH 7.4 (2.740) and 50 degrees C in 0.1M acetate buffer pH 4.0 (2.711) were similar to that at 37 degrees C (2.677). The E(a) for hydration and erosion were determined to be 42.3 and 19.4 kcal/mol, respectively. Polymer degradation was also temperature dependent and had an E(a) of 31.6 kcal/mol. Short-term in vitro release studies offer the possibility of correlation with long-term release, thereby reducing the time and expense associated with long-term studies. Accelerated release methodology could be useful in the prediction of long-term release from extended release microsphere dosage forms and may serve as a quality control tool for the release of clinical or commercial batches.

IVIVC from Long Acting Olanzapine Microspheres.

In this study, four PLGA microsphere formulations of Olanzapine were characterized on the basis of their in vitro behavior at 37°C, using a dialysis based method, with the goal of obtaining an IVIVC. In vivo profiles were determined by deconvolution (Nelson-Wagner method) and using fractional AUC. The in vitro and in vivo release profiles exhibited the same rank order of drug release. Further, in vivo profiles obtained with both approaches were nearly superimposable, suggesting that fractional AUC could be used as an alternative to the Nelson-Wagner method. A comparison of drug release profiles for the four formulations revealed that the in vitro profile lagged slightly behind in vivo release, but the results were not statistically significant (P < 0.0001). Using the four formulations that exhibited different release rates, a Level A IVIVC was established using the deconvolution and fractional AUC approaches. A nearly 1 : 1 correlation (R (2) > 0.96) between in vitro release and in vivo measurements confirmed the excellent relationship between in vitro drug release and the amount of drug absorbed in vivo. The results of this study suggest that proper selection of an in vitro method will greatly aid in establishing a Level A IVIVC for long acting injectables.

Development of Risperidone PLGA Microspheres.

The aim of this study was to design and evaluate biodegradable PLGA microspheres for sustained delivery of Risperidone, with an eventual goal of avoiding combination therapy for the treatment of schizophrenia. Two PLGA copolymers (50 : 50 and 75 : 25) were used to prepare four microsphere formulations of Risperidone. The microspheres were characterized by several in vitro techniques. In vivo studies in male Sprague-Dawley rats at 20 and 40 mg/kg doses revealed that all formulations exhibited an initial burst followed by sustained release of the active moiety. Additionally, formulations prepared with 50 : 50 PLGA had a shorter duration of action and lower cumulative AUC levels than the 75 : 25 PLGA microspheres. A simulation of multiple dosing at weekly or 15-day regimen revealed pulsatile behavior for all formulations with steady state being achieved by the second dose. Overall, the clinical use of Formulations A, B, C, or D will eliminate the need for combination oral therapy and reduce time to achieve steady state, with a smaller washout period upon cessation of therapy. Results of this study prove the suitability of using PLGA copolymers of varying composition and molecular weight to develop sustained release formulations that can tailor in vivo behavior and enhance pharmacological effectiveness of the drug.

In vitro-in vivo correlation from lactide-co-glycolide polymeric dosage forms.

The objective of this study was to compare the in vitro behavior of four long-acting subcutaneous risperidone formulations with in vivo performance, with the intent of establishing an IVIVC. Two copolymers of PLGA (50:50 and 75:25) were used to prepare four microsphere formulations of risperidone, an atypical antipsychotic. In vitro behavior was assessed at the physiological temperature (37 °C) using the 'modified dialysis' technique. The in vitro release profile demonstrated rank order behavior with Formulations A and B, prepared using the 50:50 copolymer, exhibiting rapid drug release, while Formulations C and D, prepared using 75:25 PLGA, released drug in a slower manner. In vivo profiles were obtained by two approaches, i.e., deconvolution using the Nelson-Wagner equation (the FDA recommended approach) and using fractional AUC. With both in vivo approaches, the 50:50 PLGA preparations released drug faster than the 75:25 PLGA microspheres, exhibiting the same rank order observed in vitro. Additionally, profiles for the four formulations obtained using the deconvolution approach were nearly superimposable with fractional AUC, implying that the latter procedure could be used as a substitute for the Nelson-Wagner method. A comparison of drug release profiles for the four formulations revealed that in three of the four formulations, in vivo release was slightly faster than that in vitro, but the results were not statistically significant (P > 0.0001). An excellent linear correlation (R2 values between 0.97 and 0.99) was obtained when % in vitro release for each formulation was compared with its corresponding in vivo release profile, obtained by using fraction absorbed (Nelson-Wagner method) or fractional AUC. In summary, using the four formulations that exhibited different release rates, a Level A IVIVC was established using the FDA-recommended deconvolution method and fractional AUC approach. The excellent relationship between in vitro drug release and the amount of drug absorbed in vivo in this study was corroborated by the nearly 1:1 correlation (R2 greater than 0.97) between in vitro release and in vivo performance. Thus, the results of the current study suggest that proper selection of an in vitro method to assess drug release from long-acting injectables will aid in obtaining a Level A IVIVC.

Microsphere delivery of Risperidone as an alternative to combination therapy.

The purpose of this study was to develop a parenteral delivery system of Risperidone that would provide initial and extended drug release and thereby avoid the need for co-administration of oral tablets. Key formulation parameters utilized to achieve desired therapeutic levels in vivo were particle size and drug loading. Three poly (D,L-lactide-co-glycolide) (PLGA) microsphere formulations (Formulations A, B, and C) that encapsulated Risperidone were prepared by varying particle size (19-49 µm) and drug loading parameters (31-37%) but with a uniform bulk density (0.66-0.69)g/cc and internal porosity, utilizing the solvent extraction/evaporation method. The microspheres were characterized for drug content by HPLC, particle size by laser diffractometry, surface morphology by scanning electron microscopy (SEM), and in vivo drug release. In vivo studies were performed in male Sprague-Dawley rats, and levels of the active moiety (Risperidone and its metabolite, 9-hydroxyrisperidone) were assessed. In vivo release profiles from the three microsphere formulations were dependent on particle size and drug loading. The smaller sized microspheres (Formulation A) exhibited a large initial burst and a shorter duration of action, while the larger particles exhibited a smaller initial burst (Formulations B and C) but released drug for a much longer period in vivo. Extended duration of drug release was ascribed to higher drug content in the microspheres. A biweekly simulation of multiple dosing revealed that Formulation C, the selected formulation, with a high load and large particle size would provide adequate initial and maintenance levels of the active moiety (Risperidone and its metabolite, 9-hydroxyrisperidone). A comparison of biweekly dosing in vivo of Formulation C with the marketed product showed that at steady state, though average concentrations for both preparations were similar, the time taken to achieve steady state was much faster for Formulation C. The delay in attaining steady state with Risperdal Consta® was attributed to the 3 week latency in drug release from the microspheres and was in accordance with previous studies indicating a good corroboration with clinical findings. Calculated cumulative AUC (area under the curve) levels for Formulation C were similar to the Risperdal Consta®, though there were marked differences in AUC levels at the early time points. Comparison of Risperidal Consta® and Formulation C by multiple dosing in vivo experiments revealed that the marketed preparation demonstrated a substantial delay in providing an initial loading dose, continuous circulating levels, and attainment of steady state; all of which were observed rapidly with Formulation C. Findings from the current study strongly suggest that a microsphere dosage form of Risperidone can be formulated with an optimum particle size and drug loading to provide an initial bolus followed by maintenance levels, thereby eliminating combination therapy and improving patient compliance.

Preparation, Characterization, and In Vivo Evaluation of Olanzapine Poly(D,L-lactide-co-glycolide) Microspheres.

The aim of this study was to prepare injectable depot formulations of Olanzapine using four poly(D,L-lactide-co-glycolide) (PLGA) polymers of varying molecular weight and copolymer composition, and evaluate in vivo performance in rats. In vivo release profiles from the formulations were governed chiefly by polymer molecular weight and to a lesser extent, copolymer composition. Formulations A and B, manufactured using low molecular weight PLGA and administered at 10 mg/kg dose, released drug within 15 days. Formulation C, prepared from intermediate molecular weight PLGA and administered at 20 mg/kg dose, released drug in 30 days, while Formulation D, manufactured using a high molecular weight polymer and administered at 20 mg/kg dose, released drug in 45 days. A simulation of multiple dosing at 7- and 10-day intervals for Formulations A and B revealed that steady state was achieved within 7-21 days and 10-30 days, respectively. Similarly, simulations at 15-day intervals for Formulations C and D indicated that steady state levels were reached during days 15-45. Overall, steady state levels for 7-, 10-, or 15-day dosing ranged between 45 and 65 ng/mL for all the formulations, implying that Olanzapine PLGA microspheres can be tailored to treat patients with varying clinical needs.

Stability of antimicrobial decapeptide (KSL) and its analogues for delivery in the oral cavity.

PURPOSE: To investigate the stability of KSL, an antimicrobial decapeptide, and its analogues, in human saliva and simulated gastric fluid for delivery in the oral cavity. MATERIALS AND METHODS: The degradation products of KSL in human saliva and simulated gastric fluid were separated by reversed-phase HPLC and their structures were identified by electrospray ionization-mass spectrometry. Analogues of KSL were synthesized by solid-phase synthesis procedure. Their enzymatic stabilities and antimicrobial activities were studied. RESULTS: KSL was degraded by the peptide bond cleavages at Lys(6)-Val(7) in the human saliva and Phe(5)-Lys(6) in simulated gastric fluids. Three analogues of KSL were synthesized; the Lys(6) residue was either methylated (KSL-M), or replaced with Trp (KSL-W), or the d-form of Lys (KSL-D). The KSL analogues were much more stable than the native KSL, with the rank order of stability being KSL-D > KSL-W > KSL-M > KSL in human saliva. However, in simulated gastric fluid, while KSL-D was still stable, KSL-W was significantly degraded. In addition, KSL-D significantly lost the antimicrobial activity, whereas KSL-W completely preserved the activity against several oral bacteria. In a chewing gum formulation, KSL-W showed a more sustained release profile as compared with the native KSL. CONCLUSION: This study suggests that KSL-W could be used as an antiplaque agent in a chewing gum formulation.