Yeast Cell Wall Particles Research Articles

Tumor lysate particle only vaccine (TLPO) vs. Tumor lysate particle-loaded, dendritic cell vaccine (TLPLDC) to prevent recurrence in resected stage III/IV melanoma patients: Results of a phase I/IIa trial

BackgroundThe autologous tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine is produced from dendritic cells (DC) loaded ex vivo with autologous tumor lysate (TL). TLPLDC has been shown to decrease recurrence in resected Stage III/IV melanoma patients in a Phase IIb trial. The TL particle only (TLPO) vaccine is produced by loading of yeast cell wall particles with autologous TL and direct injection allowing for in vivo DC loading. We have compared the TLPO and TLPLDC vaccines in an embedded Phase I/IIa trial of a larger Phase IIb trial of the TLPLDC vaccine. MethodsPatients rendered clinically disease-free after surgery were randomized 2:1 to receive the TLPO or TLPLDC vaccine and followed for recurrence and death. Patients had scheduled intradermal inoculations at 0, 1, 2, 6, 12, and 18 months after enrollment. Kaplan-Meier and log-rank analysis were used to compare disease-free survival (DFS) and overall survival (OS) in an intention-to-treat (ITT) analysis. ResultsSixty-three patients were randomized, 43 TLPO and 20 TLPLDC. Patients randomized to the TLPO arm were more likely to be female (37.2% vs. 10.0 %, p = 0.026), but otherwise no significant clinicopathological differences were identified. No differences in related adverse events (AE) were found between treatment arms. At a median follow-up of 20.5 months, the DFS (60.8% vs. 58.7 %, p = 0.714) and OS (94.6% vs. 93.8 %, p = 0.966) were equivalent between the TLPO and TLPLDC groups, respectively. No statistical differences were found in subgroup analyses between vaccine types, which accounted for receipt of immunotherapy and the use of G-CSF pre-blood draw. ConclusionsIn a randomized, double-blind Phase I/IIa trial, there were no differences in DFS or OS in resected Stage III/IV melanoma patients receiving adjuvant TLPO versus TLPLDC vaccines. Given manufacturing advantages, further efficacy testing of TLPO is warranted in a Phase III trial.

Integrated oral microgel system ameliorates renal fibrosis by hitchhiking co-delivery and targeted gut flora modulation

BackgroundRenal fibrosis is a progressive process associated with chronic kidney disease (CKD), contributing to impaired kidney function. Active constituents in traditional Chinese herbs, such as emodin (EMO) and asiatic acid (AA), exhibit potent anti-fibrotic properties. However, the oral administration of EMO and AA results in low bioavailability and limited kidney accumulation. Additionally, while oral probiotics have been accepted for CKD treatment through gut microbiota modulation, a significant challenge lies in ensuring their viability upon administration. Therefore, our study aims to address both renal fibrosis and gut microbiota imbalance through innovative co-delivery strategies.ResultsIn this study, we developed yeast cell wall particles (YCWPs) encapsulating EMO and AA self-assembled nanoparticles (NPYs) and embedded them, along with Lactobacillus casei Zhang, in chitosan/sodium alginate (CS/SA) microgels. The developed microgels showed significant controlled release properties for the loaded NPYs and prolonged the retention time of Lactobacillus casei Zhang (L. casei Zhang) in the intestine. Furthermore, in vivo biodistribution showed that the microgel-carried NPYs significantly accumulated in the obstructed kidneys of rats, thereby substantially increasing the accumulation of EMO and AA in the impaired kidneys. More importantly, through hitchhiking delivery based on yeast cell wall and positive modulation of gut microbiota, our microgels with this synergistic strategy of therapeutic and modulatory interactions could regulate the TGF-β/Smad signaling pathway and thus effectively ameliorate renal fibrosis in unilateral ureteral obstruction (UUO) rats.ConclusionIn conclusion, our work provides a new strategy for the treatment of renal fibrosis based on hitchhiking co-delivery of nanodrugs and probiotics to achieve synergistic effects of disease treatment and targeted gut flora modulation.Graphical

Edible antimicrobial yeast-based coating with basil essential oil for enhanced food safety

Foodborne pathogen contamination is recognized as a critical public health burden. The application of conventional antimicrobial strategies is significantly restricted in the food industry due to concerns over potential impacts on food quality and human health safety. Herein, this study developed a scalable methodology for encapsulating basil essential oil (BO) into yeast cell wall particles (YCWPs), creating edible antimicrobial coating suspensions for effective pathogen control on real food matrices. Thermal stability and long-term durability of the antimicrobial BO were enhanced after being encapsulated into the YCWPs, which served as a protective barrier, reducing the evaporation of small molecules in the BO mixture. In addition, benefiting from the specific affinity between the YCWP microcarriers and pathogens, the encapsulated BO with a high local concentration around the pathogens could avoid the unexpected consumption caused by organic pollutants encountered in the food system. The obtained BO@YCWPs demonstrated promising antimicrobial efficiency, even under high chemical oxygen demand (COD) conditions, achieving approximately 6 log CFU reductions of S. aureus and E. coli within 30 and 15 min, respectively. Moreover, the antimicrobial BO@YCWPs were suspended in an aqueous solution and then spray-coated onto the fresh basil leaves, significantly inhibiting the proliferation of various microorganisms and demonstrating the great potential application of the BO@YCWPs in ensuring food safety.

Prospective, randomized, double-blind phase 2B trial of the TLPO and TLPLDC vaccines to prevent recurrence of resected stage III/IV melanoma: a prespecified 36-month analysis

BackgroundThe tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine is made by ex vivo priming matured autologous dendritic cells (DCs) with yeast cell wall particles (YCWPs) loaded with autologous tumor lysate...

Cell-based carriers incorporated antimicrobial coatings on diverse food contact surfaces for preventing cross-contamination of fresh produce

Cross-contamination is a key food safety risk during both harvesting and post-harvest processing of fresh produce. Very few effective intervention approaches are available for reducing cross-contamination risks from diverse food contact surfaces during harvesting and post-harvest processing. Therefore, developing an effective antimicrobial coating that can be simply applied on a diversity of food contact surfaces is highly desirable. This work demonstrates a cost-effective and one-step antimicrobial coating approach that can be applied on diverse food contact surfaces, such as plastic films, Sparta brushes, and polyfoam sponges. The incorporation of chlorine charged antimicrobial yeast cell wall particles (YCWPs) into chitosan-based coatings provides unique advantages in achieving a high concentration of chlorine on food contact surface and extending the stability of chlorine. Approximately 583 nmol/cm2 of active chlorine bound to the coated poly(vinyl alcohol-co-ethylene) (PVA-co-PE) film surface after 60 min of incubation with bleach at room temperature, which enabled a significant antimicrobial activity of this coating. An inactivation of approximately 3-log and 5-log units of Listeria innocua with organic load (COD = 500 mg/L) were achieved after treatment for 2 min and 5 min, respectively. This antimicrobial coating with chlorine charged YCWPs effectively reduce the cross-contamination between fresh produce and plastic films. Baby spinach leaves were selected as a model fresh produce. The results represent that chitosan-YCWPs coatings led to more than 3 log reduction in the bacterial population transferred to leaf surface upon 10 min of contact with contaminated plastic films, while native chitosan without chlorine charged YCWPs coating did not result in any significant reduction. The results in this study also demonstrate the feasibility of using this antimicrobial coating on contaminated sponges and brushes to reduce cross-contamination of fresh produce during washing steps. No bacterial cells were detected from the leaf surface after contact with contaminated sponges or brush samples coated with chitosan and chlorine-charged YCWPs. This cost-effective and one-step antimicrobial coating approach with a high potential of scale-up will significantly enhance the efficacy of surface sanitation and prevent the cross-contamination of fresh produce.

Safety and efficacy of autologous tumor lysate particle-loaded dendritic cell vaccination in combination with systemic therapies in patients with recurrent and metastatic melanoma.

Immunotherapy has revolutionized the treatment of melanoma, yet survival remains poor for patients with metastatic disease. The autologous tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine has been shown to be safe adjuvant therapy for patients with resected stage III/IV melanoma who complete the primary vaccine series. Here, we describe an open-label trial of patients with metastatic melanoma treated with TLPLDC vaccine in addition to standard of care (SoC) therapies. The TLPLDC vaccine is created by loading autologous tumor lysate into yeast cell wall particles, which are phagocytosed by autologous dendritic cells ex vivo. Patients who recurred while enrolled in a phase IIb trial of adjuvant TLPLDC vaccine (crossover cohort) and patients with measurable metastatic melanoma cohort were offered TLPLDC vaccine along with SoC therapies. Tumor response was measured by RECIST 1.1 criteria. Overall survival (OS) and progression-free survival (PFS) were estimated by intention-to-treat analysis. Fifty-four patients were enrolled (28 in crossover cohort; 26 in metastatic melanoma cohort). The vaccine was well-tolerated with no grade ≥3 adverse events when given with SoC therapies to include checkpoint inhibitors, BRAF/MEK inhibitors, tyrosine kinase inhibitors, intralesional therapy and/or radiation. In the crossover arm, OS was 76.5% and PFS was 57.1% (median follow-up of 13.9 months). In the metastatic melanoma arm, OS was 85.7% and PFS was 52.2% (median follow-up 8.5 months). The TLPLDC vaccine is well-tolerated and safe in combination with SoC therapies. Future trials will determine the efficacy of TLPLDC in combination with SoC therapies in metastatic melanoma.

Incorporation of Antimicrobial Bio-Based Carriers onto Poly(vinyl alcohol-co-ethylene) Surface for Enhanced Antimicrobial Activity.

A biobased rechargeable antimicrobial modification approach was developed using a covalent immobilization of food grade yeast cell wall particles on a model plastic film. We demonstrate the applications of this modification approach on poly(vinyl alcohol-co-ethylene) surface to inactivate inoculated bacteria with or without the presence of organic content, reducing the cross-contamination between food contact surface and model fresh produce, and inhibiting the growth of biofilms on the film surface. These biobased cell wall particle modified plastic films can enhance the binding of chlorine to the plastic surface in the form of N-halamine, extend the stability of chlorine against high organic content and ambient storage, and improve the rechargeability of the plastic films. Upon charging with chlorine, these modified plastic films inactivated 5 log of model Gram-negative bacteria (Escherichia coli O157:H7) and Gram-positive bacteria (Listeria innocua used as a surrogate of pathogenic Listeria monocytogenes) within 2 min of surface inoculation in water and within 20 min in an organic-rich aqueous environment. The modified plastic films prevented the transfer of bacteria and eliminated cross-contamination from the contaminated films to a spinach leaf surface, while 3 log CFU/leaf of bacteria were transferred from a contaminated native film to a noninoculated spinach surface. In addition, these modified plastic films reduced the adhesion of L. innocua cells by 2.7-3.6 log CFU/cm2 compared with control films during extended incubation for biofilm formation. Overall, this study demonstrates the feasibility of this biobased food grade modification approach to reduce microbial contamination and improve produce safety in the food processing industry.

The influence of harvest method on dendritic cell function and clinical outcomes in a randomized trial of a dendritic cell vaccine to prevent recurrences in high-risk melanoma.

9548 Background: A randomized, double-blind, placebo-controlled phase IIb trial of the tumor lysate, particle loaded, dendritic cell (TLPLDC) vaccine was conducted to prevent recurrence in patients (pts) with resected stage III/IV melanoma. Two methods for dendritic cell (DC) harvest were used for vaccine production, including no pretreatment or pretreatment with granulocyte-colony stimulating factor (G-CSF) in an attempt to reduce blood draw volumes. This analysis investigates differences in clinical outcomes and RNA gene expression between these DC harvest methods for TLPLDC vaccine creation. Methods: The TLPLDC vaccine is created by loading autologous tumor lysate into yeast cell wall particles (YCWPs) and exposing them to phagocytosis by DCs. By investigator/pt choice, pts had 120mL of blood drawn for DC harvest, or pts received 300μg of G-CSF for pre-DC mobilization and a 50-70 mL blood draw 24-48 hours later. Total vaccine production time was 72 hrs. Pts were randomized 2:1 to receive TLPLDC or placebo (DCs exposed to empty YCWPs). 1-1.5 x10^6 cells/dose were injected intradermally at 0, 1, 2, 6, 12, and 18 months. Differences in disease free survival (DFS) and overall survival (OS) were evaluated by Kaplan Meier analysis between pts who did not receive pretreatment (TLPLDC), pts who did receive pretreatment with G-CSF (TLPLDC+G), and pts receiving placebo. RNA-seq analysis was performed on the total RNA of pts’ prepared TLPLDC vaccines to assess gene expression. Relative RNA expression (RRE) was compared between TLPLDC and TLPLDC+G. Results: As previously reported, 144 pts were randomized: 103 received TLPLDC (46 TLPLDC, 57 TLPLDC+G) and 41 received placebo. There were no significant clinicopathologic or treatment differences between the three treatment arms. Survival was significantly improved in TLPLDC compared to TLPLDC+G or placebo, including 36-month OS (92.9% vs 62.8% vs 72.3% respectively, p = 0.022) and DFS (51.8% vs 23.4% vs 27.1%, p = 0.027). When compared to TLPLDC+G (n = 3) vaccine, RNA-seq from TLPLDC vaccine (n = 3) showed upregulation of genes associated with DC maturation, including HLA-DMB (RRE: 3.60), IFIT1 (3.38), CD27 (3.26), IFI44L (3.24), MX1 (2.96), HLA-DQA1 (2.67), HLA-DRA (2.40), CD49D (2.34) and CD74 (2.09), while downregulated genes were associated with DC suppression or immaturity including SERPINA1 (RRE:7.8), TLR2 (6.65), CCR1 (5.11), IL10 (4.19), CD93 (3.84) and CD14 (3.25). Conclusions: Pts receiving TLPLDC vaccine had significantly improved OS and DFS, while outcomes remained similar between those who received TLPLDC+G vs placebo. Pts who did not receive G-CSF had higher expression of genes linked to DC maturation and antigen processing and presentation, likely explaining the improvement in clinical efficacy. A phase III trial to further assess the TLPLDC vaccine to prevent recurrence is planned. Clinical trial information: NCT02301611.

Utilization of PLGA nanoparticles in yeast cell wall particle system for oral targeted delivery of exenatide to improve its hypoglycemic efficacy

Oral delivery of exenatide (EXE), a high-efficiency therapeutic peptide, is urgently needed for long-term treatment of diabetes. In this study, a polylactide-co-glycoside (PLGA) nanoparticles (NPs) in yeast cell wall particle (YCWP) system was built to improve the intestinal absorption of EXE by efficient protection of EXE against gastrointestinal degradation and intestinal phagocytic cell targeted delivery. The EXE-loaded PLGA NPs were prepared by a double emulsion solvent diffusion method and exhibited a uniformly spherical appearance, a nano size (92.4 ± 4.6 nm) and a positive surface charge (+32.3 ± 3.8 mV). And then, the NPs were successfully loaded into the YCWPs by a solvent hydration − lyophilization cycle method to obtain the EXE-PLGA NPs @YCWPs, which was verified by scanning electron microscope and confocal laser scanning microscopy. An obvious sustained drug release and a reduced burst release were achieved by this nano-in-micro carrier. Moreover, the gastrointestinal stability of EXE in PLGA NPs @YCWPs was significantly higher than that in PLGA NPs in the simulated gastrointestinal environment, which were useful in enhancing the intestinal absorption of EXE. In biodistribution study, the EXE-PLGA NPs @YCWPs could quickly reached the root of the villi, and even partly entered the inner of the villi, especially in ileum and Peyer’s patches. In vitro cell evaluation demonstrated an efficient β-glucan receptor mediated endocytosis and transport of EXE-PLGA NPs @YCWPs by the macrophage RAW 264.7 cells, suggesting a potential intestinal macrophage targeted absorptive pathway. The in vivo pharmacokinetic study showed a preferred hypoglycemic effect and an increased pharmacological availability (13.7 ± 4.1%) after oral administration of the EXE-PLGA NPs @YCWPs. It is believed that the PLGA nanoparticles in YCWP system could become an efficient strategy to orally deliver therapeutic peptide drugs.

Liquid-Phase Peak Force Infrared Microscopy for Chemical Nanoimaging and Spectroscopy.

Peak force infrared (PFIR) microscopy is an emerging atomic force microscopy that bypasses Abbe's diffraction limit in achieving chemical nanoimaging and spectroscopy. The PFIR microscopy mechanically detects the infrared photothermal responses in the dynamic tip-sample contact of peak force tapping mode and has been applied for a variety of samples, ranging from soft matters, photovoltaic heterojunctions, to polaritonic materials under the air conditions. In this article, we develop and demonstrate the PFIR microscopy in the liquid phase for soft matters and biological samples. With the capability of controlling fluid compositions on demand, the liquid-phase peak force infrared (LiPFIR) microscopy enables in situ tracking of the polymer surface reorganization in fluids and detecting the product of click chemical reaction in the aqueous phase. Both broadband spectroscopy and infrared imaging with ∼10 nm spatial resolution are benchmarked in the fluid phase, together with complementary mechanical information. We also demonstrate the LiPFIR microscopy on revealing the chemical composition of a budding site of yeast cell wall particles in water as an application on biological structures. The label-free, nondestructive chemical nanoimaging and spectroscopic capabilities of the LiPFIR microscopy will facilitate the investigations of soft matters and their transformations at the solid/liquid interface.

Targeted Photodynamic Treatment of Bacterial Biofilms Using Curcumin Encapsulated in Cells and Cell Wall Particles.

Efficient inactivation and removal of pathogenic biofilms in food and biomedical environments remain a significant challenge for food safety applications and medical facilities. This research aims to develop food-grade microcarriers for the targeted delivery of a photosensitizer, curcumin, and photodynamic inactivation of a model pathogenic bacterial biofilm. The microcarriers evaluated in this study include alive yeast cell carriers, deactivated yeast cell carriers, and yeast cell wall particles. The microcarriers were evaluated based on the encapsulation yield of a model photosensitizer (curcumin), binding of the microcarriers to biofilms, and inactivation of the bacteria in the biofilms. The results illustrate that the combination of binding affinity, encapsulation yield, and the intracellular composition of the microcarriers influenced the overall inactivation of bacteria in the biofilms. All of the selected compositions achieved more than 93% inactivation of the bacteria in the biofilm using the photodynamic treatment, and the yeast cell wall particles with curcumin achieved over 99% inactivation of the bacteria in the biofilm matrix. In addition, all of the selected compositions demonstrated significant potential to remove the biofilm from the plastic surface, suggesting the role of binding affinity of the microcarriers in removal of the biofilm from surfaces. Overall, this study developed biomaterial formulations for targeted photodynamic inactivation and potential removal of biofilms.

Abstract 6536: Safety and efficacy of autologous tumor lysate particle loaded dendritic cell (TLPLDC) vaccination in combination with systemic therapies in patients with metastatic melanoma

Abstract Background: The treatment of melanoma has changed drastically with the advent of immunotherapy, particularly checkpoint inhibition (CPI). Unfortunately, only 50-60% of tumors respond to current immunotherapy. The autologous tumor lysate, particle loaded, dendritic cell (TLPLDC) vaccine utilizes the whole antigenic panel of the patient's tumor to stimulate T-cells and may work synergistically with other immunotherapies. We have previously demonstrated both safety and efficacy of TLPLDC vaccine as adjuvant therapy for patients (pts) with resected stage III or IV melanoma who complete the primary vaccine series. Here, we describe results in pts with metastatic melanoma treated with the TLPLDC vaccine together with other approved therapies. Methods: The TLPLDC vaccine is created by loading autologous tumor lysate into yeast cell wall particles which are then phagocytosed by autologous dendritic cells ex-vivo. Pts who recurred while enrolled in a randomized phase IIb trial of adjuvant TLPLDC vaccine (crossover arm) and pts with metastatic melanoma with measurable disease (MM arm) were offered TLPLDC vaccine in an open-label fashion in addition to other approved therapies as determined by their treating physician. The vaccine is given via intradermal injection monthly x4 followed by boosters at six and nine months. Tumor response is measured by RECIST 1.1 criteria. Results: To date, 53 pts have been enrolled (28 in the crossover arm and 25 in the MM arm). Within the crossover arm, 18 pts had received TLPLDC as part of the phase IIb trial, 8 were from the placebo group, and 2 patients withdrew prior to vaccination. Within the MM arm, 2 were excluded for no evidence of disease at time of vaccination and 1 withdrew. The vaccine was well tolerated, with 24% of patients experiencing a treatment-related adverse event (AE), with no grade ≥3 AE. Concurrent therapies were varied in the crossover arm, but 36% received CPI, compared with 68% in the MM arm. Of those receiving CPI, 59% had >1 immune-related AE, none of which were grade ≥3. In the crossover arm, 12-month overall survival (OS) was 83.3% in the vaccine-naïve cohort at median follow-up of 6.0 months vs. 94.7% in the re-vaccinated cohort at median follow-up of 9.0 months (p=0.961). In the MM arm, 12-month OS was 61.2% at a median follow-up of 7.7 months. In a pooled analysis of 33 evaluable pts, vaccine-naïve pts (n=21) had a 42.9% non-progression rate compared to a 66.7% non-progression rate in re-vaccinated pts (n=12). Conclusion: Vaccination with the TLPLDC vaccine in combination with approved systemic therapies in MM pts is well tolerated, with acceptable rates of immune-related AE when combined with CPI. Additionally, vaccination, and specifically re-vaccination, may provide clinical benefit particularly if used in combination with other approved therapies. Citation Format: Robert Connor Chick, Annelies T. Hickerson, Guy Travis Clifton, Phillip M. Kemp Bohan, Tommy A. Brown, Jessica L. Cindass, John W. Myers, Timothy J. Vreeland, Doreen O. Jackson, Diane F. Hale, Kaitlin M. Peace, Garth S. Herbert, Xianzhong Yu, Thomas Wagner, George E. Peoples. Safety and efficacy of autologous tumor lysate particle loaded dendritic cell (TLPLDC) vaccination in combination with systemic therapies in patients with metastatic melanoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6536.

Simultaneous Nanoscale Imaging of Chemical and Architectural Heterogeneity on Yeast Cell Wall Particles.

Particles extracted from yeast cell walls are naturally occurring immunomodulators with significant therapeutic applications. Their biological function has been thought to be a consequence of the overall chemical composition. In contrast, here we achieve direct nanoscale visualization of the compositional and structural heterogeneity of yeast cell wall particles and demonstrate that such nanoscale heterogeneity directly influences the receptor function of immune cells. By combining peak force infrared (PFIR) microscopy with super-resolution fluorescence microscopy, we achieve simultaneous chemical, topographical, and mechanical mapping of cell wall particles extracted from the yeast Saccharomyces cerevisiae with ≈6 nm resolution. We show that polysaccharides (β-glucan and chitin) and proteins are organized in specific nonuniform structures, and their heterogeneous spatial organization leads to heterogeneous recruitment of receptors on immune cell membranes. Our findings indicate that the biological function of yeast cell wall particles depends on not only their overall composition but also the nanoscale distribution of the different cell wall components.

Multi-institutional, prospective, randomized, double-blind, placebo-controlled phase IIb trial of the tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine to prevent recurrence in high-risk melanoma patients: A subgroup analysis.

63 Background: A novel vaccine strategy may prevent recurrence in high-risk melanoma patients (pts). The TLPLDC vaccine uses yeast cell wall particles (YCWP) to load tumor lysate into autologous dendritic cells (DC). In this phase IIb trial of TLPLDC vs. placebo in resected stage III/IV pts, TLPLDC increased 24 month (mo) disease free survival (DFS) in the per treatment (PT) population. Here, we present a 24mo DFS subgroup analysis and estimated overall 36mo DFS. Methods: Disease-free pts were randomized 2:1 to the TLPLDC vaccine vs. unloaded YCWP+DC at 0, 1, 2, 6, 12, and 18mo. The protocol was amended to allow concurrent adjuvant checkpoint inhibitor (CPI) therapy once approved. The pre-specified PT population included only pts completing the primary vaccine/placebo series (PVS) at 6 mo. Kaplan-Meier estimates of DFS were used to compare treatment arms by stage (III or IV) and CPI therapy (yes/no) in the ITT and PT populations. Results: 144 pts were randomized (103 TLPLDC, 41 placebo); 98 pts (66 TLPLDC, 32 placebo) completed the PVS. There were no clinicopathologic differences between treatment groups. There was no difference in 24mo DFS in stage III pts (n = 112), but in stage IV pts (n = 32), the 24mo DFS was 44% vs 0% (TLPLDC vs placebo) (p = 0.41) in ITT and 73.3% vs. 0% (HR 0.14, p = 0.002) in PT. Stage IV pts were more likely to receive CPI than stage III pts (50% vs. 30%, p = 0.003). There was no difference in 24mo DFS for pts who did not receive CPI (n = 102), but in pts who received CPI (n = 42), the 24mo DFS was 49.3% vs. 31.3% (p = 0.71) in ITT and 68.8% vs. 41.7% (HR 0.46, p = 0.28) in PT, showing a trend toward improved DFS in pts who completed the PVS and received CPI (n = 31). Overall, the 36mo estimated DFS was 34.2% vs. 21.6% (p = 0.89) for ITT and 56.9% vs. 27.9% (p = 0.021) for PT. Conclusions: The TLPLDC vaccine improved DFS in patients completing the PVS at 24 and 36 mos, particularly in the resected stage IV subset. The apparent synergistic effect with TLPLDC + CPI will be confirmed in a phase III study evaluating adjuvant TLPLDC + CPI vs. CPI alone in resected stage IV melanoma pts. Clinical trial information: NCT02301611.

Yeast Cell wall Particle mediated Nanotube-RNA delivery system loaded with miR365 Antagomir for Post-traumatic Osteoarthritis Therapy via Oral Route.

Post-traumatic osteoarthritis (PTOA) is an acute injury-induced joint inflammation followed by a gradual degradation of articular cartilage. However, there is no FDA-approved Disease-Modifying Osteoarthritis Drug currently. Although gene therapy with microRNA can improve PTOA progression, there is no effective gene delivery vehicle for orally deliver therapeutics due to the harsh environment of the gastrointestinal tract. In this study, we investigated the effect of yeast cell wall particle (YCWP) mediated nanotube-RNA delivery system on PTOA therapy via oral route.Methods: Nontoxic and degradable AAT and miRNA365 antagomir was self-assembled into miR365 antagomir/AAT (NPs). Then NPs-YCWP oral drug delivery system was constructed by using NPs and non-pathogenic YCWP which can be specifically recognized by macrophages. Moreover, surgical destabilization of the medial meniscus induced PTOA mice model was established to evaluate the therapeutic effect of NPs-YCWP via oral route.Results: Compared with control group, NPs showed higher gene inhibition efficiency both in chondrogenic cell line and primary chondrocytes in vitro. Treatment of macrophages with fluorescently labeled NPs-YCWP, the results showed that NPs-YCWP was successfully engulfed by macrophages and participated in the regulation of gene expression in vitro. Under the protection of YCWP, miR365 antagomir/AAT passes through the gastrointestinal tract without degradation after oral administration. After NPs-YCWP therapy, the results of histological staining, gene and protein expression showed that miR365 antagomir/NPs-YCWP improved the symptom of PTOA.Conclusion: Here, we constructed a biodegradable drug delivery system based on non-pathogenic YCWP and nanotubes, which can be used for PTOA therapy via the oral route. It suggests a new gene therapy strategy with YCWP mediated oral nano drug delivery system may serve as a platform for joint degeneration treatment.

Oral treatment with T6-loaded yeast cell wall particles reduces the parasitemia in murine visceral leishmaniasis model

Yeast cell wall particles isolated from Saccharomyces cerevisiae (scYCWPs) have a rich constitution of β-glucan derived from the cell wall. After removing intracellular contents, β-glucan molecules are readily recognized by dectin-1 receptors, present on the cytoplasmic membrane surface of the mononuclear phagocytic cells and internalized. Leishmania spp. are obligate intracellular parasites; macrophages are its primary host cells. An experimental murine model of visceral leishmaniasis caused by L. infantum was used to evaluate the antileishmanial activity of oral administration of these particles. A low-water soluble thiophene previously studied in vitro against L. infantum was entrapped into scYCWPs to direct it into the host cell, in order to circumvent the typical pharmacokinetic problems of water-insoluble compounds. We found that scYCWPs + T6 reduced the parasitic burden in the liver and spleen. There was an increase in IFN-γ levels related to nitric oxide production, explaining the reduction of the L. infantum burden in the tissue. Histological analysis did not show signals of tissue inflammation and biochemical analysis from plasma did not indicate signals of cytotoxicity after scYCWPs + T6 treatment. These findings suggested that scYCWPs + T6 administered through oral route reduced the parasitic burden without causing toxic effects, satisfying requirements for development of new strategies to treat leishmaniasis.

Bioaccessibility of curcumin encapsulated in yeast cells and yeast cell wall particles

This study evaluates the role of intracellular composition and integrity of cell walls in modulating the release and bioaccessibility of phytochemicals during simulated gastrointestinal digestion of cell-based carriers. Native yeast and yeast cell wall particles (YCWPs) were used as model cell-based carriers with distinct intracellular composition and curcumin was a model encapsulated bioactive. The results highlight the essential role of gastric treatments and the presence of bile salts in the release and bioaccessibility of encapsulated compound from cell-based carriers. YCWPs with significantly reduced intracellular contents have a significantly faster intestinal release after gastric digestion. Multimodal imaging approaches confirm the release of encapsulated curcumin in intestinal compartment without any significant changes in the cellular structure including cell walls. Based on these results, the study proposes a model that illustrate the significance of gastric digestion in influencing the release of curcumin during gastrointestinal digestion.

Initial Safety and Production Efficacy of a Autologous Tumor Lysate Loaded Yeast Cell Wall Particle Vaccine

Initial Safety and Production Efficacy of a Autologous Tumor Lysate Loaded Yeast Cell Wall Particle Vaccine

Abstract CT208: Prior vaccination with the autologous Tumor Lysate Particle Loaded Dendritic Cell (TLPLDC) Vaccine may impact clinical outcomes in melanoma patients treated with systemic therapies and re-vaccination

Abstract Background: Melanoma is an immunogenic cancer, yet only 50-60% of tumors respond to current immunotherapy. The autologous tumor lysate, particle loaded, dendritic cell (TLPLDC) vaccine may potentiate an immune response by stimulating T-cells. We are conducting a phase IIb double-blinded randomized trial of TLPLDC to prevent recurrence after resection. Patients who recurred were offered enrollment in an open-label TLPLDC vaccine trial along with standard therapy. Here, we describe characteristics and clinical outcomes of recurred patients that continued on open label TLPLDC vaccination. Methods: Patients with resected no evidence of disease (NED) stage III/IV melanoma were randomized to TLPLDC vs. empty yeast cell wall particle (YCWP) loaded DC in a 2:1 fashion. TLPDC is created by loading autologous tumor lysate into YCWP. This is then introduced ex vivo to the patient’s DC for phagocytosis. 1-1.5x106TLPLDCs are given via intradermal injection monthly x4 followed by boosters at 6 and 9 months. Patients who recurred after receiving TLPLDC (vaccine group, VG) or after empty YCWP (control group, CG) on the trial (primary endpoint) were offered open label TLPLDC vaccination with same dosing schedule along with standard therapy as determined by the patient’s treatment team. Disease status is measured by RECIST criteria. Results: To date, 23 (14 VG and 11 CG) patients have enrolled and been vaccinated, with median follow up (fu) of 7.86 months (mo). The patients received the following additional therapies: surgery, gamma-knife, check point inhibitor, radiation, TVEC, BRAF/MEK, and imantinib. No related toxicities > grade 2 were observed. Of 14 VG patients, 5 were NED when re-enrolled, 9 had measurable disease (MD). Of these, 11 have followed up data, 9 (82%) show disease control (5/5 NED patients remain NED, 4/6 with MD show SD), while 2 (18%) have progressed. Of 9 CG patients, 4 were NED and 5 had measurable disease at time of re-enrollment. Of these, 8 have followed up, 5 (62%) show disease control (2/4 NED remain NED, 1/4 with MD shows regression, 2/4 SD), while 3 (38%) have progressed (2/4 NED, 1/3 with MD). Conclusion: Open label administration of TLPLPDC vaccine after recurrence is demonstrated as a safe therapy and shows potential clinical benefit when combined SOC therapy. Further study is warranted to determine if prior exposure to TLPLDC may provide clinical benefit after the first episode of disease recurrence. Citation Format: Annelies T. Hickerson, Guy T. Clifton, Tommy A. Brown, Jessica L. Cindass, John W. Myers, Timothy J. Vreeland, Diane F. Hale, Kaitlin M. Peace, Doreen O. Jackson, Garth Herbert, Xianzhong Yu, Thomas E. Wagner, George E. Peoples. Prior vaccination with the autologous Tumor Lysate Particle Loaded Dendritic Cell (TLPLDC) Vaccine may impact clinical outcomes in melanoma patients treated with systemic therapies and re-vaccination [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT208.

Activity and Cell-Death Pathway in Leishmania infantum Induced by Sugiol: Vectorization Using Yeast Cell Wall Particles Obtained From Saccharomyces cerevisiae.

Visceral leishmaniasis, caused by Leishmania infantum, is a neglected tropical disease, to which efforts in the innovation of effective and affordable treatments remain limited, despite the rising incidence in several regions of the world. In this work, the antileishmanial effects of sugiol were investigated in vitro. This compound was isolated from the bark of Cupressus lusitanica and showed promising activity against L. infantum. In spite of the positive results, it is known that the compound is a poorly water-soluble diterpene molecule, which hinders further investigation, especially in preclinical animal studies. Thus, in an alternative delivery method, sugiol was entrapped in glucan-rich particles obtained from Saccharomyces cerevisiae yeast cell walls (YCWPs). To evaluate the activity of sugiol, the experiments were divided into two parts: (i) the in vitro investigation of antileishmanial activity of free sugiol against L. infantum promastigotes after 24, 48, and 72 h of treatment and (ii) the evaluation of antileishmanial activity of sugiol entrapped in glucan-rich particles against intracellular L. infantum amastigotes. Free sugiol induced the cell-death process in promastigotes, which was triggered by enhancing cytosolic calcium level and promoting the autophagy up to the first 24 h. Over time, the presence of autophagic vacuoles became rarer, especially after treatment with lower concentrations of sugiol, but other cellular events intensified, like ROS production, cell shrinkage, and phosphatidylserine exposure. Hyperpolarization of mitochondrial membrane potential was found at 72 h, induced by the mitochondria calcium uptake, causing an increase in ROS production and lipid peroxidation as a consequence. These events resulted in the cell death of promastigotes by secondary necrosis. Sugiol entrapped in glucan-rich particles was specifically recognized by dectin-1 receptor on the plasma membrane of macrophages, the main host cell of Leishmania spp. Electron micrographs revealed particles containing sugiol within the infected macrophages and these particles were active against the intracellular L. infantum amastigotes without affecting the host cell. Therefore, the YCWPs act like a Trojan horse to successfully deliver sugiol into the macrophage, presenting an interesting strategy to deliver water-insoluble drugs to parasitized cells.