Modifying the Fc Asn297 Glycan of Human IgG2 Subclass for Improved Antibody Therapeutics and Design of Site-Specific Antibody Drug Conjugates

Immunoglobulin G (IgG) is a complex glycoprotein that is largely being used in the development of antibody-based therapeutics to treat a variety of diseases such as cancers, autoimmune diseases, and infectious diseases. A major challenge in developing such therapeutics originates from the heterogeneity of the Asn297 glycan of IgG. This sugar part (Asn297 glycan) of IgG has been shown to play an important role in the antibody properties such as antibody stability, effector functions, solubility, pharmacokinetics, and immunogenicity. To study the effect of Asn297 glycan composition on the properties of the human IgG2 (a preferred IgG subclass for developing antibody-based therapeutics when the antibody effector functions are not required), we developed an approach to produces homogeneous IgG2 Fc glycoforms. Studying these homogeneous IgG2 Fc glycoforms suggested that enriching the core fucose within IgG2 Asn297 glycan could add more advantages to developing IgG2-based therapeutics by providing more reduction in any possible undesirable effector functions of these therapeutics. The in vitro enzymatic synthesis developed here also enabled studies of different glycosylation sites accessibility to glycosyltransferase and expanded our understanding of the glycosylation heterogeneity. The Asn297 glycan also offers a great opportunity in developing antibody drug conjugates (ADCs), a therapeutic modality where a toxic payload is conjugated to IgG. The early ADC development relied on conjugating the payload non-specifically through either the lysine or cysteine residues of IgG using conventional chemistries. Producing ADCs in this way, results in heterogeneous products. ADC heterogeneity usually translates into poor pharmacokinetics, stability, and efficacy. In this work, we developed a chemoenzymatic synthesis platform to functionalize the Asn297 glycan in a novel way and use it as a specific site for conjugation. The work involved using and optimizing click chemistry reactions for conjugating different linkers on IgG2 Fc. This approach enabled us to produce conjugates with excellent homogeneity. Subsequent studies of these conjugates showed the utility of the approach followed here in making of site-specific conjugates with good properties and the possibility of following this approach in designing and testing site-specific ADCs during the process of developing next generation ADCs

Preparation of Idebenone as a Thermosetting Nasal Gel for Better Bioavailability and Histopathological Effect

Background Idebenone is an extensively metabolized drug with poor water solubility that is used to treat Leiber’s hereditary optical neuropathy. Objective This study aims to prepare idebenone nanoemulsion as a poloxamer-based nasal gel to overcome the extensive rate of hepatic metabolism for better bioavailability and lower histopathological effect on the nasal mucosa. Methods The formulation strategy was based on eliciting mutual concentration reduction between the nanoemulsion and the carrier gel by setting their gelation temperature between 30-32°C to overcome the mucociliary dose washout. The o/w nanoemulsions rely on cremophor EL and transcutol as an emulsifying system to stabilize idebenone-loaded lemongrass oil. The spontaneous emulsification method was used to prepare nanoemulsions that were characterized by zeta sizer while the thermosensitive hydrogels were prepared using the cold method. In-vitro dissolution test and ex-vivo permeation study through excised sheep nasal mucosa were performed to evaluate the enhanced permeation ratio, rate of permeation, and permeation coefficient. The histopathological effect of direct application on sheep nasal mucosa was studied using optical microscopy to evaluate cellular toxicity. Results The formula prepared from NE1 with poloxamer 407: poloxamer188 in concentrations 10:3% w/w respectively showed almost complete drug release in 120 minutes due to complete polymers blend erosion. Furthermore, thermosensitive nano-emulgel at a temperature of gelation 31.8°C was obtained at much lower concentrations of poloxamer 407 (10%) compared to previous studies. Nanoemulsions retained their globular size below 100nm due to further gel entrapment stabilization. Conclusions Drug permeation through excised sheep nasal mucosa elicited an increase in enhanced permeation ratio to 20.3 times and other flux kinetics parameters compared to those of IDB oil dispersion. Direct cellular toxicity showed a minor inflammatory response characterized by serous infiltration of inflammatory cells and edema. In contrast, most of the epithelial cells retained their histological characteristics compared to control slides.   Received: Mar. 2023 Accepted: Aug 2023 Published: Oct.202

Topical Propranolol Hydrochloride Nanoemulsion: A Promising Approach Drug Delivery for Infantile Hemangiomas

Infantile hemangioma (IH) is a prevalent vascular tumor that affects up to 10% of infants worldwide. If left untreated, IH can lead to a permanent notable physical disfigurement due to skin lesions' ulceration and scarring. Oral propranolol hydrochloride (PHCl) is the only (FDA) approved, first-line treatment for infantile hemangioma (IH), but systemic exposure to propranolol HCl can cause serious adverse reactions. To reduce the risk of adverse effects, topical delivery is preferred due to its ability to provide high local drug concentration and fewer systematic side effects. However, PHCl is (BCS class I) is highly hydrophilic (LogP 1.2), which makes it challenging to be dermally delivered and penetrate the stratum corneum (SC) barrier. Oil in water o/w nanoemulsion (NE) was the chosen pharmaceutical technique to be utilized to solve this obstacle and counteract the difficulty of topical application. Propranolol was formulated as a NE employing clove as an oil phase, Tween 20 as surfactant and )PEG400 or PG( as a co-surfactants. Ten formulas were prepared, and different tests were accomplished to ensure the stability of the NEs, such as electrical conductivity, globule size, polydispersity index, percent of light transmittance (T%), dilution test, pH, zeta potential, drug content, viscosity and in-vitro drug release. Results of characterization revealed that PHCl nanoemulsion (F5) with (clove oil 10%) (Smix 60% of Tween20:PEG400 in a ratio of 3:1) and (DI DW 30%) is the optimum formula, since it has an electrical conductivity of (79 µS/cm), globule size of (14.87 nm), low PDI (0.282), light transmittance T% of (98.09), superior dilution without turbidity, pH value of (6.63), zeta potential (-24.91 mV), percent of drug content (99.79 %) with high viscosity (314.3 mpa.s at 6 rpm), and complete slow-release of the drug after (2hr). The preferred formula (F5) was subjected to further analysis by FTIR spectrophotometer. The findings indicated that the distinguishing peaks for PHCl remained unaffected, and we can conclude that there was no interaction between the drug and other NE constituents. In conclusion, the NE can be considered as an innovative approach to enhance the transport of drug molecules efficiently by overcoming the barrier properties of the skin (IH), despite the high hydrophilic properties of such drugs. الورم الوعائي الطفلي هو ورم وعائي واسع الانتشار يصيب ما يصل إلى 10٪ من الأطفال حول العالم. إذا تركت دون علاج، يمكن أن يؤدي إلى تشوه دائم ملحوظ بسبب تقرح وتندب الجلد. بروبرانولول هايدروكلورايد الفموي هو العلاج الوحيد المعتمد من إدارة الغذاء والدواء (FDA) للورم الوعائي الطفلي، ولكن التعرض الجهازي يمكن أن يسبب ردود فعل سلبية خطيرة. لتقليل مخاطر الآثار الضارة، يُفضل التوصيل الموضعي نظرًا لقدرته على توفير تركيز موضعي عالٍ وآثار جانبية جهازية أقل. ومع ذلك، فإن العقار (فئة BCS I) شديد المحبة للماء (LogP 1.2)، مما يجعل من الصعب توصيله عن طريق الجلد واختراق حاجز الطبقة المتقرنة (SC). المستحلب النانوي زيت في ماء، كان الأسلوب الصيدلاني المختار لاستخدامه في حل هذه العقبة ومواجهة صعوبة الاستعمال الموضعي. لقد تم تصييغ بروبرانولول على أنه مستحلب نانوي باستخدام زيت القرنفل كوسط زيتي، توين 20 كخافض للتوتر السطحي وPEG 400 اوPG كمواد مضادة للشد السطحي. تم تحضير عشرة تراكيب سائلة، وتم إجراء اختبارات مختلفة لها لضمان ثبات المستحلب النانوي مثل، التوصيل الكهربائي، حجم القطيرات، مؤشر التشتت المتعدد، ونسبة نفاذية الضوء (T٪)، اختبار التخفيف، درجة الحموضة، جهد زيتا، تقدير محتوى العقار، واللزوجة ونسبة تحرر العقار في المختبر. أظهرت نتائج التوصيف أن التركيبة (5) لمستحلب بروبرانولول التي تحتوي على (زيت القرنفل 10٪) (Smix 60٪ من توين 20: PEG 400 بنسبة 3:1) و (DI DW 30٪) هي التركيبة الافضل، لأنها تمتاز بموصلية كهربائية (79 µS / سم)، حجم قطيرات (14.87 نانومتر)، مؤشر تشتت منخفض (0.282)، نفاذية الضوء عالية T٪ (98.09)، تخفيف فائق بدون تعكر (تخفيفه بـ 20 جزءًا مائيا)، درجة الحموضة (6.63)، جهد زيتا (-24.91 مللي فولت)، محتوى العقار (99.79٪) مع لزوجة عالية (314.3 ميجا باسكال عند 6 دورة في الدقيقة)، وتحرر بطيء كامل للدواء بعد (ساعاتين). خضعت التركيبة المفضلة (5) لمزيد من التحليل مثل دراسة التوافق مع المكونات الاخرى بواسطة الفحص بالأشعة تحت الحمراء. أشارت النتائج إلى أن القمم المميزة للعقار ظلت غير متأثرة ويمكن أن نستنتج أنه لم يكن هناك تفاعل بين الدواء ومكونات المستحلب النانوي وكانت متسقة. في الختام، وجد أن المستحلب النانوي هو، وسيلة مبتكرة لتعزيز نقل جزيئات الدواء من خلال التغلب على خصائص حاجز الجلد (الورم الوعائي)، على الرغم من خصائص العقار العالية المحبة للماء.

Formulation and in vitro evaluation of meloxicam as a self-microemulsifying drug delivery system [version 2; peer review: 2 approved]

Background: The nonsteroidal anti-inflammatory medication meloxicam (MLX) belongs to the oxicam family and is used to reduce inflammation and pain. The aim of this study was to improve MLX's dispersibility and stability by producing it as a liquid self-microemulsifying drug delivery system since it is practically insoluble in water. Methods: Five different formulations were made by adjusting the amounts of propylene glycol, Transcutol P, Tween 80, and oleic acid oil and establishing a pseudo-ternary diagram in ratios of 1:1, 1:2, 1:3, 1:4, and 3:4, respectively. All of the prepared formulations were tested for a variety of properties, including thermodynamic stability, polydispersity index, particle size distributions, dilution resistance, drug contents, dispersibility, in vitro solubility of the drug, and emulsification time. Results: F5 was chosen as the optimal MLX liquid self-microemulsion due to its higher drug content (99.8%), greater in vitro release (100% at 40 min), smaller droplet size (63 nm), lower polydispersity index (PDI) value (0.3), and higher stability (a zeta potential of -81 mV). Conclusions: According to the data provided here, the self-microemulsifying drug delivery system is the most practical method for improving the dispersibility and stability of MLX

INVESTIGATING THE EFFECT OF DIFFERENT GRADES AND CONCENTRATIONS OF PH‑SENSITIVE POLYMER ON PREPARATION AND CHARACTERIZATION OF LIDOCAINE HYDROCHLORIDE AS IN SITU GEL BUCCAL SPRAY

Objective: The present study was aimed to develop a pH-triggered in situ gel for local release of lidocaine hydrochloride (lidocaine HCL) in the buccal cavity to improve the anesthetic effect of this amino amide drug which has very high water solubility. The formulations were introduced to the oral cavity as a spray to improve compliance and for easier administration.Methods: In this work, two grades of carbopol (934 and 940)-based in situ gel spray were designed. The formulations containing lidocaine HCl 5% were prepared by mixing different concentrations of carbopol with xanthan gum. Eight formulations were investigated and evaluated for gelation capacity, spray angle, volume of solution delivered per each actuation, rheological properties, and release kinetic model. Similarity factor (f2) was used for the comparison of dissolution profiles.Results: The prepared formulations undergo gelation after it had been actuated to the buccal cavity as a spray solution. The results showed that, as the concentration of polymer was increased, the release of drug decreased and the viscosity increased for both grades. The spray angle and volume of solution delivered per each actuation varied according to the composition of each formulation. The in situ gel containing 0.3% carbopol 934 and 0.2% xanthan gum regarded as a better candidate which had a good gelation and release property compared to other formulations. Drug release from optimized in situ gel spray followed Korsmeyer–Peppas model and was mediated by Fickian diffusion mechanism.Conclusion: Lidocaine HCl-loaded pH-sensitive in situ gel was successfully developed using carbopol 934 as polymer to be applied to the buccal cavity as spray solution for more effective anesthetic effect and painless treatment