Biology subjects

Parker, A. L.

Publications and source records attributed to Parker, A. L..

9 recordsLinked to original sources

bioRxiv2026

Pulsed priming with the FAK inhibitor narmafotinib enhances both gemcitabine/Abraxane and FOLFIRINOX chemotherapy response in pancreatic cancer

BackgroundPancreatic ductal adenocarcinoma (PDAC) is a particularly lethal malignancy with few treatment options available. Extensive remodelling of extracellular matrix (ECM) generates a highly fibrotic tumour landscape, which impairs therapeutic response. ObjectiveWe investigated whether stromal priming via the highly specific Focal Adhesion Kinase (FAK) inhibitor narmafotinib (AMP945) in combination with the two major standard-of-care chemotherapies in PDAC, gemcitabine/Abraxane and FOLFIRINOX, reduces fibrosis and enhances treatment efficacy. Design3D organotypic matrices, intravital imaging, and in vivo subcutaneous and orthotopic PDAC models were used to provide a rationale for a first-line priming regimen of narmafotinib prior to chemotherapy. ResultsNeoadjuvant chemotherapy induces fibrosis in PDAC indicating a need for upfront first-line priming of the ECM to normalise the stroma for optimal treatment response. Narmafotinib is a new potent small molecule FAK inhibitor. Phase I safety data shows excellent safety, tolerability, and pharmacokinetics following oral administration in humans. We reveal that narmafotinib treatment during early ECM remodelling ( priming) reduces fibrosis, while limiting subsequent PDAC invasion. Moreover, intravital imaging demonstrates real-time FAK inactivation and cell cycle stalling, leading to improved chemotherapeutic efficacy upon narmafotinib priming in vivo. Long-term assessment in patient-derived models shows that narmafotinib priming prior to gemcitabine/Abraxane or FOLFIRINOX reduces PDAC progression and extends survival in both chemotherapy settings. ConclusionsOur results using these Phase II-ready drug combinations strongly support the clinical assessment of narmafotinib in PDAC. Narmafotinib is currently in Phase Ib/IIa trials, assessing a pulsed dosing regimen prior to gemcitabine/Abraxane, and warrants further clinical assessment in combination with FOLFIRINOX. SIGNIFICANCE OF THIS STUDYO_ST_ABSWhat is already known on this topicC_ST_ABSO_LIPancreatic cancer (PC) is one of the most lethal malignancies and is characterised by a dense, fibrotic stroma, which impairs chemotherapy efficacy. C_LIO_LIThe non-receptor tyrosine kinase FAK is known to promote cancer fibrosis and therefore represents a therapeutic target to normalise the PC stroma and to improve chemotherapy performance. C_LI What this study addsO_LINeoadjuvant chemotherapy induces early fibrosis indicating a need for upfront first-line priming of the ECM to blunt or normalise stromal fibrosis for optimal response to therapy. C_LIO_LIThe small molecule inhibitor narmafotinib (which is currently under Phase Ib/IIa clinical trial assessment) shows high specificity towards FAK as well as desirable pharmacokinetics and pharmacodynamics in healthy human volunteers. C_LIO_LIEarly short-term narmafotinib priming reduces fibrosis and improves the efficacy of subsequent standard-of-care gemcitabine/Abraxane chemotherapy. C_LIO_LIFOLFIRINOX (oxaliplatin, irinotecan, leucovorin and 5-fluorouracil) is a multi-agent chemotherapy preferentially used in PDAC patients with good performance status. Our results demonstrate that narmafotinib priming also improves FOLFIRINOX efficacy, leading to extended survival in patient-derived PDAC models. C_LI How this study might affect research, practice, or policyO_LIThis study supports the clinical development of narmafotinib in combination with both gemcitabine/Abraxane (ACCENT trial) and further FOLFIRINOX standard-of-care chemotherapies for PDAC patient treatment. C_LIO_LIThe first-line priming strategy and early ECM normalisation used in this study may also be applicable to other combination therapy settings and warrants further investigation in ongoing clinical studies. C_LI

cancer biology↗

bioRxiv2025

Identification of a novel structural motif and overexpression of key transcripts elucidated in Adenovirus 10

Adenoviruses are widely used as vectors for subunit vaccines and oncolytic therapies. Efficient vectors must infect target cells and deliver therapeutic transgenes at high levels. Species D adenoviruses, such as human adenovirus type 10 (HAdV-D10), are promising candidates due to low seroprevalence in humans. Here, we present the cryo-electron microscopy structure of the HAdV-D10 capsid alongside transcriptomic profiling of infected cells to inform vector development. The fiber shaft, essential for cell entry, was resolved at 4.6 [A], revealing a previously uncharacterized umbrella motif. Viral transcript analysis using an ORF-centric pipeline uncovered key differences from HAdV-C5, including abundant expression of a transcript encoding a protein equivalent to mature protein VII. These findings highlight the importance of detailed vector characterization prior to clinical translation and support the advancement of HAdV-D10 as a next-generation platform for gene delivery and vaccine development.

microbiology↗

bioRxiv2025

Investigating endothelial cell transduction and hexon:PF4 binding of ChAdOx1 in the context of VITT

BackgroundVaccines against SARS-CoV2 have been essential in controlling COVID-19 related mortality and have saved millions of lives. Adenoviral (Ad) based vaccines have played an integral part in this vaccine campaign, with licensed vaccines based on the simian Y25 isolate (Vaxzevria, Astrazeneca) and human Ad type 26 (Jcovden, Janssen) widely adopted. As part of the largest global vaccination programme ever undertaken, ultrarare thromboembolic events have been described in approximately 1:200,000 vaccinees administered with Ad based SARS-CoV2 vaccines. ObjectivesThe mechanism underpinning these adverse events remain to be completely delineated, but is characterised by elevated autoantibodies against PF4 which, in complex with PF4, cluster, bind FC{gamma}RIIa on platelets and induce thrombus formation. Here we investigated the ability of ChAdOx1 to transduce and activate endothelial cells. MethodsUsing protein sequence alignment tools and in vitro transduction assays, the ability of ChAdOx1 to infect endothelial cells was assessed. Furthermore, the ability of ChAdOx1 infection to activate endothelial cells was determined. Finally, using surface plasmon resonance we assessed the electrostatic interactions between the ChAdOx1 hexon and PF4. Results and ConclusionsDespite lacking the primary cell entry receptor, Coxsackie and Adenovirus Receptor (CAR), ChAdOx1 efficiently transduced endothelial cells in a CAR-independent manner. This transduction did not result in endothelial cell activation. Purified hexon protein from ChAdOx1 preps did, however, bind PF4 with a similar affinity to that previously reported for the whole ChAdOx1 capsid. These data confirm the need to develop non-PF4 binding adenoviral capsids to reduce the potential adverse events associated with VITT.

immunology↗

bioRxiv2023

Broad sialic acid usage amongst species D human adenovirus.

Human adenoviruses (HAdV) are widespread pathogens causing infections of the respiratory and gastrointestinal tracts, genitourinary system and the eye. Species D (HAdV-D) are the most diverse species and cause both gastrointestinal tract infections and epidemic keratoconjunctivitis (EKC). Despite being significant pathogens, HAdV-D are understudied and knowledge around basic mechanisms of cell infection is lacking. Sialic acid (SA) usage has been proposed as a major mechanism of cell infection for EKC causing HAdV-D. Here, we provide apo state crystal structures for fiber knob proteins of 7 previously undetermined HAdV-D, and provide crystal structures of HAdV-D25, HAdV-D29 and HAdV-D53 knob proteins bound to SA. Biologically, we demonstrate that removal of cell surface SA reduced infectivity of HAdV-C5 vectors pseudotyped with HAdV-D fiber knob proteins, whilst engagement of the classical HAdV receptor, CAR was variable. Together, these data indicate an important role for SA engagement in the tropism of many HAdV-D and may facilitate the development of suitable antivirals to control EKC outbreaks.

microbiology↗

bioRxiv2023

Efficient Viral Capture and Inactivation from Bioaerosols Using Electrostatic Precipitation.

The presence of infectious viral particles in bioaerosols generated during laparoscopic surgery places surgical staff at significant risk of infection and represents a major cause of nosocomial infection. These factors contributed to the postponement and cancellation of countless surgical procedures during the early stages of the ongoing COVID-19 pandemic, causing backlogs, increased waiting times for surgical procedures and excess deaths indirectly related to the pandemic. The development and implementation of devices that effectively inactivate viral particles from bioaerosols would be beneficial in limiting or preventing the spread of infections from such bioaerosols. Here, we sought to evaluate whether electrostatic precipitation (EP) is a viable means to capture and inactivate both non-enveloped (Adenovirus) and enveloped (SARS-CoV-2 Pseudotyped Lentivirus) viral particles present in bioaerosols. We developed a closed-system model to mimic the release of bioaerosols during laparoscopic surgery. Known concentrations of each virus were aerosolised into the model system, exposed to EP using a commercially available system (UltravisionTM, Alesi Surgical Limited, UK) and collected in a BioSampler for analysis. Using qPCR to quantify viral genomes and transduction assays to quantify biological activity, we show that both enveloped and non-enveloped viral particles were efficiently captured and inactivated by EP. Both capture and inactivation could be further enhanced when increasing the voltage to 10kV, or when using two Ultravision discharge electrodes together at 8kV. This study highlights EP as an efficient means for capturing and inactivating viral particles present in bioaerosols. The use of EP may limit the spread of diseases, reducing nosocomial infections and potentially enable the continuation of surgical procedures during periods of viral pandemics. HighlightsO_LIBioaerosols released from patients during surgery have the potential to facilitate viral spread. C_LIO_LIUltravision technology works via the process of electrostatic precipitation. C_LIO_LIElectrostatic precipitation can be manipulated to capture and inactivate aerosolised viral particles, preventing viral spread. C_LIO_LIElectrostatic precipitation is effective against both enveloped and non-enveloped viral particles. C_LIO_LIElectrostatic precipitation represents a viable means to reduce nosocomial infections. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/529105v1_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@4b7fb1org.highwire.dtl.DTLVardef@ac6edaorg.highwire.dtl.DTLVardef@a1ea28org.highwire.dtl.DTLVardef@13f6c7_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2021

The Structure of ChAdOx1/AZD-1222 Reveals Interactions with CAR and PF4 with Implications for Vaccine-induced Immune Thrombotic Thrombocytopenia

Adenovirus derived vectors, based on chimpanzee adenovirus Y25 (ChAdOx1) and human adenovirus type 26 are proving critical in combatting the 2019 SARS-CoV-2 pandemic. Following emergency use authorisation, scale up in vaccine administration has inevitably revealed vaccine related adverse effects; too rare to observe even in large Phase-III clinical trials. These include vaccine-induced thrombotic thrombocytopenia (VITT), an ultra-rare adverse event in which patients develop life-threatening blood clots 5-24 days following vaccination. To investigate vector-host interactions of ChAdOx1 underpinning VITT we solved the structure of the ChAdOx1 capsid by CryoEM, and the structure of the primary receptor tropism determining fiber-knob protein by crystallography. These structural insights have enabled us to unravel key protein interactions involved in ChAdOx1 cell entry and a possible means by which it may generate misplaced immunity to platelet factor 4 (PF4), a protein involved in coagulation. We use in vitro cell binding assays to show that the fiber-knob protein uses coxsackie and adenovirus receptor (CAR) as a high affinity binding partner, while it does not form a stable interface with CD46. Computational simulations identified a putative mechanism by which the ChAdOx1 capsid interacts with PF4 by binding in the spaces between hexon proteins, with downstream implications for the causes of VITT. SummaryWe present the structure of the ChAdOx1 viral vector, derived from chimpanzee adenovirus Y25 at 4.2[A] resolution1. ChAdOx1 is in global use in the AstraZeneca vaccine, ChAdOx1 nCoV-19/AZD-1222, to combat the SARS-CoV-2 coronavirus pandemic. Recently observed, rare, adverse events make detailed mechanistic understanding of this vector key to informing proper treatment of affected patients and the development of safer viral vectors. Here, we determine a primary mechanism ChAdOx1 uses to attach to cells is coxsackie and adenovirus receptor (CAR), a protein which is identical in humans and chimpanzees. We demonstrate the vector does not form a stable CD46 interaction, a common species B adenovirus receptor, via its primary attachment protein. Further, we reveal the surface of the ChAdOx1 viral capsid has a strong electronegative potential. Molecular simulations suggest this charge, together with shape complementarity, are a mechanism by which an oppositely charged protein, platelet factor 4 (PF4) may bind the vector surface. PF4 is a key protein involved in the formation of blood clots2, and the target of auto-antibodies in heparin-induced immune thrombotic thrombocytopenia (HITT)3, an adverse reaction to heparin therapy which presents similarly to vaccine-induced immune thrombotic thrombocytopenia (VITT), a rare complication of ChAdOx1 nCoV-19 vaccination4-6. We propose a mechanism in which the ChAdOx1-PF4 complex may stimulate the production of antibodies against PF4, leading to delayed blood clot formation, as observed in VITT.

molecular biology↗

bioRxiv2020

The fiber knob protein of human adenovirus type 49 mediates highly efficient and promiscuous infection of cancer cell lines using a novel cell entry mechanism

The human adenovirus (HAdV) phylogenetic tree is diverse, divided across seven species and comprising over 100 individual types. Species D HAdV are rarely isolated with low rates of pre-existing immunity, making them appealing for therapeutic applications. Several species D vectors have been developed as vaccines against infectious diseases where they induce robust immunity in pre-clinical models and early phase clinical trials. However, many aspects of the basic virology of species D HAdV, including their basic receptor usage and means of cell entry, remain understudied. Here, we investigated HAdV-D49, which previously has been studied for vaccine and vascular gene transfer applications. We generated a pseudotyped HAdV-C5 presenting the HAdV-D49 fiber knob protein (HAdV-C5/D49K). This pseudotyped vector was efficient at infecting cells devoid of all known HAdV receptors, indicating HAdV-D49 uses an unidentified cellular receptor. Conversely, a pseudotyped vector presenting the fiber knob protein of the closely related HAdV-D30 (HAdV-C5/D30K), differing in four amino acids to HAdV-D49, failed to demonstrate the same tropism. These four amino acid changes resulted in a change in isoelectric point of the knob protein, with HAdV-D49K possessing a basic apical region compared to a more acidic region in HAdV-D30K. Structurally and biologically we demonstrate that HAdV-D49 knob protein is unable to engage CD46, whilst potential interaction with CAR is extremely limited by extension of the DG loop. HAdV-C5/49K efficiently transduced cancer cell lines of pancreatic, breast, lung, oesophageal and ovarian origin, indicating it may have potential for oncolytic virotherapy applications, especially for difficult to transduce tumour types. ImportanceAdenoviruses are powerful tools experimentally and clinically. To maximise efficacy, the development of serotypes with low pre-existing levels of immunity in the population is desirable. Consequently, attention has focussed on those derived from species D, which have proven robust vaccine platforms. This widespread usage is despite limited knowledge in their basic biology and cellular tropism. We investigated the tropism of HAdV-D49, demonstrating it uses a novel cell entry mechanism that bypasses all known HAdV receptors. We demonstrate, biologically, that a pseudotyped HAdV-C5/D49K vector efficiently transduces a wide range of cell lines, including those presenting no known adenovirus receptor. Structural investigation suggests that this broad tropism is the result of a highly basic electrostatic surface potential, since a homologous pseudotyped vector with a more acidic surface potential, HAdV-C5/D30K, does not display a similar pan-tropism. Therefore, HAdV-C5/D49K may form a powerful vector for therapeutic applications capable of infecting difficult to transduce cells.

microbiology↗

bioRxiv2019

Adenovirus serotype 26 utilises sialic acid bearing glycans as a primary cell entry receptor

Adenoviruses are clinically important agents. They cause respiratory distress, gastroenteritis, and epidemic keratoconjunctivitis (EKC). As non-enveloped, double stranded DNA viruses, they are easily manipulated, making them popular vectors for therapeutic applications, including vaccines. Species D adenovirus serotype 26 (HAdV-D26) is both a cause of EKC and other disease, and a promising vaccine vector. HAdV-D26 derived vaccines are under investigation as protective platforms against HIV, Zika, RSV infections and are in Phase-III clinical trials for Ebola.\n\nWe recently demonstrated that HAdV-D26 does not utilise CD46 or desmoglein 2 as entry receptors, whilst the putative interaction with Coxsackie and Adenovirus Receptor (CAR) is low affinity and unlikely to represent the primary cell receptor.\n\nHere, we definitively establish sialic acid as the primary entry receptor utilised by HAdV-D26. We demonstrate removal of cell surface sialic acid inhibits HAdV-D26 infection and provide a high-resolution crystal structure of HAdV-D26 fiber-knob in complex with sialic acid.

microbiology↗

bioRxiv2018

Diversity within the adenovirus fiber knob hypervariable loops influences primary receptor interactions.

Adenovirus based vectors are of increasing importance for wide ranging therapeutic applications. As vaccines, vectors derived from human adenovirus species D serotypes 26 and 48 (HAdV-D26/48) are demonstrating promising efficacy as protective platforms against infectious diseases. Significant clinical progress has been made, yet definitive studies underpinning mechanisms of entry, infection, and receptor usage are currently lacking. Here, we performed structural and biological analysis of the receptor binding fiber-knob protein of HAdV-D26/48, reporting crystal structures, and modelling putative interactions with two previously suggested attachment receptors, CD46 and Coxsackie and Adenovirus Receptor (CAR). We provide evidence of a low affinity interaction with CAR, with modelling suggesting affinity is attenuated through extended, semi-flexible loop structures, providing steric hindrance. Conversely, in silico and in vitro experiments are unable to provide evidence of interaction between HAdV-D26/48 fiber-knob with CD46, or with Desmoglein 2. Our findings provide new insight to the cell-virus interactions of HAdV-D26/48, with important implications for the design and engineering of optimised Ad-based therapeutics.

microbiology↗