Biology subjects

Colazo, J. M.

Publications and source records attributed to Colazo, J. M..

5 recordsLinked to original sources

bioRxiv2024

A programmable arthritis-specific receptor for guided articular cartilage regenerative medicine

ObjectiveInvestigational cell therapies have been developed as disease-modifying agents for the treatment of osteoarthritis (OA), including those that inducibly respond to inflammatory factors driving OA progression. However, dysregulated inflammatory cascades do not specifically signify the presence of OA. Here, we deploy a synthetic receptor platform that regulates cell behaviors in an arthritis-specific fashion to confine transgene expression to sites characterized by cartilage degeneration. MethodsAn scFv specific for type II collagen (CII) was used to produce a synthetic Notch (synNotch) receptor that enables "CII-synNotch" mesenchymal stromal cells (MSCs) to recognize CII fibers exposed in damaged cartilage. Engineered cell activation by both CII-treated culture surfaces and on primary tissue samples was measured via inducible reporter transgene expression. TGF{beta}3-expressing cells were assessed for cartilage anabolic gene expression via qRT-PCR. In a co-culture with CII-synNotch MSCs engineered to express IL-1Ra, ATDC5 chondrocytes were stimulated with IL-1, and inflammatory responses of ATDC5s were profiled via qRT-PCR and an NF-kB reporter assay. ResultsCII-synNotch MSCs are highly responsive to CII, displaying activation ranges over 40-fold in response to physiologic CII inputs. CII-synNotch cells exhibit the capacity to distinguish between healthy and damaged cartilage tissue and constrain transgene expression to regions of exposed CII fibers. Receptor-regulated TGF{beta}3 expression resulted in upregulation of Acan and Col2a1 in MSCs, and inducible IL-1Ra expression by engineered CII-synNotch MSCs reduced pro-inflammatory gene expression in chondrocytes. ConclusionThis work demonstrates proof-of-concept that the synNotch platform guides MSCs for spatially regulated, disease-dependent delivery of OA-relevant biologic drugs.

bioengineering↗

bioRxiv2023

Albumin-binding RNAi Conjugate for Carrier Free Treatment of Arthritis

Osteoarthritis (OA) and rheumatoid arthritis (RA) are joint diseases that are associated with pain and lost quality of life. No disease modifying OA drugs are currently available. RA treatments are better established but are not always effective and can cause immune suppression. Here, an MMP13-selective siRNA conjugate was developed that, when delivered intravenously, docks onto endogenous albumin and promotes preferential accumulation in articular cartilage and synovia of OA and RA joints. MMP13 expression was diminished upon intravenous delivery of MMP13 siRNA conjugates, consequently decreasing multiple histological and molecular markers of disease severity, while also reducing clinical manifestations such as swelling (RA) and joint pressure sensitivity (RA and OA). Importantly, MMP13 silencing provided more comprehensive OA treatment efficacy than standard of care (steroids) or experimental MMP inhibitors. These data demonstrate the utility of albumin hitchhiking for drug delivery to arthritic joints, and establish the therapeutic utility of systemically delivered anti-MMP13 siRNA conjugates in OA and RA. Editorial summaryLipophilic siRNA conjugates optimized for albumin binding and "hitchhiking" can be leveraged to achieve preferential delivery to and gene silencing activity within arthritic joints. Chemical stabilization of the lipophilic siRNA enables intravenous siRNA delivery without lipid or polymer encapsulation. Using siRNA sequences targeting MMP13, a key driver of arthritis-related inflammation, albumin hitchhiking siRNA diminished MMP13, inflammation, and manifestations of osteoarthritis and rheumatoid arthritis at molecular, histological, and clinical levels, consistently outperforming clinical standards of care and small molecule MMP antagonists.

bioengineering↗

bioRxiv2021

ROS-Degradable Polythioketal Urethane Foam Dressings to Promote Porcine Skin Wound Repair

Impaired skin healing and progression into chronic wounds is a prevalent and growing medical problem. Porous, resorbable biomaterials can be used as temporary substrates placed into skin defects to support cell infiltration, neo-tissue formation, and remodeling of nonhealing wounds. Naturally-derived biomaterials have promising healing benefits, but their low mechanical properties and exuberant costs limit their performance and use. Synthetic materials can be affordably manufactured and tuned across a broader range of physiochemical properties, but opportunities remain for tailoring them for ideal host immune and regenerative responses. Polyesters are the most clinically-tested class of synthetic biomaterials, but their hydrolysis releases acidic degradation products that can cause autocatalytic degradation processes that are poorly controlled and are not tied to cellular or other biologic activities. Here, we systemically explored a series of ROS-degradable polythioketal (PTK) urethane (UR) foams with varied hydrophilicity as an alternative class of synthetic biomaterials for wound healing. It was found that the most hydrophilic PTK- UR variant, which had 7 ethylene glycol (EG7) repeats flanking each side of each thioketal bond, had the highest ROS reactivity of the PTK-URs tested. In an in vivo porcine excisional skin wound healing model, hydrophilic EG7 PTK-UR foams more effectively promoted tissue integration, ECM deposition, and re- epithelialization of full-thickness skin wound compared to more hydrophobic PTK-UR variants. Resolution of type 1 inflammation and lower foreign body response to scaffold remnants was also observed for EG7 versus more hydrophobic PTK-UR scaffolds. Finally, porcine wound healing studies showed that EG7 PTK-UR foams had similar wound healing response to a collagen-based clinical gold standard product, Integra Bilayer Wound Matrix (BWM), while outperforming polyester UR foam-based NovoSorb Biodegradable Temporizing Matrix (BTM) with respect to increased ECM production, vascularization, and biomaterial-associated immune phenotype. In sum, PTK-UR foams warrant further development toward a new class of synthetic biomaterial foams for skin wound healing applications.

bioengineering↗

bioRxiv2021

Galectin-anchored indoleamine 2,3-dioxygenase suppresses local inflammation

Summary paragraphChronic inflammation underlies the onset, progression and associated pain of numerous diseases.(1) Current anti-inflammatory treatments administered systemically are associated with moderate-to-severe side effects, while locally administered drugs have short-lived efficacy, and neither approach successfully modifies the underlying causality of disease.(2) We report a new way to locally modulate inflammation by fusing the enzyme indoleamine 2,3-dioxygenase 1 (IDO) to galectin-3 (Gal3). A general regulator of inflammation(3), IDO is immunosuppressive(4), catabolizing the essential amino acid tryptophan into kynurenine.(5) Recently we demonstrated that extracellular exogenous IDO regulates innate immune cell function(6), and envisioned delivering IDO into specific tissues would provide control of inflammation. However, proteins problematically diffuse away from local injection sites. Addressing this, we recently established that fusion to Gal3 anchors enzymes to tissues(7) via binding to extracellular glycans. Fusion protein IDO-Gal3 was retained in injected tissues and joints for up to a week or more, where it suppressed local inflammation in rodent models of endotoxin-induced inflammation, psoriasis, periodontal disease and osteoarthritis. Amelioration of local inflammation, disease progression and inflammatory pain were concomitant with homeostatic preservation of tissues without global immune suppression. Thus, IDO-Gal3 presents a new concept of anchoring immunomodulatory enzymes for robust control of focal inflammation in multiple disease settings.

bioengineering↗

bioRxiv2020

Matrix-targeted Nanoparticles for MMP13 RNA Interference Blocks Post-Traumatic Osteoarthritis.

Osteoarthritis (OA) is a debilitating and prevalent chronic disease, but there are no approved disease modifying OA drugs (DMOADs), only pharmaceuticals for pain management. OA progression, particularly for post-traumatic osteoarthritis (PTOA), is associated with inflammation and enzymatic degradation of the extracellular matrix. In particular, Matrix Metalloproteinase 13 (MMP13) breaks down collagen type 2 (CII), a key structural component of cartilage extracellular matrix, and consequently, matrix degradation fragments perpetuate inflammation and a degenerative cycle that leads to progressive joint pathology. Here, we tested targeted delivery of endosome-escaping, MMP13 RNA interference (RNAi) nanoparticles (NPs) as a DMOAD. The new targeting approach pursued here deviates from the convention of targeting specific cell types (e.g., through cell surface receptors) and instead leverages a monoclonal antibody (mAbCII) that targets extracellular CII that becomes uniquely accessible at early OA focal defects. Targeted mAbCII-siNPs create an in situ NP depot for retention and potent activity within OA joints. The mAbCII-siNPs loaded with MMP13 siRNA (mAbCII-siNP/siMMP13) potently suppressed MMP13 expression (95% silencing) in TNF-stimulated chondrocytes in vitro, and the targeted mAbCII-siNPs had higher binding to trypsin-damaged porcine cartilage than untargeted control NPs. In an acute mechanical injury mouse model of PTOA, mAbCII-siNP/siMMP13 achieved 80% reduction in MMP13 expression (p = 0.00231), whereas a non-targeted control achieved only 55% silencing. In a more severe, PTOA model, weekly mAbCII-siNP/siMMP13 long-term treatment provided significant protection of cartilage integrity (0.45+/- .3 vs 1.6+/-.5 on the OARSI scale; p=0.0166), and overall joint structure (1.3+/-.6 vs 2.8+/-.2 on the Degenerative Joint Disease scale; p<0.05). Intra-articular mAbCII-siNPs better protected articular cartilage (OARSI score) relative to either single or weekly treatment with the clinical gold stand steroid treatment methylprednisolone. Finally, multiplexed gene expression analysis of 254 inflammation-related genes showed that MMP13 inhibition suppressed clusters of genes associated with tissue restructuring, angiogenesis (associated with synovial inflammation and thickening), innate immune response, and proteolysis. This work establishes the new concept of targeting unique local extracellular matrix signatures to sustain retention and increase delivery efficacy of biologics with intracellular activity and also validates the promise of MMP13 RNAi as a DMOAD in a clinically-relevant therapeutic context. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=148 SRC="FIGDIR/small/925321v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@1803795org.highwire.dtl.DTLVardef@c1254dorg.highwire.dtl.DTLVardef@78233eorg.highwire.dtl.DTLVardef@151d96f_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOAbstract Figure:C_FLOATNO PTOA targeted delivery of MMP13 siRNA to block disease progressionThe top left schematic illustrates the progression (left to right) from healthy knee joint, through inflammation induction following traumatic injury, to cartilage loss and degenerative joint disease (including synovial response). Degradation of cartilage enhances inflammation, inducing a degenerative cycle (middle right). The bottom of the graphic illustrates the concept of the matrix targeted nanocarriers for enhanced retention and activity of MMP13 siRNA at sites of cartilage injury. C_FIG

bioengineering↗