Search results for: signaling cascade

Authors: Rajeswari Raguraman, Sowmya Parameswaran, Krishnakumar Subramanian, Jagat Kanwar, Rupinder Kanwar

Abstract:

Background: Tumor microenvironment has been implicated in several cancers to regulate cell growth, invasion and metastasis culminating in outcome of therapy. Tumor stroma consists of multiple cell types that are in constant cross-talk with the tumor cells to favour a pro-tumorigenic environment. Not much is known about the existence of tumor microenvironment in the pediatric intraocular malignancy, Retinoblastoma (RB). In the present study, we aim to understand the multiple stromal cellular subtypes and tumor stromal interactions expressed in RB tumors. Materials and Methods: Immunohistochemistry for stromal cell markers CD31, CD68, alpha-smooth muscle (α-SMA), vimentin and glial fibrillary acidic protein (GFAP) was performed on formalin fixed paraffin embedded tissues sections of RB (n=12). The differential expression of stromal target molecules; fibroblast activation protein (FAP), tenascin-C (TNC), osteopontin (SPP1), bone marrow stromal antigen 2 (BST2), stromal derived factor 2 and 4 (SDF2 and SDF4) in primary RB tumors (n=20) and normal retina (n=5) was studied by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and Western blotting. The differential expression was correlated with the histopathological features of RB. The interaction between RB cell lines (Weri-Rb-1, NCC-RbC-51) and Bone marrow stromal cells (BMSC) was also studied using direct co-culture and indirect co-culture methods. The functional effect of the co-culture methods on the RB cells was evaluated by invasion and proliferation assays. Global gene expression was studied by using Affymetrix 3’ IVT microarray. Pathway prediction was performed using KEGG and the key molecules were validated using qRT-PCR. Results: The immunohistochemistry revealed the presence of several stromal cell types such as endothelial cells (CD31+;Vim+/-); macrophages (CD68+;Vim+/-); Fibroblasts (Vim+; CD31-;CD68- );myofibroblasts (α-SMA+/ Vim+) and invading retinal astrocytes/ differentiated retinal glia (GFAP+; Vim+). A characteristic distribution of these stromal cell types was observed in the tumor microenvironment, with endothelial cells predominantly seen in blood vessels and macrophages near actively proliferating tumor or necrotic areas. Retinal astrocytes and glia were predominant near the optic nerve regions in invasive tumors with sparse distribution in tumor foci. Fibroblasts were widely distributed with rare evidence of myofibroblasts in the tumor. Both gene and protein expression revealed statistically significant (P<0.05) up-regulation of FAP, TNC and BST2 in primary RB tumors compared to the normal retina. Co-culture of BMSC with RB cells promoted invasion and proliferation of RB cells in direct and indirect contact methods respectively. Direct co-culture of RB cell lines with BMSC resulted in gene expression changes in ECM-receptor interaction, focal adhesion, IL-8 and TGF-β signaling pathways associated with cancer. In contrast, various metabolic pathways such a glucose, fructose and amino acid metabolism were significantly altered under the indirect co-culture condition. Conclusion: The study suggests that the close interaction between RB cells and the stroma might be involved in RB tumor invasion and progression which is likely to be mediated by ECM-receptor interactions and secretory factors. Targeting the tumor stroma would be an attractive option for redesigning treatment strategies for RB.

Keywords: gene expression profiles, retinoblastoma, stromal cells, tumor microenvironment

Procedia PDF Downloads 361

Authors: Michelle Maurer, Antonia Last, Mark S. Gresnigt, Bernhard Hube, Alexander S. Mosig

Abstract:

The intestinal epithelium forms an essential barrier to prevent translocation of microorganisms, toxins or other potentially harmful molecules into the bloodstream. In particular, dendritic cells of the intestinal epithelium orchestrate an adapted response of immune tolerance to commensals and immune defense against invading pathogens. Systemic inflammation is typically associated with a dysregulation of this adapted immune response and is accompanied by a disruption of the epithelial and endothelial gut barrier which enables dissemination of pathogens within the human body. To understand the pathophysiological mechanisms underlying the inflammation-associated gut barrier breakdown, it is crucial to elucidate the complex interplay of the host and the intestinal microbiome. A microfluidically perfused three-dimensional intestine-on-chip model was established to emulate these processes in the presence of immune cells, commensal bacteria, and facultative pathogens. Multi-organ tissue flow (MOTiF) biochips made from polystyrene were used for microfluidic perfusion of the intestinal tissue model. The biochips are composed of two chambers separated by a microporous membrane. Each chamber is connected to inlet and outlet channels allowing independent perfusion of the individual channels and application of microfluidic shear stress. Human umbilical vein endothelial cells (HUVECs), monocyte-derived macrophages and intestinal epithelial cells (Caco-2) were assembled on the biochip membrane. Following 7 – 14 days of growth in the presence of physiological flow conditions, the epithelium was colonized with the commensal bacterium Lactobacillus rhamnosus, while the endothelium was perfused with peripheral blood mononuclear cells (PBMCs). Additionally, L. rhamnosus was co-cultivated with the opportunistic fungal pathogen Candida albicans. Within one week of perfusion, the epithelial cells formed self-organized and well-polarized villus- and crypt-like structures that resemble essential morphological characteristics of the human intestine. Dendritic cells were differentiated in the epithelial tissue that specifically responds to bacterial lipopolysaccharide (LPS) challenge. LPS is well-tolerated at the luminal epithelial side of the intestinal model without signs of tissue damage or induction of an inflammatory response, even in the presence of circulating PBMC at the endothelial lining. In contrast, LPS stimulation at the endothelial side of the intestinal model triggered the release of pro-inflammatory cytokines such as TNF, IL-1β, IL-6, and IL-8 via activation of macrophages residing in the endothelium. Perfusion of the endothelium with PBMCs led to an enhanced cytokine release. L. rhamnosus colonization of the model was tolerated in the immune competent tissue model and was demonstrated to reduce damage induced by C. albicans infection. A microfluidic intestine-on-chip model was developed to mimic a systemic infection with a dysregulated immune response under physiological conditions. The model facilitates the colonization of commensal bacteria and co-cultivation with facultative pathogenic microorganisms. Both, commensal bacteria alone and facultative pathogens controlled by commensals, are tolerated by the host and contribute to cell signaling. The human intestine-on-chip model represents a promising tool to mimic microphysiological conditions of the human intestine and paves the way for more detailed in vitro studies of host-microbiota interactions under physiologically relevant conditions.

Keywords: host-microbiota interaction, immune tolerance, microfluidics, organ-on-chip

Procedia PDF Downloads 108

Authors: Jayanwita Sarkar, Usha Chakraborty, Bishwanath Chakraborty

Abstract:

Plants are constantly interacting with various abiotic and biotic stresses. In changing climate scenario plants are continuously modifying physiological processes to adapt to changing environmental conditions which profoundly affect plant-pathogen interactions. Spot blotch in wheat is a fast-rising disease in the warmer plains of South Asia where the rise in minimum average temperature over most of the year already affecting wheat production. Hence, the study was undertaken to explore the role of elevated temperature in spot blotch disease development and modulation of antioxidative responses by plant growth promoting rhizobacteria (PGPR) for biocontrol of spot blotch at high temperature. Elevated temperature significantly increases the susceptibility of wheat plants to spot blotch causing pathogen Bipolaris sorokiniana. Two PGPR Bacillus safensis (W10) and Ochrobactrum pseudogrignonense (IP8) isolated from wheat (Triticum aestivum L.) and blady grass (Imperata cylindrical L.) rhizophere respectively, showing in vitro antagonistic activity against Bipolaris sorokiniana were tested for growth promotion and induction of resistance against spot blotch in wheat. GC-MS analysis showed that Bacillus safensis (W10) and Ochrobactrum pseudogrignonense (IP8) produced antifungal and antimicrobial compounds in culture. Seed priming with these two bacteria significantly increase growth, modulate antioxidative signaling and induce resistance and eventually reduce disease incidence in wheat plants at optimum as well as elevated temperature which was further confirmed by indirect immunofluorescence assay using polyclonal antibody raised against Bipolaris sorokiniana. Application of the PGPR led to enhancement in activities of plant defense enzymes- phenylalanine ammonia lyase, peroxidase, chitinase and β-1,3 glucanase in infected leaves. Immunolocalization of chitinase and β-1,3 glucanase in PGPR primed and pathogen inoculated leaf tissue was further confirmed by transmission electron microscopy using PAb of chitinase, β-1,3 glucanase and gold labelled conjugates. Activity of ascorbate-glutathione redox cycle related enzymes such as ascorbate peroxidase, superoxide dismutase and glutathione reductase along with antioxidants such as carotenoids, glutathione and ascorbate and osmolytes like proline and glycine betain accumulation were also increased during disease development in PGPR primed plant in comparison to unprimed plants at high temperature. Real-time PCR analysis revealed enhanced expression of defense genes- chalcone synthase and phenyl alanineammonia lyase. Over expression of heat shock proteins like HSP 70, small HSP 26.3 and heat shock factor HsfA3 in PGPR primed plants effectively protect plants against spot blotch infection at elevated temperature as compared with control plants. Our results revealed dynamic biochemical cross talk between elevated temperature and spot blotch disease development and furthermore highlight PGPR mediated array of antioxidative and molecular alterations responsible for induction of resistance against spot blotch disease at elevated temperature which seems to be associated with up-regulation of defense genes, heat shock proteins and heat shock factors, less ROS production, membrane damage, increased expression of redox enzymes and accumulation of osmolytes and antioxidants.

Keywords: antioxidative enzymes, defense enzymes, elevated temperature, heat shock proteins, PGPR, Real-Time PCR, spot blotch, wheat

Procedia PDF Downloads 144

Authors: Kuladip Jana

Abstract:

Benzo(a)pyrene [B(a)P] is an environmental toxicant present mostly in cigarette smoke and car exhaust, is an aryl hydrocarbon receptor (AhR) ligand that exerts its toxic effects on both male and female reproductive systems. In this study, the effect of B(a)P at different doses (0.1, 0.25, 0.5, 1 and 5 mg /kg body weight) was studied on male reproductive system of rat. A significant decrease in cauda epididymal sperm count and motility along with the presence of sperm head abnormalities and altered epididymal and testicular histology were documented following B(a)P treatment. B(a)P treatment resulted apoptotic sperm cells as observed by TUNEL and Annexin V-PI assay with increased ROS, altered sperm mitochondrial membrane potential (ΔΨm) with a simultaneous decrease in the activity of antioxidant enzymes and GSH status. TUNEL positive apoptotic cells also observed in testis as well as isolated germ and Leydig cells following B(a)P exposure. Western Blot analysis revealed the activation of p38MAPK, cytosolic translocation of cytochrome-c, up-regulation of Bax and inducible nitric oxide synthase (iNOS) with cleavage of PARP and down-regulation of BCl2 in testis upon B(a)P treatment. The protein and mRNA levels of testicular key steroidogenesis regulatory proteins like StAR, cytochrome P450 IIA1 (CYPIIA1), 3β HSD, 17β HSD showed a significant decrease in a dose dependent manner while an increase in the expression of cytochrome P450 1A1 (CYP1A1), Aryl hydrocarbon Receptor (AhR), active caspase- 9 and caspase- 3 following B(a)P exposure. We conclude that exposure of benzo(a)pyrene caused testicular gamatogenic and steroidogenic disorders by induction of oxidative stress, inhibition of StAR and other steroidogenic enzymes along with activation of p38MAPK and initiated caspase-3 mediated germ and Leydig cell apoptosis.The possible protective role of naturally occurring phytochemicals against B(a)P induced testicular toxicity needs immediate consideration. Curcumin and resveratrol separately were found to protect against B(a)P induced germ cell apoptosis, and their combinatorial effect was more significant. Our present study in isolated testicular germ cell population from adult male Wistar rats, highlighted their synergistic protective effect against B(a)P induced germ cell apoptosis. Curcumin-resveratrol co-treatment decreased the expression of pro-apoptotic proteins like cleaved caspase 3,8,9, cleaved PARP, Apaf1, FasL, tBid. Curcumin-resveratrol co-treatment decreased Bax/Bcl2 ratio, mitochondria to cytosolic translocation of cytochrome c and activated the survival protein Akt. Curcumin-resveratrol decreased the expression of p53 dependent apoptotic genes like Fas, FasL, Bax, Bcl2, Apaf1.Curcumin-resveratrol co-treatment thus prevented B(a)P induced germ cell apoptosis. B(a)P induced testicular ROS generation and oxidative stress were significantly ameliorated with curcumin and resveratrol. Curcumin-resveratrol co-treatment prevented B(a)P induced nuclear translocation of AhR and CYP1A1 production. The combinatorial treatment significantly inhibited B(a)P induced ERK 1/2, p38 MAPK and JNK 1/2 activation. B(a)P treatment increased the expression of p53 and its phosphorylation (p53 ser 15). Curcumin-resveratrol co-treatment significantly decreased p53 level and its phosphorylation (p53 ser 15). The study concludes that curcumin-resveratrol synergistically modulated MAPKs and p53, prevented oxidative stress, regulated the expression of pro and anti-apoptotic proteins as well as the proteins involved in B(a)P metabolism thus protected germ cells from B(a)P induced apoptosis.

Keywords: benzo(a)pyrene, germ cell, apoptosis, oxidative stress, resveratrol, curcumin

Procedia PDF Downloads 233

Authors: Pallab Datta, Jyotirmoy Chatterjee, Santanu Dhara

Abstract:

Over the past few years, phosphorous containing polymers have received widespread attention for applications such as high performance optical fibers, flame retardant materials, drug delivery and tissue engineering. Being pentavalent, phosphorous can exist in different chemical environments in these polymers which increase their versatility. In human biochemistry, phosphorous based compounds exert their functions both in soluble and insoluble form occurring as inorganic or as organophosphorous compounds. Specifically in case of biomacromolecules, phosphates are critical for functions of DNA, ATP, phosphoproteins, phospholipids, phosphoglycans and several coenzymes. Inspired by the role of phosphorous in functional biomacromolecules, design and synthesis of biomimetic materials are thus carried out by several authors to study macromolecular function or as substitutes in clinical tissue regeneration conditions. In addition, many regulatory signals of the body are controlled by phoshphorylation of key proteins present either in form of growth factors or matrix-bound scaffold proteins. This inspires works on synthesis of phospho-peptidomimetic amino acids for understanding key signaling pathways and this is extended to obtain molecules with potentially useful biological properties. Apart from above applications, phosphate groups bound to polymer backbones have also been demonstrated to improve function of osteoblast cells and augment performance of bone grafts. Despite the advantages of phosphate grafting, however, there is limited understanding on effect of degree of phosphorylation on macromolecular physicochemical and/or biological properties. Such investigations are necessary to effectively translate knowledge of macromolecular biochemistry into relevant clinical products since they directly influence processability of these polymers into suitable scaffold structures and control subsequent biological response. Amongst various techniques for fabrication of biomimetic scaffolds, nanofibrous scaffolds fabricated by electrospinning technique offer some special advantages in resembling the attributes of natural extracellular matrix. Understanding changes in physico-chemical properties of polymers as function of phosphorylation is therefore going to be crucial in development of nanofiber scaffolds based on phosphorylated polymers. The aim of the present work is to investigate the effect of phosphorous grafting on the electrospinning behavior of polymers with aim to obtain biomaterials for bone regeneration applications. For this purpose, phosphorylated derivatives of two polymers of widely different electrospinning behaviors were selected as starting materials. Poly(vinyl alcohol) is a conveniently electrospinnable polymer at different conditions and concentrations. On the other hand, electrospinning of chitosan backbone based polymers have been viewed as a critical challenge. The phosphorylated derivatives of these polymers were synthesized, characterized and electrospinning behavior of various solutions containing these derivatives was compared with electrospinning of pure poly (vinyl alcohol). In PVA, phosphorylation adversely impacted electrospinnability while in NMPC, higher phosphate content widened concentration range for nanofiber formation. Culture of MG-63 cells on electrospun nanofibers, revealed that degree of phosphate modification of a polymer significantly improves cell adhesion or osteoblast function of cultured cells. It is concluded that improvement of cell response parameters of nanofiber scaffolds can be attained as a function of controlled degree of phosphate grafting in polymeric biomaterials with implications for bone tissue engineering applications.

Keywords: bone regeneration, chitosan, electrospinning, phosphorylation

Procedia PDF Downloads 199

Authors: Kuladip Jana, Bhaswati Banerjee, Parimal C. Sen

Abstract:

Benzo(a)pyrene [B(a)P] is an environmental toxicant present mostly in cigarette smoke and car exhaust, is an aryl hydrocarbon receptor (AhR) ligand that exerts its toxic effects on both male and female reproductive systems along with carcinogenesis in skin, prostate, ovary, lung and mammary glands. Our study was focused on elucidating the molecular mechanism of B(a)P induced male reproductive toxicity and its prevention with phytochemical like resveratrol. In this study, the effect of B(a)P at different doses (0.1, 0.25, 0.5, 1 and 5 mg /kg body weight) was studied on male reproductive system of Wistar rat. A significant decrease in cauda epididymal sperm count and motility along with the presence of sperm head abnormalities and altered epididymal and testicular histology were documented following B(a)P treatment. B(a)P treatment resulted apoptotic sperm cells as observed by TUNEL and Annexin V-PI assay with increased Reactive Oxygen Species (ROS), altered sperm mitochondrial membrane potential (ΔΨm) with a simultaneous decrease in the activity of antioxidant enzymes and GSH status. TUNEL positive apoptotic cells also observed in testis as well as isolated germ and Leydig cells following B(a)P exposure. Western Blot analysis revealed the activation of p38 mitogen activated protein kinase (p38MAPK), cytosolic translocation of cytochrome-c, upregulation of Bax and inducible nitric oxide synthase (iNOS) with cleavage of poly ADP ribose polymerase (PARP) and down regulation of BCl2 in testis upon B(a)P treatment. The protein and mRNA levels of testicular key steroidogenesis regulatory proteins like steroidogenic acute regulatory protein (StAR), cytochrome P450 IIA1 (CYPIIA1), 3β hydroxy steroid dehydrogenase (3β HSD), 17β hydroxy steroid dehydrogenase (17β HSD) showed a significant decrease in a dose dependent manner while an increase in the expression of cytochrome P450 1A1 (CYP1A1), Aryl hydrocarbon Receptor (AhR), active caspase- 9 and caspase- 3 following B(a)P exposure. We conclude that exposure of benzo(a)pyrene caused testicular gamatogenic and steroidogenic disorders by induction of oxidative stress, inhibition of StAR and other steroidogenic enzymes along with activation of p38MAPK and initiated caspase-3 mediated germ and Leydig cell apoptosis. Next we investigated the role of resveratrol on B(a)P induced male reproductive toxicity. Our study highlighted that resveratrol co-treatment with B(a)P maintained testicular redox potential, increased serum testosterone level and prevented steroidogenic dysfunction with enhanced expression of major testicular steroidogenic proteins (CYPIIA1, StAR, 3β HSD,17β HSD) relative to treatment with B(a)P only. Resveratrol suppressed B(a)P-induced testicular activation of p38 MAPK, ATF2, iNOS and ROS production; cytosolic translocation of Cytochome c and Caspase 3 activation thereby prevented oxidative stress of testis and inhibited apoptosis. Resveratrol co-treatment also decreased B(a)P-induced AhR protein level, its nuclear translocation and subsequent CYP1A1 promoter activation, thereby decreased protein and mRNA levels of testicular cytochrome P4501A1 (CYP1A1) and prevented BPDE-DNA adduct formation. Our findings cumulatively suggest that resveratrol prevents activation of B(a)P by modulating the transcriptional regulation of CYP1A1 and acting as an antioxidant thus prevents B(a)P-induced oxidative stress and testicular apoptosis.

Keywords: benzo(a)pyrene, resveratrol, testis, apoptosis, cytochrome P450 1A1 (CYP1A1), aryl hydrocarbon receptor (AhR), p38 MAPK/ATF2/iNOS

Procedia PDF Downloads 200

Authors: Z. S. Haidar

Abstract:

Millions of teeth are removed annually, and dental extraction is one of the most commonly performed surgical procedures globally. Whether due to caries, periodontal disease, or trauma, exodontia and the ensuing wound healing and bone remodeling processes of the resultant socket (hole in the jaw bone) usually result in serious deformities of the residual alveolar osseous ridge and surrounding soft tissues (reduced height/width). Such voluminous changes render the placement of a proper conventional bridge, denture, or even an implant-supported prosthesis extremely challenging. Further, most extractions continue to be performed with no regard for preventing the onset of alveolar osteitis (also known as dry socket, a painful and difficult-to-treat/-manage condition post-exodontia). Hence, such serious resorptive morphological changes often result in significant facial deformities and a negative impact on the overall Quality of Life (QoL) of patients (and oral health-related QoL); alarming, particularly for the geriatric with compromised healing and in light of the thriving longevity statistics. Despite advances in tissue/wound grafting, serious limitations continue to exist, including efficacy and clinical outcome predictability, cost, treatment time, expertise, and risk of immune reactions. For cases of dry socket, specifically, the commercially available and often-prescribed home remedies are highly-lacking. Indeed, most are not recommended for use anymore. Alveogyl is a fine example. Hence, there is a great market demand and need for alternative solutions. Herein, SockGEL/PLUG (patent pending), an innovative, all-natural, drug-free, and injectable thermo-responsive hydrogel, was designed, formulated, characterized, and evaluated as an osteogenic, angiogenic, anti-microbial, and pain-soothing suture-free intra-alveolar dressing, safe and efficacious for use in fresh extraction sockets, immediately post-exodontia. It is composed of FDA-approved, biocompatible and biodegradable polymers, self-assembled electro-statically to formulate a scaffolding matrix to (1) prevent the on-set of alveolar osteitis via securing the fibrin-clot in situ and protecting/sealing the socket from contamination/infection; and (2) endogenously promote/accelerate wound healing and bone remodeling to preserve the volume of the alveolus. The intrinsic properties of the SockGEL/PLUG hydrogel were evaluated physical-chemical-mechanically for safety (cell viability), viscosity, rheology, bio-distribution, and essentially, capacity to induce wound healing and osteogenesis (small defect, in vivo) without any signaling cues from exogenous cells, growth factors or drugs. The proposed animal model of cranial critical-sized and non-vascularized bone defects shall provide new and critical insights into the role and mechanism of the employed natural bio-polymer blend and gel product in endogenous reparative regeneration of soft tissues and bone morphogenesis. Alongside, the fine-tuning of our modified formulation method will further tackle appropriateness, reproducibility, scalability, ease, and speed in producing stable, biodegradable, and sterilizable thermo-sensitive matrices (3-dimensional interpenetrating yet porous polymeric network) suitable for the intra-socket application. Findings are anticipated to provide sufficient evidence to translate into pilot clinical trials and validate the innovation before engaging the market for feasibility, acceptance, and cost-effectiveness studies.

Keywords: hydrogel, nanotechnology, bioengineering, bone regeneration, nanogel, drug delivery

Procedia PDF Downloads 87

Authors: Claire Harrison, Raajit K. Rampal, Vikas Gupta, Srdan Verstovsek, Moshe Talpaz, Jean-Jacques Kiladjian, Ruben Mesa, Andrew Kuykendall, Alessandro Vannucchi, Francesca Palandri, Sebastian Grosicki, Timothy Devos, Eric Jourdan, Marielle J. Wondergem, Haifa Kathrin Al-Ali, Veronika Buxhofer-Ausch, Alberto Alvarez-Larrán, Sanjay Akhani, Rafael Muñoz-Carerras, Yury Sheykin, Gozde Colak, Morgan Harris, John Mascarenhas

Abstract:

Myelofibrosis (MF) is characterized by bone marrow fibrosis, anemia, splenomegaly and constitutional symptoms. Progressive bone marrow fibrosis results from aberrant megakaryopoeisis and expression of proinflammatory cytokines, both of which are heavily influenced by bromodomain and extraterminal domain (BET)-mediated gene regulation and lead to myeloproliferation and cytopenias. Pelabresib (CPI-0610) is an oral small-molecule investigational inhibitor of BET protein bromodomains currently being developed for the treatment of patients with MF. It is designed to downregulate BET target genes and modify nuclear factor kappa B (NF-κB) signaling. MANIFEST-2 was initiated based on data from Arm 3 of the ongoing Phase 2 MANIFEST study (NCT02158858), which is evaluating the combination of pelabresib and ruxolitinib in Janus kinase inhibitor (JAKi) treatment-naïve patients with MF. Primary endpoint analyses showed splenic and symptom responses in 68% and 56% of 84 enrolled patients, respectively. MANIFEST-2 (NCT04603495) is a global, Phase 3, randomized, double-blind, active-control study of pelabresib and ruxolitinib versus placebo and ruxolitinib in JAKi treatment-naïve patients with primary MF, post-polycythemia vera MF or post-essential thrombocythemia MF. The aim of this study is to evaluate the efficacy and safety of pelabresib in combination with ruxolitinib. Here we report updates from a recent protocol amendment. The MANIFEST-2 study schema is shown in Figure 1. Key eligibility criteria include a Dynamic International Prognostic Scoring System (DIPSS) score of Intermediate-1 or higher, platelet count ≥100 × 10^9/L, spleen volume ≥450 cc by computerized tomography or magnetic resonance imaging, ≥2 symptoms with an average score ≥3 or a Total Symptom Score (TSS) of ≥10 using the Myelofibrosis Symptom Assessment Form v4.0, peripheral blast count <5% and Eastern Cooperative Oncology Group performance status ≤2. Patient randomization will be stratified by DIPSS risk category (Intermediate-1 vs Intermediate-2 vs High), platelet count (>200 × 10^9/L vs 100–200 × 10^9/L) and spleen volume (≥1800 cm^3 vs <1800 cm^3). Double-blind treatment (pelabresib or matching placebo) will be administered once daily for 14 consecutive days, followed by a 7 day break, which is considered one cycle of treatment. Ruxolitinib will be administered twice daily for all 21 days of the cycle. The primary endpoint is SVR35 response (≥35% reduction in spleen volume from baseline) at Week 24, and the key secondary endpoint is TSS50 response (≥50% reduction in TSS from baseline) at Week 24. Other secondary endpoints include safety, pharmacokinetics, changes in bone marrow fibrosis, duration of SVR35 response, duration of TSS50 response, progression-free survival, overall survival, conversion from transfusion dependence to independence and rate of red blood cell transfusion for the first 24 weeks. Study recruitment is ongoing; 400 patients (200 per arm) from North America, Europe, Asia and Australia will be enrolled. The study opened for enrollment in November 2020. MANIFEST-2 was initiated based on data from the ongoing Phase 2 MANIFEST study with the aim of assessing the efficacy and safety of pelabresib and ruxolitinib in JAKi treatment-naïve patients with MF. MANIFEST-2 is currently open for enrollment.

Keywords: CPI-0610, JAKi treatment-naïve, MANIFEST-2, myelofibrosis, pelabresib

Procedia PDF Downloads 167

Authors: Ziyad S. Haidar

Abstract:

Millions of teeth are removed annually, and dental extraction is one of the most commonly performed surgical procedures globally. Whether due to caries, periodontal disease or trauma, exodontia and the ensuing wound healing and bone remodeling processes of the resultant socket (hole in the jaw bone) usually result in serious deformities of the residual alveolar osseous ridge and surrounding soft tissues (reduced height/width). Such voluminous changes render the placement of a proper conventional bridge, denture or even an implant-supported prosthesis extremely challenging. Further, most extractions continue to be performed with no regard for preventing the onset of alveolar osteitis (also known as dry socket, a painful and difficult-to-treat/-manage condition post-exodontia). Hence, such serious resorptive morphological changes often result in significant facial deformities and a negative impact on the overall Quality of Life (QoL) of patients (and oral health-related QoL), alarming, particularly for the geriatric with compromised healing and in light of the thriving longevity statistics. Opportunity: Despite advances in tissue/wound grafting, serious limitations continue to exist, including efficacy and clinical outcome predictability, cost, treatment time, expertise and risk of immune reactions. For cases of dry sockets, specifically, the commercially-available and often-prescribed home remedies are highly lacking. Indeed, most are not recommended for use anymore. Alveogyl is a fine example. Hence, there is a great market demand and need for alternative solutions. Solution: Herein, SockGEL/PLUG (patent pending), an all-natural, drug-free and injectable stimuli-responsive hydrogel, was designed, formulated, characterized and evaluated as an osteogenic, angiogenic, anti-microbial and pain-soothing suture-free intra-alveolar dressing, safe and efficacious for use in several oro-dental and cranio-maxillo-facial interventional applications; for example: in fresh dental extraction sockets, immediately post-exodontia. It is composed of FDA-approved, biocompatible and biodegradable polymers, self-assembled electro-statically to formulate a scaffolding matrix to (a) prevent the onset of alveolar osteitis via securing the fibrin-clot in situ and protecting/sealing the socket from contamination/infection; and (b) endogenously promote/accelerate wound healing and bone remodeling to preserve the volume of the alveolus. Findings: The intrinsic properties of the SockGEL/PLUG hydrogel were evaluated physico-chemico-mechanically for safety (cell viability), viscosity, rheology, bio-distribution and essentially, capacity to induce wound healing and osteogenesis (small defect, in vivo) without any signaling cues from exogenous cells, growth factors or drugs. The performed animal model of cranial critical-sized and non-vascularized bone defects shall provide vitally critical insights into the role and mechanism of the employed natural bio-polymer blend and gel product in endogenous reparative regeneration of soft tissues and bone morphogenesis. Alongside, the fine-tuning of our modified formulation method will further tackle appropriateness, reproducibility, scalability, ease and speed in producing stable, biodegradable and sterilizable stimuli (thermo-sensitive and photo-responsive) matrices (3-dimensional interpenetrating yet porous polymeric network) suitable for an intra-socket application, and beyond. Conclusions and Perspective: Findings are anticipated to provide sufficient evidence to translate into pilot clinical trials and validate the bionanomaterial before engaging the market for feasibility, acceptance and cost-effectiveness studies. The SockGEL/PLUG platform is patent pending: SockGEL is a bio-inspired drug-free hydrogel; SockPLUG is a drug-loaded hydrogel designed for complex indications.

Keywords: hydrogel, injectable, dentistry, craniomaxillofacial complex, bioinspired, nanobiotechnology, biopolymer, sol-gel, stimuli-responsive, matrix, tissue engineering, regenerative medicine

Procedia PDF Downloads 46