Next-generation Drugs Research Articles

Surfactant-Free Synthesis of PNIPAM-Based Smart Microgels for Drug Delivery Using a High-Gravity Rotating Packed Bed

Poly(N-isopropylacrylamide) (PNIPAM) microgels have a number of potential biomedical applications, especially being considered as next-generation drug delivery systems. So far, nm-sized PNIPAM microgels have been commonly prepared in lab-scale experiments. In this work, nm-size controllable and monodisperse PNIPAM-based microgels were synthesized for the first time by surfactant-free precipitation polymerization using a high-gravity rotating packed bed reactor (RPB). The microgel size could be synergistically controlled by adjusting the amount of cross-linker and initiator in an RPB. Compared with the conventional stirred tank reactor, the RPB synthesis procedure could obtain a higher yield within 2 h of the reaction time. The higee level exhibited significant influence on the microgel size. Depending on the variation of the higee level, the particle size was tailored from 129 to 325 nm, and the hydrodynamic diameter was tailored from 217.7 to 805.4 nm without the usage of surfactants. In addition, different comonomers were introduced to regulate the lower critical solution temperature (LCST), achieve multiresponsiveness, and control microgel size. The in vitro DOX-loaded release demonstrated that PNIPAM-based microgels dramatically contributed to the sustained release of drugs.

Development of peptide affinity ligands for the purification of polyclonal and monoclonal Fabs from recombinant fluids

Engineered multi-specific monoclonal antibodies (msAbs) and antibody fragments offer valuable therapeutic options against metabolic disorders, aggressive cancers, and viral infections. The advancement in molecular design and recombinant expression of these next-generation drugs, however, is not equaled by the progress in downstream bioprocess technology. The purification of msAbs and fragments requires affinity adsorbents with orthogonal biorecognition of different portions of the antibody structure, namely its Fc (fragment crystallizable) and Fab (fragment antigen-binding) regions or the CH1-3 and CL chains. Current adsorbents rely on protein ligands that, while featuring high binding capacity and selectivity, need harsh elution conditions and suffer from high cost, limited biochemical stability, and potential release of immunogenic fragments. Responding to these challenges, we undertook the de novo discovery of peptide ligands that target different regions of human Fab and enable product release under mild conditions. The ligands were discovered by screening a focused library of 12-mer peptides against a feedstock comprising human Fab and Chinese hamster ovary host cell proteins (CHO HCPs). The identified ligands were evaluated via binding studies as well as molecular docking simulations, returning excellent values of binding capacity (Qmax ∼ 20 mg of Fab per mL of resin) and dissociation constant (KD = 2.16·10−6 M). Selected ligand FRWNFHRNTFFP and commercial Protein L ligands were further characterized by measuring the dynamic binding capacity (DBC10%) at different residence times (RT) and performing the purification of polyclonal and monoclonal Fabs from CHO-K1 cell culture fluids. The peptide ligand featured DBC10% ∼ 6-16 mg/mL (RT of 2 min) and afforded values of yield (93–96%) and purity (89–96%) comparable to those provided by Protein L resins.

User-designed device with programmable release profile for localized treatment.

Three-dimensional printing enables precise and on-demand manufacture of customizable drug delivery systems to advance healthcare toward the goal of personalized medicine. However, major challenges remain in realizing personalized drug delivery that fits a patient-specific drug dosing schedule using local drug delivery systems. In this study, a user-designed device is developed as implantable therapeutics that can realize personalized drug release kinetics by programming the inner structural design on the microscale. The drug release kinetics required for various treatments, including dose-dense therapy and combination therapy, can be implemented by controlling the dosage and combination of drugs along with the rate, duration, initiation time, and time interval of drug release according to the device layer design. After implantation of the capsular device in mice, the in vitro-in vivo and pharmacokinetic evaluation of the device is performed, and the therapeutic effect of the developed device is achieved through the local release of doxorubicin. The developed user-designed device provides a novel platform for developing next-generation drug delivery systems for personalized and localized therapy.

Emerging pharmaceutical therapeutics and delivery technologies for osteoarthritis therapy.

Osteoarthritis (OA) is one of the most common joint degenerative diseases in the world. At present, the management of OA depends on the lifestyle modification and joint replacement surgery, with the lifespan of prosthesis quite limited yet. Effective drug treatment of OA is essential. However, the current drugs, such as the non-steroidal anti-inflammatory drugs and acetaminophen, as well as glucosamine, chondroitin sulfate, hyaluronic acid, are accompanied by obvious side effects, with the therapeutic efficacy to be enhanced. Recently, novel reagents such as IL-1 antagonists and nerve growth factor inhibitors have entered clinical trials. Moreover, increasing evidence demonstrated that active ingredients of natural plants have great potential for treating OA. Meanwhile, the use of novel drug delivery strategies may overcome the shortcomings of conventional preparations and enhance the bioavailability of drugs, as well as decrease the side effects significantly. This review therefore summarizes the pathological mechanisms, management strategies, and research progress in the drug molecules including the newly identified active ingredient derived from medicinal plants for OA therapy, with the drug delivery technologies also summarized, with the expectation to provide the summary and outlook for developing the next generation of drugs and preparations for OA therapy.

From Promise to Reality: Bioengineering Strategies to Enhance the Therapeutic Potential of Extracellular Vesicles.

Extracellular vesicles (EVs) have been the focus of great attention over the last decade, considering their promising application as next-generation therapeutics. EVs have emerged as relevant mediators of intercellular communication, being associated with multiple physiological processes, but also in the pathogenesis of several diseases. Given their natural ability to shuttle messages between cells, EVs have been explored both as inherent therapeutics in regenerative medicine and as drug delivery vehicles targeting multiple diseases. However, bioengineering strategies are required to harness the full potential of EVs for therapeutic use. For that purpose, a good understanding of EV biology, from their biogenesis to the way they are able to shuttle messages and establish interactions with recipient cells, is needed. Here, we review the current state-of-the-art on EV biology, complemented by representative examples of EVs roles in several pathophysiological processes, as well as the intrinsic therapeutic properties of EVs and paradigmatic strategies to produce and develop engineered EVs as next-generation drug delivery systems.

27P Outcomes analysis of the effect of an educational activity on the knowledge and confidence of oncologists regarding emerging antibody drug conjugates for the treatment of breast cancer

The objective of this study was to assess the effect of an educational activity on the knowledge of oncologists regarding the rationale for using next-generation antibody drug conjugates (ADCs) for the treatment of breast cancer and the latest data from clinical trials.

Peptide-Drug Conjugates: A New Hope for Cancer Management.

Cancer remains the leading cause of death worldwide despite advances in treatment options for patients. As such, safe and effective therapeutics are required. Short peptides provide advantages to be used in cancer management due to their unique properties, amazing versatility, and progress in biotechnology to overcome peptide limitations. Several appealing peptide-based therapeutic strategies have been developed. Here, we provide an overview of peptide conjugates, the better equivalents of antibody-drug conjugates, as the next generation of drugs for required precise targeting, enhanced cellular permeability, improved drug selectivity, and reduced toxicity for the efficient treatment of cancers. We discuss the basic components of drug conjugates and their release action, including the release of cytotoxins from the linker. We also present peptide-drug conjugates under different stages of clinical development as well as regulatory and other challenges.

CD44 and CD221 directed magnetic cubosomes for the targeted delivery of helenalin to rhabdomyosarcoma cells

Confining chemotherapy to tumour sites by means of active targeting nanoparticles (NPs) may increase the treatment effectuality while reducing potential side effects. Cubosomes are one of the next-generation drug delivery nanocarriers by virtue of their biocompatibility and bioadhesion, sizeable payload encapsulation and high thermostability. Herein, an active tumour targeting system towards rhabdomyosarcoma (RMS) cells was evaluated. Cubosomes were loaded with helenalin (a secondary metabolite from Arnica plants), which we have previously shown to induce apoptosis in RMS cells. The functionalization of the cubosomes was accomplished to enable binding to membrane receptors and translocation under a magnetic field. RMS cells overexpress CD44 and CD221 on their membrane surface and, therefore, hyaluronic acid (HA, a ligand for CD44) and antibodies (Abs) against CD221 were coupled to cubosomes via electrostatic attraction and the thiol-Michael reaction, respectively. Magnetization of the cubic phase NPs was achieved by embedding superparamagnetic iron oxide NPs (SPIONPs) into the cubic matrix. Single-function and multi-function cubosomes had Im3m cubic phase structures with well-organized lattice patterns. Conjugation with 2% HA or anti-CD221 half Abs and/or 1% SPIONPs showed significantly higher uptake into RMS cells compared to unfunctionalized cubosomes. CD44 and CD221 directed magnetic (triple-function) cubosomes were capable of internalizing into RMS cells in an energy-independent mechanism. Helenalin-laden triple functionalized cubosomes showed limited impact on the viability of control fibroblast cells, while they induced a high degree cytotoxicity against RMS cells. Profound tumour cell death was observed in both two-dimensional (2D) culture and three-dimensional (3D) tumour spheroids.

Ultrasmall Folate Receptor Alpha Targeted Enzymatically Cleavable Silica Nanoparticle Drug Conjugates Augment Penetration and Therapeutic Efficacy in Models of Cancer.

To address the key challenges in the development of next-generation drug delivery systems (DDS) with desired physicochemical properties to overcome limitations regarding safety, in vivo efficacy, and solid tumor penetration, an ultrasmall folate receptor alpha (FRα) targeted silica nanoparticle (C'Dot) drug conjugate (CDC; or folic acid CDC) was developed. A broad array of methods was employed to screen a panel of CDCs and identify a lead folic acid CDC for clinical development. These included comparing the performance against antibody-drug conjugates (ADCs) in three-dimensional tumor spheroid penetration ability, assessing in vitro/ex vivo cytotoxic efficacy, as well as in vivo therapeutic outcome in multiple cell-line-derived and patient-derived xenograft models. An ultrasmall folic acid CDC, EC112002, was identified as the lead candidate out of >500 folic acid CDC formulations evaluated. Systematic studies demonstrated that the lead formulation, EC112002, exhibited highly specific FRα targeting, multivalent binding properties that would mediate the ability to outcompete endogenous folate in vivo, enzymatic responsive payload cleavage, stability in human plasma, rapid in vivo clearance, and minimal normal organ retention organ distribution in non-tumor-bearing mice. When compared with an anti-FRα-DM4 ADC, EC112002 demonstrated deeper penetration into 3D cell-line-derived tumor spheroids and superior specific cytotoxicity in a panel of 3D patient-derived tumor spheroids, as well as enhanced efficacy in cell-line-derived and patient-derived in vivo tumor xenograft models expressing a range of low to high levels of FRα. With the growing interest in developing clinically translatable, safe, and efficacious DDSs, EC112002 has the potential to address some of the critical limitations of the current systemic drug delivery for cancer management.

Antiviral activity of soybean GL 2626/96 (Glycine max) ethanolic extract against influenza A virus in vitro and in vivo

Influenza viruses cause respiratory infections in humans with high morbidity and mortality rates. Neuraminidase inhibitors such as oseltamivir and peramivir are the most commonly used drugs for influenza virus infections. However, the emergence of resistant viruses necessitates the urgent need to develop next-generation anti-influenza drugs. Soybean (Glycine max L. Merr.) is widely cultivated and used as food worldwide. In addition, soybean has long been used as a nutritional supplement and herbal medicine. However, the potential anti-influenza properties of the soybean cultivar “GL 2626/96″ (SG2626) are yet to be investigated. Herein, we determined whether the ethanolic extract of SG2626 (SG2626E) has anti-viral activity through performing SG2626E pre-, co-, and post-treatment assays, using the influenza green fluorescent protein (GFP)-tagged influenza A/PR/8/34 (A/PR/8/34-GFP) virus. SG2626E showed anti-influenza virus activity in pre- and co-treated cells in a dose-dependent manner, but not in post-treated cells. SG2626E imparted a considerable inhibitory effect on influenza A virus (IAV) infection through blocking viral attachment. SG2626E inhibited the activity of viral hemagglutinin, but not viral neuraminidase of the IAV. SG2626E inhibited IAV infection by reducing intracellular calcium levels in infected human lung epithelial A549 cells. Additionally, SG2626E reduced body weight loss, decreased mortality, and increased the survival rate through reducing viral replication in the lungs of IAV-infected mice. Overall, these results suggest that SG2626E inhibits IAV infection and is a potential novel anti-influenza agent.

Noncoding RNAs Emerging as Drugs or Drug Targets: Their Chemical Modification, Bio-Conjugation and Intracellular Regulation.

With the increasing understanding of various disease-related noncoding RNAs, ncRNAs are emerging as novel drugs and drug targets. Nucleic acid drugs based on different types of noncoding RNAs have been designed and tested. Chemical modification has been applied to noncoding RNAs such as siRNA or miRNA to increase the resistance to degradation with minimum influence on their biological function. Chemical biological methods have also been developed to regulate relevant noncoding RNAs in the occurrence of various diseases. New strategies such as designing ribonuclease targeting chimeras to degrade endogenous noncoding RNAs are emerging as promising approaches to regulate gene expressions, serving as next-generation drugs. This review summarized the current state of noncoding RNA-based theranostics, major chemical modifications of noncoding RNAs to develop nucleic acid drugs, conjugation of RNA with different functional biomolecules as well as design and screening of potential molecules to regulate the expression or activity of endogenous noncoding RNAs for drug development. Finally, strategies of improving the delivery of noncoding RNAs are discussed.

Fundamental Studies on Development of Next-generation Medium Sized Peptide Drugs

Medium-sized peptides are expected as a next-generation drug discovery modality because they combine the properties of conventional small-molecule drugs and biopharmaceuticals. Nonetheless, peptides are easily degraded by digestive enzymes such as protease in the body, which could be problematic for the development of peptide-based drugs. To overcome such a problem, peptide-based foldamers containing non-proteinogenic amino acids or cyclized peptides have been reported. In addition, peptides must form stable secondary structures and their side chains should be correctly positioned to exert their bioactivity. In our lab, bioactive peptides have been developed based on regulation of secondary structures by introducing non-proteinogenic amino acids such as acyclic α,α-disubstituted amino acids (dAAs), cyclic dAAs, cyclic β-amino acids, and side-chain stapling. Based on these knowledges, I have been performing research on the development of bioactive peptides based on the secondary structural control of peptides as categorized in the following manner: (1) rational design of antimicrobial foldamers; (2) post-functionalization of helical peptides; (3) development of carrier peptides for intracellular delivery of siRNA utilizing the helical template peptides.

Innovative Translation Strategies in the Emergence of Next-Generation Drugs in Precision Cancer Medicine

There has been a conscientious momentum to evolve more efficacious and potent chemotherapeutics that could cure advanced cancers. Although, the conventional cancer therapeutics can cause dramatic tumor regressions in some patients, many patients inexplicably reveal no benefit. This is a unique characteristic of tumor recurrence in human cancers, linked with several resistance mechanisms. Recent past years witnessed enormous ground-breaking advancements in genomic medicine symbolizing a culmination of patients' genomic, proteomic, metabolomic, including cellular profiles which have significantly transformed understanding and therapeutic options for cancer management. Furthermore, the ability of combination chemotherapy to cure advanced cancers, immensely facilitated the prospects of adjuvant chemotherapy. Many global pharmaceuticals have accelerated translational research strategies on proteomics and genomic technologies paving the way to emerge next-generation drugs to strengthen personalized cancer medicine. Numerous forthcoming cancer therapies are at most vital phase in their innovation and development for the unfulfilled clinical need to turn into futuristic cancer treatment strategies. There is a growing optimism about the innovative strategies on emergence of the new generation of immunotherapy, targeted therapies, and Oligonucleotide therapy to improve patient outcomes, and to make real dream of accomplishing personalized cancer therapy.

Venom Variation of Neonate and Adult Chinese Cobras in Captivity Concerning Their Foraging Strategies.

The venom and transcriptome profile of the captive Chinese cobra (Naja atra) is not characterized until now. Here, LC-MS/MS and illumine technology were used to unveil the venom and trascriptome of neonates and adults N. atra specimens. In captive Chinese cobra, 98 co-existing transcripts for venom-related proteins was contained. A total of 127 proteins belong to 21 protein families were found in the profile of venom. The main components of snake venom were three finger toxins (3-FTx), snake venom metalloproteinase (SVMP), cysteine-rich secretory protein (CRISP), cobra venom factor (CVF), and phosphodiesterase (PDE). During the ontogenesis of captive Chinese cobra, the rearrangement of snake venom composition occurred and with obscure gender difference. CVF, 3-FTx, PDE, phospholipase A2 (PLA2) in adults were more abundant than neonates, while SVMP and CRISP in the neonates was richer than the adults. Ontogenetic changes in the proteome of Chinese cobra venom reveals different strategies for handling prey. The levels of different types of toxin families were dramatically altered in the wild and captive specimens. Therefore, we speculate that the captive process could reshape the snake venom composition vigorously. The clear comprehension of the composition of Chinese cobra venom facilitates the understanding of the mechanism of snakebite intoxication and guides the preparation and administration of traditional antivenom and next-generation drugs for snakebite.

Plant hairy roots for the production of extracellular vesicles with antitumor bioactivity

Plant extracellular vesicles (EVs) concentrate and deliver different types of bioactive molecules in human cells and are excellent candidates for a next-generation drug delivery system. However, the lack of standard protocols for plant EV production and the natural variations of their biomolecular cargo pose serious limitation to their use as therapeutics. To overcome these issues, we set up a versatile and standardized procedure to purify plant EVs from hairy root (HR) cultures, a versatile biotechnological system, already successfully employed as source of bioactive molecules with pharmaceutical and nutraceutical relevance. Herewith, we report that HR of Salvia dominica represent an excellent platform for the production of plant EVs. In particular, EVs derived from S. dominica HRs are small round-shaped vesicles carrying typical EV-associated proteins such as cytoskeletal components, chaperon proteins and integral membrane proteins including the tetraspanin TET-7. Interestingly, the HR-derived EVs showed selective and strong pro-apoptotic activity in pancreatic and mammary cancer cells. These results reveal that plant hairy roots may be considered a new promising tool in plant biotechnology for the production of extracellular vesicles for human health.

The road ahead for applications of mechanics in drug delivery

From vaccination to cancer treatment, controlled drug delivery systems have shown great promise across a wide range of therapies. Understanding the mechanics of advanced drug delivery systems is essential to aid in the design of better treatments for patients. Mechanics has particularly emerged as a key area intersecting engineering and drug delivery. Most drug delivery systems work in direct interaction with the body (e. g. vasculature, GI tract). In this perspective, we focus on three areas highlighting the importance of mechanics for next-generation drug delivery applications. We identify challenges and opportunities for innovation. The areas explored include micro/nano mechanics, gastrointestinal mechanics, and vascular mechanics for next generation drug delivery systems and organoids. First, we review applications of micro/nano technology in fabrication of advanced drug delivery systems. We specifically highlight opportunities for using computational mechanics in drug delivery in micro/nano scales. Second, we include a discussion on mechanical considerations for the design of next-generation cancer therapeutics, drug discovery systems and cell therapy platforms. Finally, we discuss the importance of a detailed understanding of gastrointestinal mechanics in designing orally administered drug delivery systems. Taken together, we highlight areas of mechanics that could be important frontiers for innovation across length scales, disease models and organ systems for advanced therapeutics.

Incorporating HPLC as a Multi-Attribute Characterization Technology for Next-Generation Drug Development

The complexity of next-generation drug modalities and delivery systems requires new and integrated analytical approaches. A progressive design of chromatographic instrumentation, stationary phases, flexible data systems, and integrated control are needed for chromatography to remain a leading relevant characterization tool. This paper provides an overview of some current and emerging physicochemical analytical challenges associated with these sophisticated drug systems and some of the technical advances needed in chromatographic systems to enable their design, development, and manufacture.

Biocompatible Supramolecular Mesoporous Silica Nanoparticles as the Next-Generation Drug Delivery System.

Supramolecular mesoporous silica nanoparticles (MSNs) offer distinct properties as opposed to micron-sized silica particles in terms of their crystal structure, morphology–porosity, toxicity, biological effects, and others. MSN biocompatibility has touched the pharmaceutical realm to exploit its robust synthesis pathway for delivery of various therapeutic molecules including macromolecules and small-molecule drugs. This article provides a brief review of MSN history followed by special emphasis on the influencing factors affecting morphology–porosity characteristics. Its applications as the next-generation drug delivery system (NGDDS) particularly in a controlled release dosage form via an oral drug delivery system are also presented and shall be highlighted as oral delivery is the most convenient route of drug administration with the economical cost of development through to scale-up for clinical trials and market launch.

Abstract 2338: Bioluminescent assays for measuringthemetabolic state of cancer and immune cells

Abstract In the tumor microenvironment, cancer and immune cells compete for limited nutrients and both must undergo complex metabolic changes to support their function. To better understand the underlying mechanisms of tumor growth, metastasis and adaptation, a comprehensive analysis of metabolic pathways is needed. Success of such studies requires rapid and reliable approaches for monitoring key metabolic pathways, including glycolysis, the pentose phosphate pathway, fatty acid metabolism, amino acid metabolism and TCA cycle. Due to its inherent robustness, sensitivity and broad dynamic range, bioluminescence detection provides an attractive opportunity for developing such assays. Here, we implement a panel of bioluminescent metabolite assays (glucose, lactate, glutamine/glutamate, triglyceride/glycerol, cholesterol esters/cholesterol) to investigate the metabolic state of immune and cancer cells. We demonstrate flexibility of the assays in various volumes (96-, 384- well plates), sample types (2D, 3D cultures), automation platforms, and workflows, including detecting multiple metabolites in the same sample and monitoring metabolite changes over time by assaying multiple samples. We validate the performance of the assays by using metabolic pathway-specific inhibitors and demonstrating the utility of the assays as early indicators of cytotoxic T-cell activation. Finally, since metabolism is a dynamic process guided by fuel availability, we show the importance of medium composition when designing studies and suggest guidance for future work. In conclusion, understanding the crosstalk between tumor cells and their microenvironment is required for next-generation drug development. Bioluminescent metabolite assays provide the sensitivity, throughput and robustness required for rapid evaluation of changes in major metabolic pathways and are well suited for novel inhibitor screening and drug development. Citation Format: Maggie Bach, Donna Leippe, Natasha Karassina, Michael Valley, James Cali, Jolanta Vidugiriene. Bioluminescent assays for measuringthemetabolic state of cancer and immune cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2338.

The orexin story, sleep and sleep disturbances.

The orexins, also known as hypocretins, are two neuropeptides (orexin A and B or hypocretin 1 and 2) produced by a few thousand neurons located in the lateral hypothalamus that were independently discovered by two research groups in 1998. Those two peptides bind two receptors (orexin/hypocretin receptor 1 and receptor 2) that are widely distributed in the brain and involved in the central physiological regulation of sleep and wakefulness, orexin receptor 2 having the major role in the maintenance of arousal. They are also implicated in a multiplicity of other functions, such as reward seeking, energy balance, autonomic regulation and emotional behaviours. The destruction of orexin neurons is responsible for the sleep disorder narcolepsy with cataplexy (type 1) in humans, and a defect of orexin signalling also causes a narcoleptic phenotype in several animal species. Orexin discovery is unprecedented in the history of sleep research, and pharmacological manipulations of orexin may have multiple therapeutic applications. Several orexin receptor antagonists were recently developed as new drugs for insomnia, and orexin agonists may be the next-generation drugs for narcolepsy. Given the broad range of functions of the orexin system, these drugs might also be beneficial for treating various conditions other than sleep disorders in the near future.