Single-port versus multiport robotic total mesorectal excision for rectal cancer: initial experiences by case-matched analysis of short-term outcomes.

Purpose: The da Vinci single-port (SP) system has been used in various surgical fields, including colorectal surgery. However, limited experience has been reported on its safety and feasibility. This study aims to evaluate the short-term outcomes of SP robotic surgery for the treatment of rectal cancer compared with multiport (MP) robotic surgery. Methods: Rectal cancer patients who underwent curative resection in 2020 were reviewed. A total of 43 patients underwent robotic total mesorectal excision (TME), of which 26 (13 in each group, SPTME vs. MPTME) were included in the case-matched cohort for analysis. Intraoperative and postoperative outcomes and pathological results were compared between the 2 groups. Results: Median tumor height was similar between the 2 groups (SPTME vs. MPTME: 5.9 cm [range, 2.2-9.6 cm] vs. 6.7 cm [range, 3.4-10.0 cm], P = 0.578). Preoperative chemoradiotherapy was equally performed (38.5%). The median estimated blood loss was less (20.0 mL [range, 5.0-20.0 mL] vs. 30.0 mL [range, 20.0-30.0 mL], P = 0.020) and the median hospital stay was shorter (7 days [range, 6-8 days] vs. 8 days [range, 7-9 days], P = 0.055) in the SPTME group. Postoperative complications did not differ (SPTME vs. MPTME: 7.7% vs. 23.1%, P = 0.587). One patient in the SPTME group and 3 in the MPTME group experienced anastomotic leakage. Conclusion: SP robotic TME showed perioperative outcomes similar to MP robotic TME. The SP robotic system can be considered a surgical option for the treatment of rectal cancer. Further prospective randomized trials with larger cohorts are required.

Single-Port Robotic Intersphincteric Resection for the Treatment of Rectal Cancer.

BACKGROUND: The da Vinci Single-port (SP) system is designed to facilitate single-incision robotic surgery in a narrow space. We developed a new procedure of intersphincteric resection (ISR) using the SP platform and evaluated the technical safety and feasibility of this procedure for the treatment of very low rectal cancer. MATERIALS AND METHODS: Eleven rectal cancer patients who underwent SP robotic ISR between August 2020 and July 2021 were included. Patients' clinical characteristics, operative and pathologic findings of the patients were retrospectively analyzed. RESULTS: The median tumor height was 3 cm (range, 2-4 cm). A single docking was performed, and the median docking time was 3 min 10 sec (range, 2 min 50 sec-3 min 30 sec). The median total operation time was 210 min (range, 150-280 min), and the median time of pelvic dissection was 57 min (range, 45-68 min). All patients presented with negative distal resection margins [median 1 cm (range, 0.5-2.0 cm)], and only one patient had less than 1mm of circumferential resection margin (0.9 mm). CONCLUSIONS: Our initial experience suggests that SP robotic ISR is safe and feasible.

Efficacy of air leak test in detection of anastomotic leaks after rectal excision: a retrospective case-controlled study.

Purpose: Although its efficacy is uncertain, an intraoperative air leak test (ALT) is commonly used to detect mechanical defects following bowel anastomosis. This study aimed to evaluate the efficacy of ALT to detect anastomotic leakage (AL) following rectal excision. Methods: We reviewed our database for patients with rectal cancers who had undergone curative surgery between January 2012 and January 2018. Patients were grouped according to whether or not an ALT was performed. Propensity score analyses were performed to compare outcomes for groups in a 1:1 case-matched cohort. Results: In total, 1,191 patients underwent rectal excision; 438 (219 in each group) formed the case-matched cohort for analysis. The protective stoma rate was 16.0% and 14.6% in the ALT and the no-ALT groups, respectively (P = 0.791). In the ALT group, 2 patients (0.9%) showed a positive result and were treated with rectal tube drainage, resulting in no leakage. There was no significant difference in postoperative AL rate between the groups (ALT group: 4.6% vs. no-ALT group: 4.1%, P > 0.999). Conclusion: ALT played a minimal role in detecting AL following rectal excision. Further studies are warranted to validate our results and clarify whether AL can be prevented with ALT or alternative methods.

Gastrointestinal Permeation Enhancers for the Development of Oral Peptide Pharmaceuticals.

Recently, two oral-administered peptide pharmaceuticals, semaglutide and octreotide, have been developed and are considered as a breakthrough in peptide and protein drug delivery system development. In 2019, the Food and Drug Administration (FDA) approved an oral dosage form of semaglutide developed by Novo Nordisk (Rybelsus®) for the treatment of type 2 diabetes. Subsequently, the octreotide capsule (Mycapssa®), developed through Chiasma's Transient Permeation Enhancer (TPE) technology, also received FDA approval in 2020 for the treatment of acromegaly. These two oral peptide products have been a significant success; however, a major obstacle to their oral delivery remains the poor permeability of peptides through the intestinal epithelium. Therefore, gastrointestinal permeation enhancers are of great relevance for the development of subsequent oral peptide products. Sodium salcaprozate (SNAC) and sodium caprylate (C8) have been used as gastrointestinal permeation enhancers for semaglutide and octreotide, respectively. Herein, we briefly review two approved products, Rybelsus® and Mycapssa®, and discuss the permeation properties of SNAC and medium chain fatty acids, sodium caprate (C10) and C8, focusing on Eligen technology using SNAC, TPE technology using C8, and gastrointestinal permeation enhancement technology (GIPET) using C10.

Cleavage-Responsive Biofactory T Cells Suppress Infectious Diseases-Associated Hypercytokinemia.

Severe infectious diseases, such as coronavirus disease 2019 (COVID-19), can induce hypercytokinemia and multiple organ failure. In spite of the growing demand for peptide therapeutics against infectious diseases, current small molecule-based strategies still require frequent administration due to limited half-life and enzymatic digestion in blood. To overcome this challenge, a strategy to continuously express multi-level therapeutic peptide drugs on the surface of immune cells, is established. Here, chimeric T cells stably expressing therapeutic peptides are presented for treatment of severe infectious diseases. Using lentiviral system, T cells are engineered to express multi-level therapeutic peptides with matrix metallopeptidases- (MMP-) and tumor necrosis factor alpha converting enzyme- (TACE-) responsive cleavage sites on the surface. The enzymatic cleavage releases γ-carboxyglutamic acid of protein C (PC-Gla) domain and thrombin receptor agonist peptide (TRAP), which activate endothelial protein C receptor (EPCR) and protease-activated receptor-1 (PAR-1), respectively. These chimeric T cells prevent vascular damage in tissue-engineered blood vessel and suppress hypercytokinemia and lung tissue damages in vivo, demonstrating promise for use of engineered T cells against sepsis and other infectious-related diseases.

Recent Progress in Drug Release Testing Methods of Biopolymeric Particulate System.

Biopolymeric microparticles have been widely used for long-term release formulations of short half-life chemicals or synthetic peptides. Characterization of the drug release from microparticles is important to ensure product quality and desired pharmacological effect. However, there is no official method for long-term release parenteral dosage forms. Much work has been done to develop methods for in vitro drug release testing, generally grouped into three major categories: sample and separate, dialysis membrane, and continuous flow (flow-through cell) methods. In vitro drug release testing also plays an important role in providing insight into the in vivo performance of a product. In vitro release test with in vivo relevance can reduce the cost of conducting in vivo studies and accelerate drug product development. Therefore, investigation of the in vitro-in vivo correlation (IVIVC) is increasingly becoming an essential part of particulate formulation development. This review summarizes the principles of the in vitro release testing methods of biopolymeric particulate system with the recent research articles and discusses their characteristics including IVIVC, accelerated release testing methods, and stability of encapsulated drugs.

Lung-selective 25-hydroxycholesterol nanotherapeutics as a suppressor of COVID-19-associated cytokine storm.

In response to the coronavirus disease-19 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), global efforts are focused on the development of new therapeutic interventions. For the treatment of COVID-19, selective lung-localizing strategies hold tremendous potential, as SARS-CoV-2 invades the lung via ACE2 receptors and causes severe pneumonia. Similarly, recent reports have shown the association of COVID-19 with decreased 25-hydroxycholesterol (25-HC) and increased cytokine levels. This mechanism, which involves the activation of inflammatory NF-κB- and SREBP2-mediated inflammasome signaling pathways, is believed to play a crucial role in COVID-19 pathogenesis, inducing acute respiratory distress syndrome (ARDS) and sepsis. To resolve those clinical conditions observed in severe SARS-CoV-2 patients, we report 25-HC and didodecyldimethylammonium bromide (DDAB) nanovesicles (25-HC@DDAB) as a COVID-19 drug candidate for the restoration of intracellular cholesterol level and suppression of cytokine storm. Our data demonstrate that 25-HC@DDAB can selectively accumulate the lung tissues and effectively downregulate NF-κB and SREBP2 signaling pathways in COVID-19 patient-derived PBMCs, reducing inflammatory cytokine levels. Altogether, our findings suggest that 25-HC@DDAB is a promising candidate for the treatment of symptoms associated with severe COVID-19 patients, such as decreased cholesterol level and cytokine storm.

PEGylated nanoparticle albumin-bound steroidal ginsenoside derivatives ameliorate SARS-CoV-2-mediated hyper-inflammatory responses.

The rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on a global scale urges prompt and effective countermeasures. Recently, a study has reported that coronavirus disease-19 (COVID-19), the disease caused by SARS-CoV-2 infection, is associated with a decrease in albumin level, an increase in NETosis, blood coagulation, and cytokine level. Here, we present drug-loaded albumin nanoparticles as a therapeutic agent to resolve the clinical outcomes observed in severe SARS-CoV-2 patients. PEGylated nanoparticle albumin-bound (PNAB) was used to promote prolonged bioactivity of steroidal ginsenoside saponins, PNAB-Rg6 and PNAB-Rgx365. Our data indicate that the application of PNAB-steroidal ginsenoside can effectively reduce histone H4 and NETosis-related factors in the plasma, and alleviate SREBP2-mediated systemic inflammation in the PBMCs of SARS-CoV-2 ICU patients. The engineered blood vessel model confirmed that these drugs are effective in suppressing blood clot formation and vascular inflammation. Moreover, the animal model experiment showed that these drugs are effective in promoting the survival rate by alleviating tissue damage and cytokine storm. Altogether, our findings suggest that these PNAB-steroidal ginsenoside drugs have potential applications in the treatment of symptoms associated with severe SARS-CoV-2 patients, such as coagulation and cytokine storm.

Blocking microglial activation of reactive astrocytes is neuroprotective in models of Alzheimer's disease.

Alzheimer's disease (AD) is the most common cause of age-related dementia. Increasing evidence suggests that neuroinflammation mediated by microglia and astrocytes contributes to disease progression and severity in AD and other neurodegenerative disorders. During AD progression, resident microglia undergo proinflammatory activation, resulting in an increased capacity to convert resting astrocytes to reactive astrocytes. Therefore, microglia are a major therapeutic target for AD and blocking microglia-astrocyte activation could limit neurodegeneration in AD. Here we report that NLY01, an engineered exedin-4, glucagon-like peptide-1 receptor (GLP-1R) agonist, selectively blocks ß-amyloid (Aß)-induced activation of microglia through GLP-1R activation and inhibits the formation of reactive astrocytes as well as preserves neurons in AD models. In two transgenic AD mouse models (5xFAD and 3xTg-AD), repeated subcutaneous administration of NLY01 blocked microglia-mediated reactive astrocyte conversion and preserved neuronal viability, resulting in improved spatial learning and memory. Our study indicates that the GLP-1 pathway plays a critical role in microglia-reactive astrocyte associated neuroinflammation in AD and the effects of NLY01 are primarily mediated through a direct action on Aß-induced GLP-1R+ microglia, contributing to the inhibition of astrocyte reactivity. These results show that targeting upregulated GLP-1R in microglia is a viable therapy for AD and other neurodegenerative disorders.

Nano Differential Scanning Fluorimetry-Based Thermal Stability Screening and Optimal Buffer Selection for Immunoglobulin G.

Nano differential scanning fluorimetry (nanoDSF) is a high-throughput protein stability screening technique that simultaneously monitors protein unfolding and aggregation properties. The thermal stability of immunoglobulin G (IgG) was investigated in three different buffers (sodium acetate, sodium citrate, and sodium phosphate) ranging from pH 4 to 8. In all three buffers, the midpoint temperature of thermal unfolding (Tm) showed a tendency to increase as the pH increased, but the aggregation propensity was different depending on the buffer species. The best stability against aggregation was obtained in the sodium acetate buffers below pH 4.6. On the other hand, IgG in the sodium citrate buffer had higher aggregation and viscosity than in the sodium acetate buffer at the same pH. Difference of aggregation between acetate and citrate buffers at the same pH could be explained by a protein-protein interaction study, performed with dynamic light scattering, which suggested that intermolecular interaction is attractive in citrate buffer but repulsive in acetate buffer. In conclusion, this study indicates that the sodium acetate buffer at pH 4.6 is suitable for IgG formulation, and the nanoDSF method is a powerful tool for thermal stability screening and optimal buffer selection in antibody formulations.

Long-acting nanoparticulate DNase-1 for effective suppression of SARS-CoV-2-mediated neutrophil activities and cytokine storm.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a new strain of coronavirus not previously identified in humans. Globally, the number of confirmed cases and mortality rates of coronavirus disease 2019 (COVID-19) have risen dramatically. Currently, there are no FDA-approved antiviral drugs and there is an urgency to develop treatment strategies that can effectively suppress SARS-CoV-2-mediated cytokine storms, acute respiratory distress syndrome (ARDS), and sepsis. As symptoms progress in patients with SARS-CoV-2 sepsis, elevated amounts of cell-free DNA (cfDNA) are produced, which in turn induce multiple organ failure in these patients. Furthermore, plasma levels of DNase-1 are markedly reduced in SARS-CoV-2 sepsis patients. In this study, we generated recombinant DNase-1-coated polydopamine-poly(ethylene glycol) nanoparticulates (named long-acting DNase-1), and hypothesized that exogenous administration of long-acting DNase-1 may suppress SARS-CoV-2-mediated neutrophil activities and the cytokine storm. Our findings suggest that exogenously administered long-acting nanoparticulate DNase-1 can effectively reduce cfDNA levels and neutrophil activities and may be used as a potential therapeutic intervention for life-threatening SARS-CoV-2-mediated illnesses.

Bioinspired DNase-I-Coated Melanin-Like Nanospheres for Modulation of Infection-Associated NETosis Dysregulation.

The current outbreak of the beta-coronavirus (beta-Cov) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) began in December 2019. No specific antiviral treatments or vaccines are currently available. A recent study has reported that coronavirus disease 2019 (COVID-19), the disease caused by SARS-CoV-2 infection, is associated with neutrophil-specific plasma membrane rupture, and release excessive neutrophil extracellular traps (NETs) and extracellular DNAs (eDNAs). This mechanism involves the activation of NETosis, a neutrophil-specific programmed cell death, which is believed to play a crucial role in COVID-19 pathogenesis. Further progression of the disease can cause uncontrolled inflammation, leading to the initiation of cytokine storms, acute respiratory distress syndrome (ARDS), and sepsis. Herein, it is reported that DNase-I-coated melanin-like nanospheres (DNase-I pMNSs) mitigate sepsis-associated NETosis dysregulation, thereby preventing further progression of the disease. Recombinant DNase-I and poly(ethylene glycol) (PEG) are used as coatings to promote the lengthy circulation and dissolution of NET structure. The data indicate that the application of bioinspired DNase-I pMNSs reduce neutrophil counts and NETosis-related factors in the plasma of SARS-CoV-2 sepsis patients, alleviates systemic inflammation, and attenuates mortality in a septic mouse model. Altogether, the findings suggest that these nanoparticles have potential applications in the treatment of SARS-CoV-2-related illnesses and other beta-CoV-related diseases.

Dual peptide-dendrimer conjugate inhibits acetylation of transforming growth factor ß-induced protein and improves survival in sepsis.

Sepsis is a potentially fatal complication of infections and there are currently no effective therapeutic options for severe sepsis. In this study, we revealed the secretion mechanism of transforming growth factor ß-induced protein (TGFBIp) that was recently identified as a therapeutic target for sepsis, and designed TGFBIp acetylation inhibitory peptide (TAIP) that suppresses acetylation of lysine 676 in TGFBIp. To improve bioavailability and biodegradation of the peptide, TAIP was conjugated to polyamidoamine (PAMAM) dendrimers. Additionally, the cell-penetrating peptide (CPP) was conjugated to the TAIP-modified PAMAM dendrimers for the intracellular delivery of TGFBIp. The resulting nanostructures, decorated with TAIP and CPP via poly(ethylene glycol) linkage, improved the mortality and organ damage in the septic mouse model and suppressed lipopolysaccharide-activated severe vascular inflammatory responses in endothelial cells. Thus, the dendrimer-based nanostructures for delivery of TAIP using CPP show great promise in practical applications in sepsis therapy.

Emerging PEGylated non-biologic drugs.

Introduction: PEGylation is a well-established technology for improving the therapeutic value of drugs by attaching polyethylene glycol (PEG). The first PEGylated enzyme products appeared on the market in the early 1990s; currently, more than 18 PEGylated products have been approved by Food and Drug Administration, which encompass various classes of drug molecules, such as enzymes, interferons, granulocyte colony-stimulating factors, hormones, antibody fragments, coagulation factors, oligonucleotide aptamers, synthetic peptides, and small organic molecules. Areas covered: While PEGylated products mainly comprise biologic drugs, such as recombinant proteins and enzymes, non-biologic drugs have recently emerged as a target for PEGylation. This review focuses on the recent development of PEGylated non-biologic drugs, such as small organic molecules, synthetic peptides, and aptamers. Expert opinion: Several PEGylated versions of anti-cancer drugs, opioid agonists, glucagon-like peptide-1 receptor agonists, and oligonucleotide aptamers are in active development stage, and it is likely that they will have a dramatic impact on the market. Although some safety concerns about PEG in clinical trials have been recently issued, PEGylation is still a commercially attractive proposition as a half-life extension technology for long-acting drug development.

Simultaneous Determination of Cysteamine and Cystamine in Cosmetics by Ion-Pairing Reversed-Phase High-Performance Liquid Chromatography.

Cysteamine has been used in cosmetics as an antioxidant, a hair straightening agent, and a hair waving agent. However, recent studies indicate that cysteamine can act as an allergen to hairdressers. The objective of this study was to develop and validate a simple and effective reversed-phase high-performance liquid chromatography (RP-HPLC) method for the measurement of cysteamine and its dimer, cystamine. Sodium 1-heptanesulfonate (NaHpSO) was used as an ion-pairing agent to improve chromatographic performance. Separation was performed on a Gemini C18 column (250 mm × 4.6 mm, 5 µm particle size) using a mobile phase composed of 85:15 (v/v) 4 mM NaHpSO in 0.1% phosphoric acid:acetonitrile. UV absorbance was monitored at 215 nm. The RP-HPLC method developed in this study was validated for specificity, linearity, limit of detection, limit of quantitation, precision, accuracy, and recovery. Cysteamine and cystamine were chromatographically resolved from other reducing agents such as thioglycolic acid and cysteine. Extraction using water and chloroform resulted in the recovery for cysteamine and cystamine ranging from 100.2-102.7% and 90.6-98.7%, respectively. This validated RP-HPLC method would be useful for quality control and monitoring of cysteamine and cystamine in cosmetics.

Targeting of dermal myofibroblasts through death receptor 5 arrests fibrosis in mouse models of scleroderma.

Scleroderma is an autoimmune rheumatic disorder accompanied by severe fibrosis in skin and other internal organs. During scleroderma progression, resident fibroblasts undergo activation and convert to α-smooth muscle actin (α-SMA) expressing myofibroblasts (MFBs) with increased capacity to synthesize collagens and fibrogenic components. Accordingly, MFBs are a major therapeutic target for fibrosis in scleroderma and treatment with blocking MFBs could produce anti-fibrotic effects. TLY012 is an engineered human TNF-related apoptosis-inducing ligand (TRAIL) which induces selective apoptosis in transformed cells expressing its cognate death receptors (DRs). Here we report that TLY012 selectively blocks activation of dermal fibroblasts and induces DR-mediated apoptosis in α-SMA+ MFBs through upregulated DR5 during its activation. In vivo, TLY012 reverses established skin fibrosis to near-normal skin architecture in mouse models of scleroderma. Thus, the TRAIL pathway plays a critical role in tissue remodeling and targeting upregulated DR5 in α-SMA+ MFBs is a viable therapy for fibrosis in scleroderma.

Chemical chaperone-conjugated exendin-4 as a cytoprotective agent for pancreatic ß-cells.

Endoplasmic reticulum stress has been considered a major cause of pancreatic ß-cell dysfunction and apoptosis leading to diabetes. Glucagon-like peptide-1 receptor activation and chemical chaperones have been known to reduce endoplasmic reticulum stress and improve ß-cell function and survival. The purpose of this study was to prepare and evaluate the chemical chaperone tauroursodeoxycholic acid-conjugated exendin-4 as a protective agent for pancreatic ß-cells. Mono-tauroursodeoxycholic acid-Lys27-exendin-4 conjugate (TUM1-Ex4) showed better receptor binding affinity than other conjugates with strong in vitro insulinotropic activity in rat pancreatic ß-cells and in vivo hypoglycemic activity in type 2 diabetic db/db mice. In INS-1 cells under endoplasmic reticulum stress induced by thapsigargin, TUM1-Ex4 promoted cell survival in a dose-dependent manner. In western blot analysis, TUM1-Ex4 reduced the expression of the endoplasmic reticulum stress marker GRP78 and phosphorylation of the translation initiation factor eIF2α. These results reveal that TUM1-Ex4 accelerates translational recovery and contributes to ß-cell protection and survival. The present study indicates that the chemical chaperone-coupled glucagon-like peptide-1 receptor agonist is a feasible therapeutic strategy to enhance ß-cell function and survival.

Drying Technologies for the Stability and Bioavailability of Biopharmaceuticals.

Solid dosage forms of biopharmaceuticals such as therapeutic proteins could provide enhanced bioavailability, improved storage stability, as well as expanded alternatives to parenteral administration. Although numerous drying methods have been used for preparing dried protein powders, choosing a suitable drying technique remains a challenge. In this review, the most frequent drying methods, such as freeze drying, spray drying, spray freeze drying, and supercritical fluid drying, for improving the stability and bioavailability of therapeutic proteins, are discussed. These technologies can prepare protein formulations for different applications as they produce particles with different sizes and morphologies. Proper drying methods are chosen, and the critical process parameters are optimized based on the proposed route of drug administration and the required pharmacokinetics. In an optimized drying procedure, the screening of formulations according to their protein properties is performed to prepare a stable protein formulation for various delivery systems, including pulmonary, nasal, and sustained-release applications.