Publications

Triple-Negative Breast Cancer Aptamer-Targeting Porous Silicon Nanocarrier

Ankit Malhotra, Pouya Dehghankelishadi, Ishdeep Kaur, Morgan Marshall, David Rudd, Marcin Wojnilowicz, Cameron J Nowell, Alex J Fulcher, Lars Esser, Wing Yin Tong, Anna Cifuentes, Kylie M Wagstaff, Nicolas H Voelcker

ACS Applied Materials & Interfaces · 2026

Common treatment approaches for triple-negative breast cancer (TNBC) are associated with severe side effects due to the unfavorable biodistribution profile of potent chemotherapeutics. Here, we explored the potential of TNBC-targeting aptamer-decorated porous silicon nanoparticles (pSiNPs) as targeted nanocarriers for TNBC. A "salt-aging" strategy was employed to fabricate a TNBC-targeting aptamer functionalized pSiNP that was highly colloidally stable. Doxorubicin (Dox) was efficiently loaded into nanoparticles (179 ± 5 μg/mg of pSiNP) and experienced pH-dependent release kinetics. Further experiments highlighted that clathrin-mediated endocytosis was the primary route that aptamer-pSiNP conjugates take to enter the endolysosomal compartment of the MCF10Ca1h TNBC cells. A time-interval colocalization study shows the accumulation of an aptamer-decorated pSiNP conjugate in the lysosomes of TNBC cells, unlike for antibody-decorated pSiNPs, leading to particle-induced lysosomal swelling and membrane destabilization. Dox-loaded aptamer-pSiNPs efficiently reduced the viability of the TNBC cells (11.8 ± 1.5%) compared to nontargeted nanoparticles (58.2 ± 8.8%) while the developed system showed a low level of toxicity in healthy cells, both in vitro and in vivo. These findings have laid the foundation for further investigating the potential of aptamer-pSiNP conjugates as a targeted treatment strategy in preclinical TNBC models.

Endosomal escape and cytosolic delivery of cell-penetrating peptide conjugates

Kathryn J. Fincham, Cameron J. Nowell, Macgregor A. Matthews, David K. Chalmers, Philip E. Thompson, Sandra E. Nicholson, Raymond S. Norton

Bioorganic & Medicinal Chemistry · 2026

Cell-penetrating peptides (CPPs) are sequences designed to enhance the cytosolic delivery of a range of cargoes. We are interested in delivering peptides containing the sequence DINNN, which are high-affinity inhibitors of the interaction between the SPRY domain-containing SOCS (suppressor of cytokine signaling) box (SPSB) proteins 1, 2 and 4 and inducible nitric oxide synthase (iNOS), and have potential as host-directed antibiotics. In order to assess the efficacy of CPPs in delivering these peptides to the cytosol of target cells, we have utilised the split luciferase endosomal escape quantitation (SLEEQ) assay, which measures the endosomal escape and cytosolic delivery of CPP conjugates. After applying correction factors to account for luminescence differences between peptides, the SLEEQ assay was used to evaluate CPP-peptide conjugates for their ability to enter a macrophage cell line. The CPPs TAT, (RW)4 and CPP9 all successfully enhanced cytosolic delivery of DINNN-containing peptides, but did so mainly by enhancing uptake into endosomes rather than by enhancing endosomal escape. Inclusion of the fluorophore sCy5 in the peptide conjugates further enhanced cytosolic uptake in general, although the impact varied significantly depending on the CPP used. Confocal imaging of the sCy5-containing peptide conjugates was consistent with the results of the SLEEQ assay, emphasising the risk of conflating CPP efficacy and dye effects when using fluorophore-based methods to study cell uptake. Our results for total cellular association and endosomal escape argue strongly for a mechanism of CPP uptake wherein interaction with the membrane and incorporation into endosomes is the key factor driving cytosolic delivery, rather than the efficiency of endosomal escape. While CPPs did enhance the cytosolic uptake of a DINNN-containing peptide, endosomal escape was found to be very inefficient, with none of the CPPs investigated here enhancing endosomal escape of the cargo.

Transmembrane orientation and extracellular juxtamembrane residues of GP130 are critical for signal transduction

Farhad Dehkhoda, Yash Chhabra, Fiona Whelan, Cameron J Nowell, Matthew J Morris, Sebastian G B Furness, Andrew J Brooks

Journal of Immunology · 2026

Glycoprotein 130 (GP130) is an archetypal class I cytokine receptor that serves as a shared signalling subunit for a group of cytokines known as interleukin 6 (IL-6)-type cytokines. GP130 does not have an intrinsic kinase activity and relies on Janus kinases (JAK) 1, 2, or Tyrosine kinase 2 (TYK2) for phosphorylation of tyrosine residues on its cytoplasmic domain for signal transduction. In this study we investigated the molecular mechanism of GP130 activation. We utilized GP130 constructs that were held in varying states of activation by replacing the entire extracellular region of the receptor with a leucine zipper dimerisation domain and inserted alanine residues into the extracellular juxtamembrane linker sequence to induce rotation of the transmembrane domains. These chimeras differentially activated the associated JAK2 molecules which also correlated with receptor tyrosine phosphorylation status. Förster resonance energy transfer (FRET) microscopy in live cells was used to show that GP130 cytoplasmic domains were in close proximity in active chimeras compared to the inactive receptors, presumably to facilitate JAK protein activation. Coarse-grained molecular dynamics simulations provided a structural rationale for this register-dependent mechanism, showing that active chimeras preferentially maintain compact transmembrane packing and reduced membrane-proximal cytosolic spacing, whereas signalling-deficient constructs sample more heterogeneous transmembrane geometries and increased cytosolic separation. Finally, mutational analysis identified the negatively charged juxtamembrane residues E617 and E619 as critical for signal transduction; charge reversal at these positions in the full-length receptor generated a dominant-negative GP130 variant devoid of signalling capacity.

Global impact through train-the-trainer (TtT) courses

Mar García-Ferrés, Amy L Bottomley, Louise Cole, Marcela Díaz, Neftali Flores-Rodriguez, Gleb Grebnev, Adan Guerrero, Caron A Jacobs, Laurence Lejeune, Xiaoxiao Ma, Leonel Malacrida, Paul J McMillan, Alison J North, Cameron J Nowell, Naomi Okugbeni, María V Repetto, Alejandra M Ross Beraldi, Andres Hugo Rossi, Lía I Pietrasanta, Rajan Singh, Claire M Brown

Journal of Microscopy · 2026

Global BioImaging (GBI) and its regional and national partners, such as Canada BioImaging (CBI), BioImaging North America (BINA), Latin America BioImaging (LABI), the African BioImaging Consortium (ABIC) and Microscopy Australia (MA), have been working to create training opportunities for imaging scientists across the globe. During the COVID-19 pandemic, GBI and its partners developed a virtual training platform for independent learning (GBI Virtual Training Platform), with self-guided training modules spanning from microscope operation to facility management. More recently, in-person train-the-trainer (TtT) frameworks, including the Montreal Light Microscopy Course (MLMC), have been designed to deliver a multiplying effect by providing imaging scientists with the training and resources they need to run microscopy courses in their facilities, institutions, countries, and regions. Major goals of this initiative included empowering the next generation of imaging scientists by sharing resources and expertise, providing training in course logistics and organisation and building an international collaborative network of expert trainers to accelerate access to fundamental and advanced technologies. As of April 2026, the TtT project has consisted of three TtT courses, at least 11 second-generation microscopy fundamentals courses and impacted more than 400 course faculty, corporate partners and trainees from 25 countries. LAY DESCRIPTION: This manuscript demonstrates that global train-the-trainer (TtT) programmes in bioimaging create a powerful multiplier effect, rapidly expanding microscopy training capacity. By equipping imaging scientists with course materials, training, and access to a global trainer's network, it enabled second-generation courses across multiple regions, impacting over 400 individuals in 25 countries. It empowered the next generation and strengthened international collaboration to help democratise access to advanced technologies and expertise to accelerate scientific discovery and innovation. With sufficient resources, TtT is scalable across technology areas and research fields.

Mitochondrial segmentation and function prediction in live-cell images with deep learning

Yang Ding, Jintao Li, Jiaxin Zhang, Panpan Li, Hua Bai, Bin Fang, Haixiao Fang, Kai Huang, Guangyu Wang, Cameron J Nowell, Nicolas H Voelcker, Bo Peng, Lin Li, Wei Huang

Nature Communications · 2025

Mitochondrial morphology and function are intrinsically linked, indicating the opportunity to predict functions by analyzing morphological features in live-cell imaging. Herein, we introduce MoDL, a deep learning algorithm for mitochondrial image segmentation and function prediction. Trained on a dataset of 20,000 manually labeled mitochondria from super-resolution (SR) images, MoDL achieves superior segmentation accuracy, enabling comprehensive morphological analysis. Furthermore, MoDL predicts mitochondrial functions by employing an ensemble learning strategy, powered by an extended training dataset of over 100,000 SR images, each annotated with functional data from biochemical assays. By leveraging this large dataset alongside data fine-tuning and retraining, MoDL demonstrates the ability to precisely predict functions of heterogeneous mitochondria from unseen cell types through small sample size training. Our results highlight the MoDL's potential to significantly impact mitochondrial research and drug discovery, illustrating its utility in exploring the complex relationship between mitochondrial form and function within a wide range of biological contexts.

Evaluation of Serum Supplementation on the Development of Haemonchus contortus Larvae In Vitro and on Compound Screening Results

Sandani S Thilakarathne, Aya C Taki, Tao Wang, Cameron Nowell, Bill C H Chang, Robin B Gasser

International Journal of Molecular Sciences · 2025

A high-throughput platform for assessing the activity of synthetic or natural compounds on the motility and development of Haemonchus contortus larvae has been established for identifying new anthelmintic compounds active against strongylid nematodes. This study evaluated the impact of serum supplementation on larval development, motility and survival in vitro and its implications for phenotypic compound screening. Of five blood components assessed, 7.5% sheep serum significantly enhanced larval development, motility and survival compared to the original medium (LB*), leading to the formulation of an improved medium (LBS*). Proteomic analysis revealed marked differences in protein expression in larvae cultured in LBS* versus LB*, including molecules associated with structural integrity and metabolic processes. The phenotypic screening of 240 compounds ("Global Priority Box" from Medicines Malaria Venture) using LBS* yielded results distinct from those in LB*, highlighting the effect of culture conditions on screening assessments. These findings indicate/emphasise the critical need to evaluate and optimise culture media for physiologically relevant conditions in screening platforms, improving the reliability of anthelmintic discovery.

Constitutive ghrelin receptor activity enables reversal of dopamine D2 receptor signaling

Farhad Dehkhoda, Mitchell T Ringuet, Emily A Whitfield, Keith Mutunduwe, Fiona Whelan, Cameron J Nowell, Desye Misganaw, Zheng Xu, Noah B C Piper, Richard J Clark, Mohammed Akhter Hossain, Linda J Fothergill, Stuart J McDougall, John B Furness, Sebastian G B Furness

Molecular Cell · 2025

Changes in cellular output from one G protein-coupled receptor in another's presence have been attributed to dimerization and/or signal crosstalk. However, data distinguishing models are often lacking, and, in many cases, the molecular basis remains controversial. One such interaction is the reversal, inhibition to excitation, at the dopamine D2 receptor in the ghrelin receptor's presence in the spinal defecation center. This reversal is required for the physiological control of defecation, where dopamine is the relevant neurotransmitter. Using native neurons from mice and recombinant cells, we show that this reversal of dopamine D2 activity occurs downstream of calcium mobilization through a dominant signaling switch. Our results show that this reversal is a consequence of the ghrelin receptor's constitutive activity, where neither its agonism nor dimerization with the D2 receptor is required, in the systems tested. Dependence on constitutive activity may account for conservation of central-nervous-system ghrelin receptor in the absence of endogenous agonist.

Annexin-A1 deficiency uncovers female-specific pathways in blood pressure control and cardiovascular remodeling in mice

Jaideep Singh, Kristy L Jackson, Haoyun Fang, Feng Shii Tang, Cindy Gueguen, Alex M Parker, HsinErh Chen, Cameron J Nowell, Helen Kiriazis, Ekaterina Salimova, Owen L Woodman, Rebecca H Ritchie, Geoffrey A Head, David W Greening, Cheng Xue Qin

Communications Biology · 2025

Cardiovascular disease exhibits distinct sex-based differences, yet the mechanisms underlying these differences remain under-explored. The pro-resolving mediator annexin-A1 (ANXA1) is a pivotal regulator in inflammation resolution and tissue homeostasis, including within the cardiovascular system. However, the sex-specific differences in ANXA1 in blood pressure regulation have not been investigated. Here, we demonstrate that deficiency of ANXA1 exacerbates angiotensin II-induced adverse aortic and cardiac structural remodeling, mitochondrial proteome dysregulation, and impaired mitochondrial function in preclinical hypertensive models, exacerbated in females. Mechanistically, we demonstrate that estrogen upregulates ANXA1 levels, associated with dysregulation of inflammatory and mitochondrial networks, suggesting that the estrogen-ANXA1 axis plays a critical role in modulating inflammation and preventing pathological remodeling. In conclusion, this study advances the understanding of female-specific cardiac and aortic tissue and cellular alterations in hypertension, providing a platform for developing therapeutic ANXA1 mimetics that address the unique pathophysiological features of hypertension in females.

Hox-C12 coordinates β2-adrenoceptor coupling to a cAMP/calcium feedforward loop to drive invasion in triple-negative breast cancer

Terrance Lam, Bailey Cardwell, Bonan Liu, Cheng Peng, Mia Spark, Sandra Sursock, Cameron J Nowell, Andrew M Ellisdon, Aeson Chang, Alastair C Keen, Erica K Sloan, Michelle L Halls

Science Signaling · 2025

Noradrenaline released from sympathetic neurons accelerates cancer metastasis by activating β2-adrenergic receptors (β2-adrenoceptors) on tumor cells to promote invasion. We previously showed that the β2-adrenoceptor promotes invasive behavior in a metastatic triple-negative breast cancer (TNBC) cell line by activating a cAMP- and calcium-mediated feedforward loop. Here, we found this mechanism in most TNBC lines that have an active β2-adrenoceptor. Integrated analysis of transcriptomic datasets revealed HOXC12, which encodes a developmental homeobox transcription factor, as the most discriminating gene separating cell lines with the feedforward loop and those without it. The high expression of HOXC12 did not correlate with transcriptional changes in integral proteins associated with cAMP or calcium signaling, and immunostaining showed cytosolic localization of Hox-C12, suggesting that it played a nontranscriptional role. Knocking out HOXC12 prevented β2-adrenoceptor-mediated calcium signaling and invasion in cultured TNBC cells. In basal breast cancers, HOXC12 expression in tumors negatively correlated with overall and disease-free survival in patients. These findings identify a key mediator, Hox-C12, in the coordination of invasion driven by cAMP and calcium signaling in β2-adrenoceptor-responsive TNBC cells.

Contrasting roles for GLP-1R and GIPR in a model of diet-induced obesity

Jie Gao, Shelby Cree, Seungmin Ham, Cameron Nowell, Alex Parker, Peishen Zhao, Lynda Whiting, Kyle W Sloop, Ricardo J Samms, Patrick M Sexton, Denise Wootten, Dana S Hutchinson

Journal of Molecular Endocrinology · 2025

The glucagon-like peptide-1 receptor (GLP-1R) and glucose-dependent insulinotropic polypeptide receptor (GIPR) are important incretin receptors that are therapeutic targets for the treatment of type 2 diabetes and obesity. This study extensively characterised the metabolic phenotype of mice with global deletion of either the GLP-1R or GIPR side by side under identical conditions. Age-matched male wild-type (WT) C57Bl6NTac, GLP-1RKO or GIPRKO mice were placed on a high-fat or chow diet for 12 weeks, and a range of in vivo (weight gain, food intake, glucose tolerance, insulin tolerance, and whole-body energy metabolism) and ex vivo (white adipocyte lipolysis, brown adipose tissue and liver mitochondrial function, adipocyte and islet size, and hepatic steatosis) parameters were measured. While both WT and GLP-1RKO mice gained weight similarly on a HFD, obese high-fat-fed GLP-1RKO mice had altered glucose and insulin tolerance, and exhibited hepatic steatosis, highlighting the physiological importance of the GLP-1R in the regulation of blood glucose and lipid homoeostasis. In contrast, GIPRKO mice were partially resistant to diet-induced obesity compared to the WT mice, which was associated with a small reduction in food intake and intact epididymal and subcutaneous white adipocyte β-adrenoceptor-mediated lipolysis. Similarly, WT mice treated with a GIPR antagonist prevented weight gain due to a reduction in food intake on a HFD. These findings provide further support that the GLP-1R is important for normal glycaemic control, whereas the GIPR may play a role in the regulation of body weight.

Native adenosine A2A receptor solubilisation by a library of amphipathic copolymers

Michelle D Farrelly, Nazanin Mohebali, Emma Dal Maso, Cameron J Nowell, Denise Wootten, Patrick M Sexton, Lisandra L Martin, San H Thang

Biomaterials Science · 2025

Polymer nanodiscs are a research tool that allows membrane proteins (MPs) to be encapsulated by a surrounding amphipathic polymer, isolated and studied to understand their structural and physiological properties. An advantage of using polymer nanodiscs over other membrane mimetics can be found in their ability to natively solubilise membrane proteins (MPs) within an annulus of cellular phospholipids, however, potential polymer interactions with membrane constituents can hinder MP activity making the selection of a suitable polymer critical. This work demonstrates the native solubilisation of G-protein coupled A2A adenosine receptor (A2AR) by polymers with alternating units and cationic charge, poly(N-methyl-4-vinyl pyridinium iodide-co-N-alkyl-maleimides) (poly(MVP-co-AlkylMs)), and novel statistical copolymers with pseudozwitterionic charge, poly(potassium 3-sulfopropyl methacrylate-co-2-(trimethyl-amino) ethyl methacrylate-co-n-butyl methacrylate) (poly(KSPMA-co-TMAEMA-co-BMA)), both synthesised using RAFT polymerisation. After surveying a library of polymers within each class, A2AR extraction was the most efficient using poly(MVP-co-BM) (1 : 1 MVP : BM) and poly(KSPMA-co-TMAEMA-co-BMA) (1 : 1 : 1 KSPMA : TMAEMA : BMA). The optimal pH, temperature, solubilisation time, polymer concentration and ionic strength conditions required for extracting A2AR were identified and enabled a large-scale A2AR-nanodisc preparation. The yield of A2AR-poly(MVP-co-BM) was superior to A2AR-poly(KSPMA-co-TMAEMA-co-BMA) nanodiscs after affinity purification. Functional assessment of the reconstituted receptors was undertaken using fluorescence correlation spectroscopy (FCS) to determine the ligand binding capacity of A2AR stabilised within an alternating cationic poly(MVP-co-BM). These native nanodiscs retained their ability to specifically bind A2AR ligand antagonists.

Two-Photon Polymerized 3D Geometries to Study Single-Cell Mechanotransduction and Force Generation in Mesenchymal Stromal Cells

W Roshantha Perera, Aafreen Ansari, Cameron Nowell, Benjamin Geiger, Nicolas H Voelcker, Jessica E Frith, Victor J Cadarso

Small · 2025

Understanding cellular response to mechanical cues in three-dimensional (3D) environments remains a central challenge in cell biology. Shape and force distribution are key regulators of mechanosensing. In vivo, cells are embedded in 3D environments where force transmission and cytoskeletal behavior differ markedly from two-dimensional systems. However, current tools lack the resolution to precisely control single-cell geometry or quantify traction forces in defined 3D contexts. Here, a direct laser writing-based platform is presented that fabricates microscale cage structures capable of confining individual mesenchymal stem cells in defined 3D geometries while enabling high-resolution traction force measurements. The system allows independent control of cell volume and shape, and captures nanowire deflection as a readout of cell-generated forces at varying heights. Using this platform, this study reveals that 3D shape alone, modulates cytoskeletal organization, contractility, and localization of the mechanosensitive transcription factor Yes-associated protein (YAP) in a shape and time dependent manner. Square cages induced previously unreported vertical actin fibers and corner-enriched myosin accumulation, suggesting that pointed 3D geometry alters internal force distribution. Delayed YAP nuclear translocation indicates a time-sensitive mechanotransduction response to 3D confinement. Altogether, this platform offers a tunable 3D confinement tool and new insights into how shape alone direct cellular force architecture and mechanosensitive signaling.

Novel formylpeptide receptor 1/2 agonist limits hypertension-induced cardiovascular damage

Jaideep Singh, Kristy L Jackson, Haoyun Fang, Audrey Gumanti, Bethany Claridge, Feng Shii Tang, Helen Kiriazis, Ekaterina Salimova, Alex M Parker, Cameron Nowell, Owen L Woodman, David W Greening, Rebecca H Ritchie, Geoffrey A Head, Cheng Xue Qin

Cardiovascular Research · 2024

Aims: Formylpeptide receptors (FPRs) play a critical role in the regulation of inflammation, an important driver of hypertension-induced end-organ damage. We have previously reported that the biased FPR small-molecule agonist, compound17b (Cmpd17b), is cardioprotective against acute, severe inflammatory insults. Here, we reveal the first compelling evidence of the therapeutic potential of this novel FPR agonist against a longer-term, sustained inflammatory insult, i.e. hypertension-induced end-organ damage. The parallels between the murine and human hypertensive proteome were also investigated. Methods and results: The hypertensive response to angiotensin II (Ang II, 0.7 mg/kg/day, s.c.) was attenuated by Cmpd17b (50 mg/kg/day, i.p.). Impairments in cardiac and vascular function assessed via echocardiography were improved by Cmpd17b in hypertensive mice. This functional improvement was accompanied by reduced cardiac and aortic fibrosis and vascular calcification. Cmpd17b also attenuated Ang II-induced increased cardiac mitochondrial complex 2 respiration. Proteomic profiling of cardiac and aortic tissues and cells, using label-free nano-liquid chromatography with high-sensitivity mass spectrometry, detected and quantified ∼6000 proteins. We report hypertension-impacted protein clusters associated with dysregulation of inflammatory, mitochondrial, and calcium responses, as well as modified networks associated with cardiovascular remodelling, contractility, and structural/cytoskeletal organization. Cmpd17b attenuated hypertension-induced dysregulation of multiple proteins in mice, and of these, ∼110 proteins were identified as similarly dysregulated in humans suffering from adverse aortic remodelling and cardiac hypertrophy. Conclusion: We have demonstrated, for the first time, that the FPR agonist Cmpd17b powerfully limits hypertension-induced end-organ damage, consistent with proteome networks, supporting development of pro-resolution FPR-based therapeutics for treatment of systemic hypertension complications.

Lipoxin A4 improves cardiac remodeling and function in diabetes-associated cardiac dysfunction

Ting Fu, Muthukumar Mohan, Madhura Bose, Eoin P Brennan, Helen Kiriazis, Minh Deo, Cameron J Nowell, Catherine Godson, Mark E Cooper, Peishen Zhao, Barbara K Kemp-Harper, Owen L Woodman, Rebecca H Ritchie, Phillip Kantharidis, Cheng Xue Qin

Cardiovascular Diabetology · 2024

Background: Diabetic heart disease may eventually lead to heart failure, a leading cause of mortality in diabetic individuals. The lack of effective treatments for diabetes-induced heart failure may result from a failure to address the underlying pathological processes, including chronic, low-grade inflammation. Previous studies have reported that lipoxin A4 (LXA4), known to promote resolution of inflammation, attenuates diabetes-induced atherosclerosis, but its impact on diabetic hearts has not been sought. Thus, we aimed to determine whether LXA4 therapeutic treatment attenuates diabetes-induced cardiac pathology. Methods: Six-week-old male apolipoprotein E-deficient (ApoE-/-) mice were followed for 16 weeks after injection of streptozotocin (STZ, 55 mg/kg/day, i.p. for 5 days) to induce type-1 diabetes (T1DM). Treatment with LXA4 (5 μg/kg, i.p.) or vehicle (0.02% ethanol, i.p.) was administered twice weekly for the final 6 weeks. One week before endpoint, echocardiography was performed within a subset of mice from each group. At the end of the study, mice were euthanized with sodium pentobarbital (100 mg/kg i.p.) and hearts were collected for ex vivo analysis, including histological assessment, gene expression profiling by real-time PCR and protein level measurement by western blot. Results: As expected diabetic mice showed a significant elevation in plasma glycated hemoglobin (HbA1c) and glucose levels, along with reduced body weight. Vehicle-treated diabetic mice exhibited increased cardiac inflammation, macrophage content, and an elevated ratio of M1-like to M2-like macrophage markers. In addition, myocardial fibrosis, cardiomyocytes apoptosis and hypertrophy (at the genetic level) were evident, with echocardiography revealing early signs of left ventricular (LV) diastolic dysfunction. Treatment with LXA4 ameliorated diabetes-induced cardiac inflammation, pro-inflammatory macrophage polarization and cardiac remodeling (especially myocardial fibrosis and cardiomyocytes apoptosis), with ultimate improvement in cardiac function. Of note, this improvement was independent of glucose control. Conclusions: These findings demonstrated that LXA4 treatment attenuated the extent of cardiac inflammation in diabetic hearts, resulting in limited cardiac remodeling and improved LV diastolic function. This supports further exploration of LXA4-based therapy for the management of diabetic heart disease. The recent development of stable LXA4 mimetics holds potential as a novel strategy to treat cardiac dysfunction in diabetes, paving the way for innovative and more effective therapeutic strategies.

Impact of Drug Conjugation Site and Corona Chemistry on the Therapeutic Activity of Polymer Nanorod - Drug Conjugates

Nicole M Warne, Cameron J Nowell, Mai P Tran, John R Finnegan, Orlagh M Feeney, Kristian Kempe

Advanced Healthcare Materials · 2024

Biocompatible rod-shaped nanoparticles of controlled length can be produced through the heat-induced "living" seeded crystallization-driven self-assembly (CDSA) of poly(2-isopropyl-2-oxazoline)-containing block copolymers. With a hydrophilic poly(2-methyl-2-oxazine) or poly(2-methyl-2-oxazoline) corona, these nanorods have proven non-cytotoxic, non-hemolytic, and ideal for use as a polymer-based drug delivery system. This study demonstrates a facile, one-pot method for the synthesis of mycophenolic acid (MPA)-conjugated block copolymer "unimers" for use in seeded CDSA. Through altering block order during sequential monomer addition cationic ring-opening polymerization (CROP), MPA is conjugated to either the chain end of the core-forming or corona-forming block. This allows bioactive polymer nanorods to be prepared with MPA positioned at either the periphery of the corona, or at the core-corona interface of the nanorod formed during seeded CDSA. In vitro, these nanorods arrest growth in human T and B lymphocytes, with reduced effect in "off-target" monocytes when compared with unconjugated MPA. Furthermore, the conjugation of MPA to the core-corona interface of the nanorods leads to a slower release and reduced cytostatic effect. This study offers a robust investigation into the effect of steric hindrance and corona chemistry on the therapeutic potential of drug-conjugated CDSA nanorods and demonstrates the potential of poly(2-oxazoline)/poly(2-oxazine)-based CDSA nanomaterials as effective drug delivery platforms.

Impact of conjugation to different lipids on the lymphatic uptake and biodistribution of brush PEG polymers

Mohammad Abdallah, Lihuan Lin, Ian K Styles, Alexander Mörsdorf, James L Grace, Gracia Gracia, Cornelia B Landersdorfer, Cameron J Nowell, John F Quinn, Michael R Whittaker, Natalie L Trevaskis

Journal of Controlled Release · 2024

Delivery to peripheral lymphatics can be achieved following interstitial administration of nano-sized delivery systems (nanoparticles, liposomes, dendrimers etc) or molecules that hitchhike on endogenous nano-sized carriers (such as albumin). The published work concerning the hitchhiking approach has mostly focussed on the lymphatic uptake of vaccines conjugated directly to albumin binding moieties (ABMs such as lipids, Evans blue dye derivatives or peptides) and their subsequent trafficking into draining lymph nodes. The mechanisms underpinning access and transport of these constructs into lymph fluid, including potential interaction with other endogenous nanocarriers such as lipoproteins, have largely been ignored. Recently, we described a series of brush polyethylene glycol (PEG) polymers containing end terminal short-chain or medium-chain hydrocarbon tails (1C2 or 1C12, respectively), cholesterol moiety (Cho), or medium-chain or long-chain diacylglycerols (2C12 or 2C18, respectively). We evaluated the association of these materials with albumin and lipoprotein in rat plasma, and their intravenous (IV) and subcutaneous (SC) pharmacokinetic profiles. Here we fully detail the association of this suite of polymers with albumin and lipoproteins in rat lymph, which is expected to facilitate lymph transport of the materials from the SC injection site. Additionally, we characterise the thoracic lymph uptake, tissue and lymph node biodistribution of the lipidated brush PEG polymers following SC administration to thoracic lymph cannulated rats. All polymers had moderate lymphatic uptake in rats following SC dosing with the lymph uptake higher for 1C2-PEG, 2C12-PEG and 2C18-PEG (5.8%, 5.9% and 6.7% dose in lymph, respectively) compared with 1C12-PEG and Cho-PEG (both 1.5% dose in lymph). The enhanced lymph uptake of 1C2-PEG, 2C12-PEG and 2C18-PEG appeared related to their association profile with different lipoproteins. The five polymers displayed different biodistribution patterns in major organs and tissues in mice. All polymers reached immune cells deep within the inguinal lymph nodes of mice following SC dosing. The ability to access these immune cells suggests the potential of the polymers as platforms for the delivery of vaccines and immunotherapies. Future studies will focus on evaluating the lymphatic targeting and therapeutic potential of drug or vaccine-loaded polymers in pre-clinical disease models.

The oxytocin antagonist cligosiban reduces human prostate contractility: Implications for the treatment of benign prostatic hyperplasia

Beatrix Bester, Kristina Koslowa, Ann-Catherine Gronau, Andrea Mietens, Cameron Nowell, Michael R Whittaker, Adrian Pilatz, Florian Wagenlehner, Betty Exintaris, Ralf Middendorff

British Journal of Pharmacology · 2024

Background and purpose: With increasing life expectancy, benign prostatic hyperplasia (BPH) consequently affects more ageing men, illustrating the urgent need for advancements in BPH therapy. One emerging possibility may be the use of oxytocin antagonists to relax smooth muscle cells in the prostate, similar to the currently used (although often associated with side effects) α1-adrenoceptor blockers. Experimental approach: For the first time we used live-imaging, combined with a novel image analysis method, to investigate the multidirectional contractions of the human prostate and determine their changes in response to oxytocin and the oxytocin antagonists atosiban and cligosiban. Human prostate samples were obtained and compared from patients undergoing prostatectomy due to prostate cancer as well as from patients with transurethral resection of prostate tissue due to severe BPH. Key results: The two cohorts of tissue samples showed spontaneous multidirectional contractions, which significantly increased after the addition of oxytocin. Different to atosiban, which showed ambiguous effects of short duration, only long-acting cligosiban reliably prevented, as well as counteracted, any contractile oxytocin effect. Furthermore, cligosiban visibly reduced not only oxytocin-induced contractions, but also showed intrinsic activity to relax prostatic tissue. Conclusion and implications: Thus, the oxytocin antagonist cligosiban could be an interesting candidate in the search for novel BPH treatment options.

A therapy for suppressing canonical and noncanonical SARS-CoV-2 viral entry and an intrinsic intrapulmonary inflammatory response

Sandra L Leibel, Rachael N McVicar, Rabi Murad, Elizabeth M Kwong, Alex E Clark, Asuka Alvarado, Bethany A Grimmig, Ruslan Nuryyev, Randee E Young, Jamie C Lee, Weiqi Peng, Yanfang P Zhu, Eric Griffis, Cameron J Nowell, Brian James, Suzie Alarcon, Atul Malhotra, Linden J Gearing, Paul J Hertzog, Cheska M Galapate, Koen M O Galenkamp, Cosimo Commisso, Davey M Smith, Xin Sun, Aaron F Carlin, Richard L Sidman, Ben A Croker, Evan Y Snyder

PNAS · 2024

The prevalence of "long COVID" is just one of the conundrums highlighting how little we know about the lung's response to viral infection, particularly to syndromecoronavirus-2 (SARS-CoV-2), for which the lung is the point of entry. We used an in vitro human lung system to enable a prospective, unbiased, sequential single-cell level analysis of pulmonary cell responses to infection by multiple SARS-CoV-2 strains. Starting with human induced pluripotent stem cells and emulating lung organogenesis, we generated and infected three-dimensional, multi-cell-type-containing lung organoids (LOs) and gained several unexpected insights. First, SARS-CoV-2 tropism is much broader than previously believed: Many lung cell types are infectable, if not through a canonical receptor-mediated route (e.g., via Angiotensin-converting encyme 2(ACE2)) then via a noncanonical "backdoor" route (via macropinocytosis, a form of endocytosis). Food and Drug Administration (FDA)-approved endocytosis blockers can abrogate such entry, suggesting adjunctive therapies. Regardless of the route of entry, the virus triggers a lung-autonomous, pulmonary epithelial cell-intrinsic, innate immune response involving interferons and cytokine/chemokine production in the absence of hematopoietic derivatives. The virus can spread rapidly throughout human LOs resulting in mitochondrial apoptosis mediated by the prosurvival protein Bcl-xL. This host cytopathic response to the virus may help explain persistent inflammatory signatures in a dysfunctional pulmonary environment of long COVID. The host response to the virus is, in significant part, dependent on pulmonary Surfactant Protein-B, which plays an unanticipated role in signal transduction, viral resistance, dampening of systemic inflammatory cytokine production, and minimizing apoptosis. Exogenous surfactant, in fact, can be broadly therapeutic.

Evaluation of Ki-67, goblet cell and MUC2 mucin RNA expression in dogs with lymphoplasmacytic and granulomatous colitis

Chelsea Lim, Julien R S Dandrieux, Richard Ploeg, Cameron J Nowell, Simon M Firestone, Caroline S Mansfield

Veterinary Immunology and Immunopathology · 2024

Intestinal mucus barrier disruption may occur with chronic inflammatory enteropathies. The lack of studies evaluating mucus health in dogs with chronic colitis arises from inherent challenges with assessment of the intestinal mucus layer. It is therefore unknown if reduced goblet cell (GBC) numbers and/or mucin 2 (MUC2) expression, which are responsible for mucus production and secretion, correlate with inflammation severity in dogs with granulomatous colitis (GC) or lymphocytic-plasmacytic colitis (LPC). It is undetermined if Ki-67 immunoreactivity, which has been evaluated in dogs with small intestinal inflammation, similarly correlates to histologic severity in GC and LPC. Study objectives included comparing Ki-67 immunoreactivity, GBC population and MUC2 expression in dogs with GC, LPC and non-inflamed colon; and exploring the use of ribonucleic acid (RNAscope®) in-situ hybridization (ISH) to evaluate MUC2 expression in canine colon. Formalin-fixed endoscopic colonic biopsies were obtained from 48 dogs over an eight-year period. A blinded pathologist reviewed all biopsies. Dogs were classified into the GC (n=19), LPC (n=19) or no colitis (NC) (n=10) group based on final histopathological diagnosis. Ki-67 immunohistochemistry, Alcian-Blue/PAS staining to highlight GBCs, and RNAscope® ISH using customized canine MUC2-targeted probes were performed. At least five microscopic fields per dog were selected to measure Ki-67 labelling index (KI67%), GBC staining percentage (GBC%) and MUC2 expression (MUC2%) using image analysis software. Spearman's correlation coefficients were used to determine associations between World Small Animal Veterinary Association histologic score (WHS) and measured variables. Linear regression models were used to compare relationships between WHS with KI67%, GBC%, and MUC2%; and between GBC% and MUC2%. Median WHS was highest in dogs with GC. Median KI67% normalised to WHS was highest in the NC group (6.69%; range, 1.70-23.60%). Median GBC% did not correlate with colonic inflammation overall. Median MUC2% normalised to WHS in the NC group (10.02%; range, 3.05-39.09%) was two- and three-fold higher than in the GC and LPC groups respectively. With increased colonic inflammation, despite minimal changes in GBC% overall, MUC2 expression markedly declined in the LPC group (-27.4%; 95%-CI, -49.8, 5.9%) and mildly declined in the GC and NC groups. Granulomatous colitis and LPC likely involve different pathways regulating MUC2 expression. Decreased MUC2 gene expression is observed in dogs with chronic colitis compared to dogs without colonic signs. Changes in MUC2 expression appear influenced by GBC activity rather than quantity in GC and LPC.