Recombinant Human Growth Factor Research Articles

A single dose of VEGF-A circular RNA sustains in situ long-term expression of protein to accelerate diabetic wound healing

Diabetic foot ulcer (DFU), which is characterised by damage to minute blood vessels or capillaries around wounds, is one of the most serious and dreaded complications of diabetes. It is challenging to repair chronic non-healing DFU wounds. Vascular endothelial growth factor (VEGF) plays an important role in angiogenesis and promotes wound healing in DFU. However, it is difficult to sustainably deliver VEGF to the wound site owing to its poor stability and easy degradation. To overcome this challenge, lipid nanoparticles (LNP) encapsulating circular RNA (circRNA) encoding VEGF-A have been developed to continuously generate and release VEGF-A and accelerate diabetic wound healing. First, VEGF-A circRNA was synthesized using group I intron autocatalysis strategy and confirmed by enzyme digestion, polymerase chain reaction, and sequencing assay. VEGF-A circRNA was encapsulated in ionizable lipid U-105-derived LNP (U-LNP) using microfluidic technology to fabricate U-LNP/VEGF-A circRNA. For comparison, a commercially ionizable lipid ALC-0315-derived LNP (A-LNP) encapsulating circRNA (A-LNP/circRNA) was used. Dynamic light scattering and transmission electron microscopy characterization indicated that U-LNP/circRNA had spherical structure with an average diameter of 108.5 nm, a polydispersity index of 0.22, and a zeta potential of -3.31 mV. The messenger RNA (mRNA) encapsulation efficiency (EE%) of U-LNP was 87.12%. In vitro transfection data confirmed better stability and long-term VEGF-A expression of circRNA compared with linear mRNA. Assessment of cytotoxicity and innate immunity further revealed that U-LNP/circRNA was biocompatible and induced a weak congenital immune response. Cell scratch and angiogenesis tests demonstrated the bioactivity of U-LNP/VEGF-A circRNA owing to its VEGF-A expression. In situ bioluminescence imaging of firefly luciferase (F-Luc) probe and ELISA demonstrated that circRNA had long-term and strong expression of VEGF-A in the first week, and a gradual decrease in the next week at the wound site and surrounding areas. Finally, a diabetic mouse model was used to validate the healing effect of U-LNP/VEGF-A circRNA formulation. The results showed that a single dose of U-LNP/VEGF-A circRNA administered by dripping resulted in almost complete wound recovery on day 12, which was significantly superior to that of U-LNP/VEGF-A linear mRNA, and it also outperformed recombinant human vascular endothelial growth factor (rhVEGF) injection and A-LNP/circRNA dripping. Histological analysis confirmed the healing efficiency and low toxicity of U-LNP/VEGF-A circRNA formulation. Together, VEGF-A circRNA delivered by U-105-derived LNP showed good performance in wound healing, which was ascribed to the long-term expression and continuous release of VEGF-A, and has potential applications for the treatment of diabetic foot ulcer wounds.

Enhancing diabetic wound healing through anti-bacterial and promoting angiogenesis using dual-functional slow-release microspheres-loaded dermal scaffolds

Bacterial infections and the degeneration of the capillary network comprise the primary factors that contribute to the delayed healing of diabetic wounds. However, treatment modalities that cater to effective diabetic wounds healing in clinical settings are severely lacking. Herein, a dual-functional microsphere carrier was designed, which encapsulates polyhexamethylene biguanide (PHMB) or recombinant human vascular endothelial growth factor (rhVEGF) together. The in vitro release experiments demonstrated that the use of the microspheres ensured the sustained release of the drugs (PHMB or rhVEGF) over a period of 12 days. Additionally, the integration of these controlled-release microspheres into a dermal scaffold (DS-PLGA@PHMB/rhVEGF) imbued both antibacterial and angiogenic functions to the resulting material. Accordingly, the DS-PLGA@PHMB/rhVEGF scaffold exhibited potent antibacterial properties, effectively suppressing bacterial growth and providing a conducive environment for wound healing, thereby addressing the drawbacks associated with the susceptibility of rhVEGF to deactivation in inflammatory conditions. Additionally, the histological analysis revealed that the use of the DS-PLGA@PHMB/rhVEGF scaffold accelerated the process of wound healing by inhibiting inflammatory reactions, stimulating the production of collagen formation, and enhancing angiogenesis. This provides a novel solution for enhancing the antibacterial and vascularization capabilities of artificial dermal scaffolds, providing a beacon of hope for improving diabetic wound healing.

Zinc Iodide Dimethyl Sulfoxide Reduces Collagen Deposition by Increased Matrix Metalloproteinase-2 Expression and Activity in Lung Fibroblasts.

Chronic inflammatory lung diseases are characterized by disease-specific extracellular matrix accumulation resulting from an imbalance of matrix metalloproteinases (MMPs) and their inhibitors. Zinc is essential for the function of MMPs, and zinc deficiency has been associated with enhanced tissue remodeling. This study assessed if zinc iodide (ZnI) supplementation through dimethyl sulfoxide (DMSO) modifies the action of MMPs in isolated human lung fibroblasts. The expression and activity of two gelatinases, MMP-2 and MMP-9, were determined by gelatin zymography and enzyme-linked immuno-sorbent assay (ELISA). Collagen degradation was determined by cell-based ELISAs. Collagen type I and fibronectin deposition was stimulated by human recombinant tumor growth factor β1 (TGF-β1). Untreated fibroblasts secreted MMP-2 but only minute amounts of MMP-9. TGF-β1 (5 ng/mL) reduced MMP-2 secretion, but stimulated collagen type I and fibronectin deposition. All the effects of TGF-β1 were significantly reduced in cells treated with ZnI-DMSO over 24 h, while ZnI and DMSO alone had a lower reducing effect. ZnI-DMSO alone did not increase MMP secretion but enhanced the ratio of active to inactive of MMP-2. ZnI alone had a lower enhancing effect than ZnI-DMSO on MMP activity. Furthermore, MMP-2 activity was increased by ZnI-DMSO and ZnI in the absence of cells. Soluble collagen type I increased in the medium of ZnI-DMSO- and ZnI-treated cells. Blocking MMP activity counteracted all the effects of ZnI-DMSO. Conclusion: The data suggest that the combination of ZnI with DMSO reduces fibrotic processes by increasing the degradation of collagen type I by up-regulating the activity of gelatinases. Thus, the combination of ZnI with DMSO might be considered for treatment of fibrotic disorders of the lung. DMSO supported the beneficial effects of ZnI.

Injectable bioactive poly(propylene fumarate) and polycaprolactone based click chemistry bone cement for spinal fusion in rabbits.

Degenerative spinal pathology is a widespread medical issue, and spine fusion surgeries are frequently performed. In this study, we fabricated an injectable bioactive click chemistry polymer cement for use in spinal fusion and bone regrowth. Taking advantages of the bioorthogonal click reaction, this cement can be crosslinked by itself eliminating the addition of a toxic initiator or catalyst, nor any external energy sources like UV light or heat. Furthermore, nano-hydroxyapatite (nHA) and microspheres carrying recombinant human bone morphogenetic protein-2 (rhBMP-2) and recombinant human vascular endothelial growth factor (rhVEGF) were used to make the cement bioactive for vascular induction and osteointegration. After implantation into a rabbit posterolateral spinal fusion (PLF) model, the cement showed excellent induction of new bone formation and bridging bone, achieving results comparable to autograft control. This is largely due to the osteogenic properties of nano-hydroxyapatite (nHA) and the released rhBMP-2 and rhVEGF growth factors. Since the availability of autograft sources is limited in clinical settings, this injectable bioactive click chemistry cement may be a promising alternative for spine fusion applications in addressing various spinal conditions.

Abstract 3178: Vitamin B1 derivative, fursultiamine, prevents colon cancer growth

Abstract Colon cancer is the second deadliest type of cancer in the United States, with surgical resection being the primary treatment modality for localized early-stage colon cancer. Still, more treatment and prevention options are needed to help improve the outcomes for colon cancer as they are currently limited to invasive procedures. One promising option is fursultiamine, a disulfide lipid-soluble vitamin B1 derivative, which has anti-oxidative and anti-inflammatory activity that could aid in preventing colon cancer. We hypothesize that supplementation of fursultiamine could be an effective chemopreventive treatment to control the growth and spread of colon carcinogenesis. Human colorectal adenocarcinoma (SW480) cell lines were incubated with complete media (CM) or human recombinant endothelial growth factor (EGF 20 ng/ml) in the absence and presence of fursultiamine. The cell viability was examined by MTT assay in a concentration- (0-150 uM) and time (24 and 48 h) - dependent manner. Our results indicate that fursultiamine prevents SW480 cell death in a dose-dependent manner. Apoptosis was detected by using Annexin-V staining and live and death cell assay. Our results suggest that fursultiamine prevents SW480 cell proliferation by inducing apoptotic cell death. The production of reactive oxygen species (ROS) and activation of caspase-3 were examined by specific assay kits. Treatment with fursultiamine resulted in the decreased production of ROS and activation of caspase-3. The expression of anti-apoptotic, pro-apoptotic, and pro-inflammatory factors was analyzed using antibody arrays. The results demonstrated that treatment with fursultiamine increased the expression of pro-apoptotic factors such as Bad, Bax, cleaved caspase 3, Fas/CD95, and FADD. Further, fursultiamine increased expression of the tumor suppressor genes p53, p27, and p21. We next planned to examine this vitamin B1 derivative's effect in preventing cancer growth in nude mice xenograft models. In conclusion, our in vitro results indicate that fursultiamine, a vitamin B1 derivative, prevents colon cancer growth through increased regulation of pro-apoptotic pathways and tumor suppressor proteins and thus has the potential for further development as a chemopreventative agent. Citation Format: Sudeep Poludasu, Christian Pompoco, Kota Ramana. Vitamin B1 derivative, fursultiamine, prevents colon cancer growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3178.

Bioactive Moldable Click Chemistry Polymer Cement with Nano-Hydroxyapatite and Growth Factor-Enhanced Posterolateral Spinal Fusion in a Rabbit Model.

Spinal injuries or diseases necessitate effective fusion solutions, and common clinical approaches involve autografts, allografts, and various bone matrix products, each with limitations. To address these challenges, we developed an innovative moldable click chemistry polymer cement that can be shaped by hand and self-cross-linked in situ for spinal fusion. This self-cross-linking cement, enabled by the bioorthogonal click reaction, excludes the need for toxic initiators or external energy sources. The bioactivity of the cement was promoted by incorporating nanohydroxyapatite and microspheres loaded with recombinant human bone morphogenetic protein-2 and vascular endothelial growth factor, fostering vascular induction and osteointegration. The release kinetics of growth factors, mechanical properties of the cement, and the ability of the scaffold to support in vitro cell proliferation and differentiation were evaluated. In a rabbit posterolateral spinal fusion model, the moldable cement exhibited remarkable induction of bone regeneration and effective bridging of spine vertebral bodies. This bioactive moldable click polymer cement therefore presents a promising biomaterial for spinal fusion augmentation, offering advantages in safety, ease of application, and enhanced bone regrowth.

Development of a prototype of a theranostic system based on silica nanoparticles with immobilized fluorescent dyes and VEGF targeting ligand.

Background. Administration of certain drugs causes undesirable effects associated with the systemic effect of the active substance on the entire body. Selective targeting of the drug to the affected tissue promotes a selective increase in the concentration of the substance in the area of interest, thereby reducing the systemic effect and enhancing the local therapeutic effect.Objective. Development of a targeted delivery system for theranostic agents using recombinant human vascular endothelial growth factor type A (rhVEGF-A121) as a targeting ligand.Design and method. To create the theranostic complex, commercially available reagents were used: the recombinant protein rhVEGF-A121 (cat.#: PSG140-10, LLC CyStorLab, Skolkovo, Russia) and fumed silica Aerosil (A-200, Degussa AG, Germany). The tosyl spacer that interconnects both components was synthesized in the laboratory. Protein conjugation with fluorophores was also carried out in-house. Indocyanine green (ICG; Sigma-Aldrich, USA) and rhodamine B (JSC Lenreaktiv, St. Petersburg, Russia) were taken for immobilization.Results. In the course of the work, functionalization of silica nanoparticles (SiNPs) with a tosyl spacer was carried out, conjugates of SiNPs with rhVEGF-A121 were synthesized, and theranostic constructs based on SiNPs were obtained, including rhVEGF-A121 as a targeting ligand, and ICG/Rhodamine B as a visualizing label.Conclusion. In the presented study, a prototype of a complex for targeted delivery of a theranostic agent to tissues with an active angiogenesis process, for example, to tumor and ischemic tissues, was developed. To solve the problem, we immobilized on the surface of SiNP a recombinant protein of human vascular endothelial growth factor (rhVEGF) to use as a guide ligand. Such a synthetic construct will help to deliver diagnostic and/ or medicinal substances packed in SiNP directly to cells that overexpress extracellular specific receptors of the VEGFR family. In subsequent in vivo experiments, delivery efficiency will be assessed by tissue accumulation of the fluorophores ICG and rhodamine B, which have been conjugated to the targeting ligand protein. The physicochemical characteristics of the obtained samples were studied by the methods of spectrophotometry and dynamic light scattering.

Evaluating the efficacy of recombinant human growth factors in scar remodelling for patients with facial soft tissue injuries.

Facial soft tissue injuries, often resulting in scarring, pose a challenge in reconstructive and aesthetic surgery due to the need for functional and aesthetic restoration. This study evaluates the efficacy of recombinant human growth factors (rhGFs) in scar remodelling for such injuries. A retrospective evaluation was conducted from January 2020 to January 2023, involving 100 patients with facial soft tissue injuries. Participants were divided equally into a control group, receiving standard cosmetic surgical repair, and an observation group, treated with rhGFs supplemented cosmetic surgery. The study assessed scar characteristics (pigmentation, pliability, vascularity, height), hospital stay duration, tissue healing time, complication rates and patient satisfaction. The observation group demonstrated significant improvements in all scar characteristics, with notably better pigmentation, pliability, vascularity and height compared with the control group. The rhGF treatment also resulted in reduced hospital stay duration and faster tissue healing. Notably, the total complication rate was significantly lower in the observation group (10%) compared with the control group (34%). Additionally, patient satisfaction levels were higher in the observation group, with 98% combined satisfaction compared with 76% in the control group. The application of rhGFs in treating facial soft tissue injuries significantly enhances scar remodelling, expedites healing, reduces complications and improves patient satisfaction. These findings establish rhGFs as a valuable tool in the management of facial soft tissue injuries, highlighting their potential in improving both functional and aesthetic outcomes.

Preconditioning Single High-Dose Palifermin Alters the Posttransplant Inflammatory Cytokine Profile

Palifermin is a truncated form of human recombinant keratinocyte growth factor (KGF) that binds to the FGF receptor 2b expressed in epithelia including that of the epidermis, oral and GI mucosa, and thymus. In animal models, the cytoprotective and regenerative effects of KGF showed efficacy in controlling acute and chronic graft-versus-host disease (A/C GVHD), but these were not demonstrated in subsequent clinical studies using the FDA-approved dosing schedule for oral mucositis prevention (60 µg/kg/day for 3 consecutive days). The current study aims to investigate alterations in immune cell reconstitution and cytokine expression among patients (n=31) participating in the ongoing prospective open-label phase 1/2 trial NCT02356159. This trial evaluates the incidence of severe CGVHD (primary objective of phase 2) following the addition of a single high dose of palifermin to GVHD prophylaxis with TMS (tacrolimus, methotrexate, sirolimus). Four different dose levels of palifermin (DL1: 180 µg/kg; DL2: 360 µg/kg; DL3: 540 µg/kg; DL4: 720 µg/kg) were tested on day −7 in the phase 1 part of the trial, with the recommended phase 2 dose being 720 µg/kg. All patients received cyclophosphamide (total 4.8 g/m 2) and fludarabine (total 120 mg/m 2) as conditioning on days −6 to −3 followed by infusion of an 8/8 HLA-matched unrelated donor peripheral blood cell graft on day 0. Results of correlative studies were compared with patients of the identical TMS treatment arm of the NCT00520130 study without palifermin (n=31). In the DL4 group, a previous interim analysis revealed no cases of classic (day +100) AGVHD II-IV in 16 patients (P=0.014), whereas the TMS group showed a 22.6% (95% CI: 9.8% to 38.6%) incidence (Schulz et al. Cell Ther Transplant. 2023;29(2):S258). T, B and NK cells were routinely measured by flow cytometry at days +14, +28, +60, +100, +180, +365, +730. The V-PLEX human cytokine 36-plex kit and U-PLEX assays of human BAFF, CXCL9 and IL1RL1/ST2 (all MSD, Rockville, MD) were used to measure cytokine levels in plasma at days −8, 0, +14, +28, +60, +100. The measures at various time points were compared between two treatment groups using multiple Mann-Whitney tests, and the false discovery rate approach was applied with Q=0.05. Comparison between the four DL groups and the TMS group was performed using the Kruskal-Wallis H-test and Dunn's post-hoc test, with a significance threshold set at P<0.05. The statistical analyses were conducted using GraphPad Prism 9. In comparison to patients receiving TMS only, those treated with KGF exhibited reduced NK cell counts at day +28 (Q=0.0155) and increased B cell counts from day +60 (Q<0.01) onwards. However, there were no differences in the absolute numbers of CD3+, CD4+, and CD8+ T cells between patients from both trials. Additionally, no apparent dose-response relationship was observed. KGF-treated patients showed significantly reduced plasma levels of proinflammatory cytokines, including TNF-α at day 0, TNF-β at days 0, +14, +28, IL-12/IL23p40 at days 0 and +14, and CXCL9 at days 0 and +14, compared to patients receiving TMS only. Notably, levels of homeostatic IL-7, previously linked to AGVHD, were significantly lower in KGF-treated patients from day +28 (TMS vs. DL4, P=0.002) to +100. There was a trend towards lower levels of IL-22 at days 0 and +14 (both unadjusted P<0.05, Q>0.05) in KGF-treated patients. Other proinflammatory biomarkers showed increased levels in KGF-treated patients: IL-15 at day 0 (Q=0.0121), BAFF at days +14 and +28 (both Q<0.01), and ST2 at day 0 and day +14 (both Q<0.01). As reported previously by others, patients who developed classic or late onset AGVHD II-IV without prior relapse up to day +180 had higher ST2 levels at day +14 compared to patients without AGVHD II-IV (P=0.03). Here, this association was mainly observed in patients from the TMS and DL1 groups (6/8 tested patients with any AGVHD II-IV). The administration of a single high dose of palifermin (KGF) prior to conditioning treatment resulted in a notably distinct cytokine profile, characterized by a reduction in several GVHD associated pro-inflammatory cytokines, consistent with KGF's proposed cytoprotective effect. Further investigations focusing on GI microbiome and immune cell subsets are needed to elucidate the consequences of cytokine alterations induced by KGF on immune cell reconstitution. This research was supported by the Intramural Research Program of the NIH, NCI, CCR.

Клінічний випадок успішного лікування пацієнта з перфорацією дивертикула сигмовидної кишки, ускладненого сепсисом на тлі COVID-19

описано випадок успішного лікування пацієнта, у якого після перенесеного COVID-19виникла перфорація дивертикула сигмовидної кишки, що ускладнилась перитонітом та сепсисом,спочатку бактеріальним, потім грибковим. На фоні проведеної консервативної терапії в перед іпісляопераційному періоді та частого застосування нестероїдних протизапальних препаратів,зокрема метамізолу натрію у хворого розвинулась важка лейкопенія та тромбоцитопенія, щобули частково рефрактерними до глюкокортикоїдів. Через відсутності можливості призначеннярекомбінантних факторів росту у даного пацієнта був застосований нестандартний підхід якийполягав у призначенні ціанокобаламіну в комбінації з фолієвою кислотою, що в комплексі з іншимизаходами дозволили нормалізувати рівень лейкоцитів та тромбоцитів.

Twenty-five years of recombinant human growth factors rhPDGF-BB and rhBMP-2 in oral hard and soft tissue regeneration.

Contemporary oral tissue engineering strategies involve recombinant human growth factor approaches to stimulate diverse cellular processes including cell differentiation, migration, recruitment, and proliferation at grafted areas. Recombinant human growth factor applications in oral hard and soft tissue regeneration have been progressively researched over the last 25 years. Growth factor-mediated surgical approaches aim to accelerate healing, tissue reconstruction, and patient recovery. Thus, regenerative approaches involving growth factors such as recombinant human platelet-derived growth factor-BB (rhPDGF-BB) and recombinant human bone morphogenetic proteins (rhBMPs) have shown certain advantages over invasive traditional surgical approaches in severe hard and soft tissue defects. Several clinical studies assessed the outcomes of rhBMP-2 in diverse clinical applications for implant site development and bone augmentation. Current evidence regarding the clinical benefits of rhBMP-2 compared to conventional therapies is inconclusive. Nevertheless, it seems that rhBMP-2 can promote faster wound healing processes and enhance de novo bone formation, which may be particularly favorable in patients with compromised bone healing capacity or limited donor sites. rhPDGF-BB has been extensively applied for periodontal regenerative procedures and for the treatment of gingival recessions, showing consistent and positive outcomes. Nevertheless, current evidence regarding its benefits at implant and edentulous sites is limited. The present review explores and depicts the current applications, outcomes, and evidence-based clinical recommendations of rhPDGF-BB and rhBMPs for oral tissue regeneration.

Recombinant VEGF-C (Cys156Ser) improves mesenteric lymphatic drainage and gut immune surveillance in experimental cirrhosis

Lymphatic vessels (LVs) are crucial for maintaining abdominal fluid homoeostasis and immunity. In cirrhosis, mesenteric LVs (mLVs) are dilated and dysfunctional. Given the established role of vascular endothelial growth factor-C (VEGF-C) in improving LVs, we hypothesised that VEGF-C treatment could ameliorate the functions of mLVs in cirrhosis. In this study, we developed a nanoformulation comprising LV-specific growth factor, recombinant human VEGF-C (Cys156Ser) protein (E-VEGF-C) and delivered it orally in different models of rat cirrhosis to target mLVs. Cirrhotic rats were given nanoformulation without VEGF-C served as vehicles. Drainage of mLVs was analysed using tracer dye. Portal and systemic physiological assessments and computed tomography were performed to measure portal pressures and ascites. Gene expression and permeability of primary mesenteric lymphatic endothelial cells (LyECs) was studied. Immune cells in mesenteric lymph nodes (MLNs) were quantified by flow cytometry. Endogenous and exogenous gut bacterial translocation to MLNs was examined. In cirrhotic rats, mLVs were dilated and leaky with impaired drainage. Treatment with E-VEGF-C induced proliferation of mLVs, reduced their diameter, and improved functional drainage. Ascites and portal pressures were significantly reduced in E-VEGF-C rats compared with vehicle rats. In MLNs of E-VEGF-C animals, CD8+CD134+ T cells were increased, whereas CD25+ regulatory T cells were decreased. Both endogenous and exogenous bacterial translocation were limited to MLNs in E-VEGF-C rats with reduced levels of endotoxins in ascites and blood in comparison with those in vehicle rats. E-VEGF-C treatment upregulated the expression of vascular endothelial-cadherin in LyECs and functionally improved the permeability of these cells. E-VEGF-C treatment ameliorates mesenteric lymph drainage and portal pressure and strengthens cytotoxic T-cell immunity in MLNs in experimental cirrhosis. It may thus serve as a promising therapy to manage ascites and reduce pathogenic gut bacterial translocation in cirrhosis. A human recombinant pro-lymphangiogenic growth factor, VEGF-C, was encapsulated in nanolipocarriers (E-VEGF-C) and orally delivered in different models of rat liver cirrhosis to facilitate its gut lymphatic vessel uptake. E-VEGF-C administration significantly increased mesenteric lymphatic vesselproliferation and improved lymph drainage, attenuating abdominal ascites and portal pressures in the animal models. E-VEGF-C treatment limited bacterial translocation to MLNs only with reduced gut bacterial load and ascitic endotoxins. E-VEGF-C therapy thus holds the potential to manage ascites and portal pressure and reduce gut bacterial translocation in patients with cirrhosis.

Development of a Silk Fibroin-Small Intestinal Submucosa Small-Diameter Vascular Graft with Sequential VEGF and TGF-β1 Inhibitor Delivery for In Situ Tissue Engineering.

Proper endothelialization and limited collagen deposition on the luminal surface after graft implantation plays a crucial role to prevent the occurrence of stenosis. To achieve these conditions, a biodegradable graft with adequate mechanical properties and the ability to sequentially deliver therapeutic agents isfabricated. In this study, a dual-release system is constructed through coaxial electrospinning by incorporating recombinant human vascular endothelial growth factor (VEGF) and transforming growth factor β1 (TGF-β1) inhibitor into silk fibroin (SF) nanofibers to form a bioactive membrane. The functionalized SF membrane as the inner layer of the graft is characterized by the release profile, cell proliferation and protein expression. It presents excellent biocompatibility and biodegradation, facilitating cell attachment, proliferation, and infiltration. The core-shell structure enables rapid VEGF release within 10 days and sustained plasmid delivery for 21 days. A 2.0-mm-diameter vascular graft is fabricated by integrating the SF membrane with decellularized porcine small intestinal submucosa (SIS), aiming to facilitate the integration process under a stable extracellular matrix structure. The bioengineered graft is functionalized with the sequential administration of VEGF and TGF-β1, and with the reinforced and compatible mechanical properties, thereby offers an orchestrated solution for stenosis with potential for in situ vascular tissue engineering applications.

A phase Ib trial of the VEGF fusion protein HB002.1T plus chemotherapy in patients with advanced solid tumors.

2530 Background: HB002.1T is a recombinant human vascular endothelial growth factor receptor (VEGF)-IgG Fc fusion protein, which functions as a decoy receptor to bind VEGF-A, VEGF-B and placental growth factor (PlGF). In phase Ia, HB002.1T monotherapy showed encouraging anti-tumor activity in gallbladder cancer. Methods: This study aimed to evaluate efficacy and safety of HB002.1T plus chemotherapy (mainly based on platinum, paclitaxel, gemcitabine, etc.) in advanced solid tumors. The tumor assessment was evaluated by investigator according to RECIST v1.1. The study included part 1 and part 2, part 1 study was dose escalation (12mg/kg, 16mg/kg Q3W) and dose expansion conducted in patients (pts) with advanced solid tumors, part 2 study was dose expansion (16mg/kgQ3W) in specified tumors (cohort 1: ovarian and cervical, cohort 2: pancreatic, gallbladder and bile duct cancer). Results: As of Dec 20,2022,68 (42F/26M) advanced solid tumor pts were enrolled in the study with 32 pts in part 1 and 36 pts in part 2. 33.3% pts failed at least 3 lines of therapy before enrollment. 63(92.6%) pts had stage IV disease. Median age was 58. It is to be note that 12 efficacy assessment evaluablepts with ovarian cancer (1 with 12mg/kg in part 1,11with 16mg/kg in part 2) were all stage IV, which included 3 complete response (CR),6 partial responses (PR) and 3 stable disease (SD). Median progression free survival (PFS) was 7.3 months and 6-month PFS rate was 58.3%. The ORR and DCR were 75%, 100% respectively. The most common treatment related adverse events (TRAEs) included neutropenia (9/12, 75.0%) and leukocytopenia (8/12, 66.7%). 8 (8/12, 66.7%) pts had grade ≥ 3 TRAEs, which neutropenia (4/12, 33.3%) and leukocytopenia (3/12, 25.0%) were commonly observed. Part 1: In 28 pts whose tumor assessment were available, including 1 gastric cancer patient achieved CR, 11 pts achieved PR (2 gastric, 2 ovarian, 2 cervical, 1 maxillary, 1 lung, 1 parotid, 1 nasopharynx, 1 pancreatic) and 15 achieved SD. Part 2: 32 pts were available for tumor assessment, including 4 achieved CR (3 ovarian, 1 cervical), 10 achieved PR (4 ovarian, 2 cervical, 3 bileduct, 1 pancreatic), and 16 achieved SD. Overall, in Part 1 and 2, TRAEs occurred in 64 (64/68, 94.1%) pts. The most common TRAEs were leukocytopenia (28/68, 41.2%), neutropenia (26/68, 38.2%) and proteinuria (25/68, 36.8%). 34 (34/68, 50.0%) pts had grade ≥ 3 TRAEs, which neutropenia (15/68, 22.1%) and leukocytopenia (11/68, 16.2%) were commonly observed. TRAE leading to discontinuation occurred in 3 pts. No death was reported related with HB002.1T. Conclusions: The Pts were well tolerated to 12mg/kg and 16mg/kg HB002.1T dose combined with chemotherapy, and HB002.1T showed acceptable safety profile. As well as, promising anti-tumor activity was observed in various tumors, especially in advanced ovarian cancer. More data is being accumulated, and confirmatory clinical trials are being prepared. Clinical trial information: NCT04802980 .

Abstract 174: Organoids developed from synovial sarcoma patient-derived xenografts

Abstract Background: Soft tissue sarcoma (STS) is a heterogeneous family of rare malignancies. Despite this heterogeneity, systemic treatment options for patients with advanced STS are rather uniform and have very limited efficacy. There is an unmet medical need to develop more active, subtype-specific treatment options. Three-dimensional (3D) cell culture models (organoids) have been successfully used for drug testing in carcinoma research. However, the development of STS organoids is lagging behind. We attempted to identify the best growth conditions for organoids derived from synovial sarcoma (SySa), which is one of the more common translocation-related subtypes of STS. Methods: Human synovial sarcoma tissue was implanted subcutaneously into immunodeficient mice and grown to create a patient-derived xenograft (PDX). Fresh tumor tissue from the PDX model was collected and enzymatically and mechanically dissociated into small cell clusters. The cells were seeded into a 3D scaffold with advanced Dulbecco's modified Eagle’s medium (DMEM)/F12 and different growth factors were added. In order to obtain optimal results, different scaffolds and growth factors were compared. The growth of the 3D cultures was evaluated by bright-field microscopy and a cell viability assay using PrestoBlue. The organoids were characterized by histopathological stainings and the presence of the SSX-SS18 translocation was molecularly confirmed. Finally, we attempted to create organoids from PDX-derived material stored in liquid nitrogen. Results: A scaffold consisting of 50% collagen type 1, 30% Matrigel, 20% DMEM 10x and sodium hydroxide was superior to Matrigel alone. The addition of human recombinant epidermal growth factor (EGF, 10 ng/ml), human insulin-like growth factor 1 (IGF-1, 20 ng/ml), human recombinant fibroblast growth factor 2 (FGF-2, 10 ng/ml), and N-acetylcysteine (NAC, 500 mmol) to advanced DMEM/F12 stimulated the growth of SySa organoids the best. The Rho-kinase inhibitor Y-27632 was added to prevent apoptosis during early passages. So far, attempts to create organoids from frozen material have been unsuccessful. Conclusions: At present, the success rate of establishing organoids from synovial sarcoma cells is limited if the culture conditions are not adapted to their needs. We identified that growth factors such as EGF, FGF-2, IGF-1, and NAC are beneficial for growth. The availability of fresh samples plays an important role in the development of these 3D models. Despite these limitations, organoids can be an interesting tool for studying this rare disease and aid in the development of new drugs. Citation Format: Lore De Cock, Agnieszka Wozniak, Kimberly Verbeeck, Patrick Schöffski. Organoids developed from synovial sarcoma patient-derived xenografts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 174.

Albumin-fused long-acting FGF21 analogue for the treatment of non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease (NAFLD) currently affects about 25% of the world's population, and the numbers continue to rise as the number of obese patients increases. However, there are currently no approved treatments for NAFLD. This study reports on the evaluation of the therapeutic effect of a recombinant human serum albumin-fibroblast growth factor 21 analogue fusion protein (HSA-FGF21) on the pathology of NAFLD that was induced by using two high-fat diets (HFD), HFD-60 and STHD-01. The HFD-60-induced NAFLD model mice with obesity, insulin resistance, dyslipidemia and hepatic lipid accumulation were treated with HSA-FGF21 three times per week for 4weeks starting at 12weeks after the HFD-60 feeding. The administration of HSA-FGF21 suppressed the increased body weight, improved hyperglycemia, hyperinsulinemia, and showed a decreased accumulation of plasma lipid and hepatic lipid levels. The elevation of C16:0, C18:0 and C18:1 fatty acids in the liver that were observed in the HFD-60 group was recovered by the HSA-FGF21 administration. The increased expression levels of the hepatic fatty acid uptake receptor (CD36) and fatty acid synthase (SREBP-1c, FAS, SCD-1, Elovl6) were also suppressed. In adipose tissue, HSA-FGF21 caused an improved adipocyte hypertrophy, a decrease in the levels of inflammatory cytokines and induced the expression of adiponectin and thermogenic factors. The administration of HSA-FGF21 to the STHD-01-induced NAFLD model mice resulted in suppressed plasma ALT and AST levels, oxidative stress, inflammatory cell infiltration and fibrosis. Together, HSA-FGF21 has some potential for use as a therapeutic agent for the treatment of NAFLD.

THERAPY OF PERIODONTITIS WITH VASCULAR ENDOTHELIAL GROWTH FACTOR BASED ON HYDROGEL: IMMUNOHISTOCHEMICAL CHARACTERISTICS

Regeneration of lost periodontal tissues is the ultimate goal of periodontal treatment. One of the promising approaches to stimulating periodontal regeneration is the use of hydrogels and various growth factors that contribute to the colonization of the scaffold with fibroblast populations, followed by the formation of mature connective tissue of the periodontal ligament. Patients with chronic generalized periodontitis of mild and moderate severity (n=70, age 49-69 years) were treated in the Department of maxillofacial surgery of the UKB No. 4. Treatment was carried out with a preparation from a complex of bioresorbable carbohydrates and recombinant human vascular endothelial growth factor (VEGF-A). The biopsy material was taken immediately before the introduction of the drug into the periodontal pocket and 14 days after the procedure. Prior to treatment, pronounced chronic inflammation with a predominance of lymphocytes, moderate edema and divergence of the fibers of the stromal component were noted in the gum preparations, signs of chronic moderate active inflammation were observed. After treatment, collagen fibers constructed from type III collagen and type I collagen were found in the structured fibers of the intercellular substance, which is characteristic of the formed connective tissue. Thus, the therapy of chronic periodontitis with a hydrogel with VEGF-A is a promising direction for further research, since it has the potential to influence not only stimulating the regeneration of periodontal tissues, but also the effect of local anti-inflammatory treatment.

Long non-coding RNA PVT1 promotes the proliferation, migration and EMT process of ovarian cancer cells by regulating CTGF.

Ovarian cancer remains one of the most common gynecological malignancies with a poor prognosis. The present study investigated the roles of long non-coding RNA plasmacytoma variant translocation 1 (lncRNA PVT1) in the regulation of the malignant phenotype of ovarian cancer cells, including cell proliferation, migration, invasion and epithelial-mesenchymal transition (EMT). SKOV3 and CAOV3 cells were transfected with small interfering RNA (siRNA) targeting lncRNA PVT1 (si-PVT1) or control siRNA and the si-PVT1 transfected cells were co-cultured with recombinant human connective tissue growth factor (rhCTGF). The proliferation, migration and invasion abilities of the cells were examined via Cell Counting Kit-8, colony formation, wound-healing and Transwell assays. The relative expression levels of lncRNA PVT1, CTGF, E-cadherin and vimentin were analyzed using reverse transfection-quantitative polymerase chain reaction, and western blotting was employed to detect the protein levels of CTGF, E-cadherin and vimentin. The expression of lncRNA PVT1 was significantly reduced in SKOV3 and CAOV3 cells following transfection with si-PVT1. In addition, the proliferation, migration and invasion abilities of SKOV3 and CAOV3 cells were repressed following lncRNA PVT1 knockdown. The knockdown of lncRNA PVT1 also reduced the expression of CTGF and vimentin, and increased the expression of E-cadherin. The changes in the proliferation, migration and invasion of the cells induced by transfection with si-PVT1 were partially attenuated in the presence of rhCTGF. Furthermore, co-culture with rhCTGF reversed the si-PVT1-induced changes in the expression of EMT-associated proteins. In conclusion, lncRNA PVT1 promotes the proliferation, migration, invasiveness and EMT process of ovarian cancer cells, and CTGF contributes to the effect of lncRNA PVT1.

Optimizing cell proliferation and function for immunotherapy with recombinant growth factors and cytokines

As more and new immune cell therapies continue to be developed, the demand for Animal Component-Free (ACF) and GMP-quality recombinant human growth factors and cytokines has increased. Cytokine therapies have become extremely popular due to their effectiveness in the treatment of cancer and autoimmune conditions.The use of GMP products is the gold standard for Cell & Gene therapy manufacturing. Especially the use of GMP recombinant human interleukins as companies prepare for clinical trials or scale-up manufacturing of their immune cell therapies. Interleukins (ILs) are a group of cytokines that regulate immune and inflammatory responses. Interleukins also regulate cell growth, differentiation, and motility. The discovery of Interleukin (IL-2) as a “T-cell growth factor” (TCGF) in 1976 revolutionized the fields of basic immunology research and immunotherapy for human cancers.

A Net Meta-Analysis of the Effectiveness of Different Types of Dressings in the Treatment of Diabetic Foot.

Objective This is an analysis of the impact of a new dressing commonly used to treat diabetic foot (DFU). Methods Chinese and English databases were searched to collect clinical randomized controlled studies (RCTs) of various types of dressings for the treatment of DFU, and the healing rates of the different dressings were combined by reticulation meta-analysis. Results The aggregate of the 36 RCTs included in this study analysed the healing rates of nine dressings: conventional dressing, alginate dressing, chitosan dressing, hyaluronic acid dressing, platelet-rich plasma dressing, amniotic membrane dressing, honey dressing, human recombinant growth factor dressing, and silver ionomer dressing. Conclusion Hyaluronic acid dressing, amniotic membrane dressing, honey dressing, and platelet-rich plasma dressing are the ideal materials for topical treatment of DFU.