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Dr Sheraz Gul examines how patient-driven imaging strategies can be utilised to aid the translation of initial research all the way into the clinic.

Pathology-based approaches are commonly used to diagnose and classify many diseases. Historically, this entailed qualitative or semi-quantitative histological analysis of patient samples for disease-specific markers. With advances in technologies such as digital pathology, multiplex spatial and temporal imaging, it is now possible to dissect disease pathophysiology in the context of mechanism of action, patient stratification and optimised treatment regimens. This strategy now has the potential to be used in a patient-driven approach to bridge basic translational research, drug discovery and clinical research.

Analysis of patient biopsies using immunohistochemistry is often used in a clinical setting to identify biomarkers for accurate disease diagnosis, characterisation and companion diagnostics during therapy. Such patient‑centric approaches are now also being implemented in preclinical drug discovery to de-risk the therapeutic agent that will enter clinical trials. This is largely being driven by technological advances such as multiplex spatial and temporal high-content imaging1 in combination with omics approaches. This can provide valuable insights into compound mechanism of action and potential insights into undesired side effects. The underlying causes of the latter are usually due to compound polypharmacology, which can be difficult to predict but may come to light when using multi-pronged spatial and temporal techniques in conjunction with omics methods to capture aspects of the genome, epigenome, metabolome, proteome and transcriptome.

Importantly, the above approaches could allow the early identification of potentially undesirable events in the clinic. This is elegantly illustrated by the marketed histone deacetylase (HDAC) inhibitor panobinostat, which has been shown in the clinic to increase the level of phenylalanine and decrease the level of tyrosine in a condition termed tyrosinemia.2 A proteomics‑based approach using panobinostat immobilised onto beads and incubated with cell extracts allowed the isolation of panobinostat-bound proteins. This led to the identification of its expected primary targets (HDAC) but also phenylalanine hydroxylase (PAH), tetratricopeptide repeat domain 38 (TTC38) and fatty acid desaturase (FADS1 and FADS2) with inhibition of the former being the cause of the tyrosinemia observed in the clinic. If this type of study was performed prior to panobinostat entering clinical trials, it could have undergone further optimisation to design a more selective compound with potentially no tyrosinemia side effects. This serves as a justification to perform comprehensive mechanism of action studies as early as possible in the drug discovery value chain. 

Chinese researchers discover a mutation, referred to as the “Shanghai APP” mutation, which has been linked to late-onset Alzheimer’s disease and offers fresh insights into the disease’s underlying molecular mechanisms.

Alzheimer’s disease, a progressive neurodegenerative disorder and the leading cause of dementia worldwide, affects millions of individuals. A recent breakthrough study, from Ruijin Hospital, affiliated with Shanghai Jiao Tong University School of Medicine, China, has revealed a newly discovered mutation, referred to as the “Shanghai APP” mutation, which has been linked to late-onset Alzheimer’s disease (LOAD) and offers fresh insights into the disease’s underlying molecular mechanisms.

Published in Genes & Diseases, The investigation centered around a Chinese patient who experienced memory decline in his mid-70s. Neuroimaging techniques confirmed the presence of widespread amyloid β deposition, a critical characteristic of Alzheimer’s.

Through molecular dynamics simulation and in vitro experiments, the researchers observed that the E674Q mutation in the APP gene increased the processing of amyloid precursor protein (APP), leading to the production of amyloid β—a toxic protein closely associated with Alzheimer’s. Furthermore, biochemical aggregation experiments indicated that the E674Q peptide exhibited a higher propensity for aggregation than the wild-type peptide, particularly forming filaments that linked multiple fibrils.

To assess the mutation’s impact in vivo, the researchers introduced the E674Q mutant APP gene into the hippocampi of two-month-old mice via adeno-associated virus (AAV) gene transfer. The findings demonstrated that the E674Q mutation led to impaired learning behavior and an increased pathological burden in the mouse model, validating its pathogenic role in Alzheimer’s. Remarkably, the E674Q substitution within the amyloid processing sequence is the sole known pathogenic mutation causing LOAD.

This discovery carries significant implications as it opens up new avenues for understanding the development of LOAD and potentially paves the way for more effective treatments. By comprehending how the specific E674Q mutation contributes to the onset and progression of Alzheimer’s, scientists hope to develop targeted therapies or interventions capable of slowing or halting the disease’s advancement.

The identification of the novel Shanghai APP mutation provides researchers with a unique opportunity to delve deeper into the molecular mechanisms underlying LOAD. Further investigation into the effects of the E674Q mutation is crucial to explore potential avenues for the development of therapies or interventions. This advancement has the potential to improve the lives of tens of millions of patients and their families by offering new strategies for preventing or treating this devastating form of Alzheimer’s.

The research team’s groundbreaking findings shed light on the complex nature of Alzheimer’s and may inspire future studies aimed at unraveling additional genetic and molecular factors contributing to the disease. With each new discovery, scientists move closer to understanding and combating this global health challenge.

New imaging approach reveals that changes in retinal microcirculation may indicate cerebrovascular diseases that involve reduced blood flow.

The brain is one of the most metabolically active organs in the human body. Although it represents only about 2 percent of the human body’s weight, it receives 15 to 20 percent of the body’s total blood supply. Disrupted blood flow to the brain over a long period of time, a condition known as “chronic cerebral hypoperfusion” (CCH), can lead to serious cerebrovascular diseases such as white matter disease.

CCH manifests as lesions in the white matter, a brain region vulnerable to problems with blood supply. Unfortunately, CCH has no available cure. An early diagnosis by visualising the microvascular changes in the brain that occur prior to lesion development is thus crucial. However, such a diagnosis is challenging with the available imaging techniques.

In a recent study published in Neurophotonics, researchers from USA and China led by Baoqiang Li, Associate Professor at the Chinese Academy of Sciences, investigated whether blood flow in the retina at the microscopic level could be used to predict cerebrovascular diseases involving hypoperfusion. To test this hypothesis, the team developed an innovative imaging approach based on two-photon microscopy.

Conveniently, insights into the microvasculature of the brain may come from our eyes. The retina at the back of the eye is a peripheral part of the central nervous system and shares many similarities with cerebral brain matter. But it has fewer nerve cell types and a simpler structure, making it an excellent target for studying neural circuitry and neurovascular coupling.

The researchers first induced CCH in mice by slightly blocking their carotid arteries. A week after, they studied one of their eyes directly under a two-photon microscope. They observed and counted red blood cells (RBCs) circulating within individual capillaries in the retinal microvasculature of mice by labelling their blood plasma with a fluorescent tag.

The objective of these experiments was to quantify the flux of RBCs in as many capillaries as possible. Accordingly, the researchers compared their results with those of similar measurements made on cerebral grey and white matter in a previous study performed under similar experimental conditions.

Following careful statistical analysis, they found that the mean capillary RBC flux in the retina was more significantly affected by CCH than those in the white and grey matter. While the mean RBC flux decreased by 56 percent in the retina of CCH mice compared to that in normal mice, the corresponding reductions were only 36 percent and 6 percent in their white and grey matter, respectively.

Overall, the findings of this study indicate that microcirculation in the retina could be a promising predictor of CCH, and potentially serve as an early diagnostic biomarker for cerebrovascular diseases. Moreover, the imaging approach developed by the researchers is efficient, has high signal quality, and can be implemented with a standard commercial two-photon microscope.

Neurophotonics Associate Editor Andy Shih of the Seattle Children’s Research Institute, US, remarks, “Being able to image retinal vascular physiology in vivo without adaptive optics is innovative. The results from this study may encourage further application of conventional two-photon imaging to retinal research.”

Researchers have identified distinct differences among the cells comprising a tissue in the retina, findings that could help develop precise therapies for retinal diseases.

In a new study, researchers form the US National Eye Institute (NEI) have identified distinct differences among the cells comprising a tissue in the retina that is vital to human visual perception. The scientists discovered five subpopulations of retinal pigment epithelium (RPE), a layer of tissue that nourishes and supports the retina’s light-sensing photoreceptors. Using artificial intelligence (AI), the researchers analysed images of RPE at single-cell resolution to create a reference map that locates each subpopulation within the eye. The ground-breaking research, which was recently published in Proceedings of the National Academy of Science, could help find more precise cell and gene therapies for retinal diseases.

Age and disease can cause metabolic changes in RPE cells that can lead to photoreceptor degeneration. The impact on vision from these RPE changes varies dramatically by severity and where the RPE cells reside within the retina. For example, late-onset retinal degeneration (L-ORD) affects mostly peripheral retina and, therefore, peripheral vision. Age-related macular degeneration (AMD), a leading cause of vision loss, primarily affects RPE cells in the macula, which is crucial for central vision.

The researchers aimed to determine if there are different RPE subpopulations that might explain the wide spectrum of retinal disease phenotypes. They used AI to analyse RPE cell morphometry, the external shape and dimensions of each cell. They trained a computer using fluorescently labelled images of RPE to analyse the entire human RPE monolayer from nine cadaver donors with no history of significant eye disease.

Morphometry features were calculated for each RPE cell – on average, about 2.8 million cells per donor; 47.6 million cells were analysed in total. The algorithm assessed each cell’s area, aspect ratio (width to height), hexagonality, and number of neighbours. Previous studies had suggested that RPE function is tied to the tightness of cellular junctions; the more crowded, the better for indicating cellular health.

Based on morphometry, they identified five distinct RPE cell subpopulations, referred to as P1-P5, organised in concentric circles around the fovea, which is the centre of the macula and the most light-sensitive region of the retina. Compared to RPE in the periphery, foveal RPE tend to be perfectly hexagonal and more compactly situated, with higher numbers of neighbouring cells.

Next, they analysed RPE from cadavers with AMD. Foveal (P1) RPE tended to be absent due to disease damage, and the differences among cells in the P2-P5 subpopulations were not statistically significant. Overall, the AMD RPE subpopulations tended to be elongated relative to RPE cells not affected by AMD.

To further test the hypothesis that different retinal degenerations affect specific RPE subpopulations, they analysed ultrawide-field fundus autofluorescence images from patients affected by chloridaemia, L-ORD, or a retinal degeneration with no identified molecular cauase. While these studies were conducted at a single point in time, they still demonstrated that different RPE subpopulations are vulnerable to different types of retinal degenerative diseases.

Age-related morphometric changes also may appear in some RPE subpopulations before they’re detectable in others. These finding could help inform future studies using non-invasive imaging technologies, such as adaptive optics, which resolve retinal cells in unprecedented detail and could potentially be used to predict changes in RPE health in living patients.

Researchers found that ‘rational vaccinology’ increases potency by changing the structural location of antigens and adjuvants.

Scientists globally are investigating vaccines for cancer, some with ambitious goals. One such group of scientists at from the International Institute for Nanotechnology (IIN) at Northwestern University, US.

The study was recently published in Nature Biomedical Engineering.  

“The work shows that vaccine structure and not just the components is a critical factor in determining vaccine efficacy,” said lead investigator Chad Mirkin, director of the IIN. “Where and how we position the antigens and adjuvant within a single architecture markedly changes how the immune system recognises and processes it. 

This new heightened emphasis on structure has the potential to improve the effectiveness of conventional cancer vaccines, which historically have not worked well, Mirkin said.

Mirkin’s team has studied the effect of vaccine structure in the context of seven different types of cancer to date, including triple-negative breast cancer, papillomavirus-induced cervical cancer, melanoma, colon cancer and prostate cancer to determine the most effective architecture to treat each disease.  

Conventional vaccines take a blender approach

With most conventional vaccines, the antigen and the adjuvant are blended and injected into a patient. There is no control over the vaccine structure, and, consequently, limited control over the trafficking and processing of the vaccine components. Thus, there is no control over how well the vaccine works. 

“A challenge with conventional vaccines is that out of that blended mish mosh, an immune cell might pick up 50 antigens and one adjuvant or one antigen and 50 adjuvants,” said study author Assistant Professor Michelle Teplensky. “But there must be an optimum ratio of each that would maximise the vaccine’s effectiveness.”

Enter SNAs (spherical nucleic acids), which are the structural platform — invented and developed by Mirkin — used in this new class of modular vaccines. SNAs allow scientists to pinpoint exactly how many antigens and adjuvants are being delivered to cells. SNAs also enable scientists to tailor how these vaccine components are presented, and the rate at which they are processed. Such structural considerations, which greatly impact vaccine effectiveness, are largely ignored in conventional approaches. 

Vaccines developed through ‘rational vaccinology’ offer precise dosing for maximum effectiveness 

This approach to systematically control antigen and adjuvant locations within modular vaccine architectures was created by Mirkin, who coined the term rational vaccinology to describe it. It is based on the concept that the structural presentation of vaccine components is as important as the components themselves in driving efficacy.  

“Vaccines developed through rational vaccinology deliver the precise dose of antigen and adjuvant to every immune cell, so they are all equally primed to attack cancer cells,” said Mirkin.

Building an (even) better vaccine 

The team developed a cancer vaccine that doubled the number of cancer antigen-specific T cells and increased the activation of these cells by 30 percent by reconfiguring the architecture of the vaccine to contain multiple targets to help the immune system find tumour cells. 

The team investigated differences in how well two antigens were recognised by the immune system depending on their placement — on the core or perimeter — of the SNA structure. For an SNA with optimum placement, they could increase the immune response and how quickly the nanovaccine triggered cytokine (an immune cell protein) production to boost T cells attacking the cancer cells. The scientists also studied how the different placements affected the immune system’s ability to remember the invader, and whether the memory was long-term. 

“Where and how we position the antigens and adjuvant within a single architecture markedly changes how the immune system recognises and processes it,” Mirkin said.

US researchers administered a therapy to Alzheimer’s patient-derived neurons in the lab, eliminating deteriorating cells, leading to positive consequences for the remaining healthy cells.

Scientists from the Salk Institute, US, have found that neurons from people with Alzheimer’s disease show deterioration and undergo a late-life stress process called senescence. These neurons have a loss of functional activity, impaired metabolism, and increased brain inflammation.

The researchers’ discovery, published in Cell Stem Cell, found that targeting the deteriorating neurons with therapeutics could be an effective strategy for preventing or treating Alzheimer’s disease.

“Our study clearly demonstrates that these non-replicating cells are going through the deterioration process of senescence and that it is directly related to neuroinflammation and Alzheimer’s disease,” said Professor Rusty Gage, president of the Salk Institute and Chair for Research on Age-Related Neurodegenerative Disease.

As cells age, they can undergo cellular senescence, which contributes to tissue dysfunction and age-related disorders. Senescence is also thought to play a role in cellular stress, molecular damage, and cancer initiation. However, scientists previously believed that senescence primarily occurred in dividing cells, not in neurons. Little was known about the senescence-like state of ageing human neurons. 

 In this study, the team took skin samples from people with Alzheimer’s disease and converted those cells directly into neurons in the lab. They tested these neurons to see if they undergo senescence and examined the mechanisms involved in the process.

They also explored senescence markers and gene expression of post-mortem brains from 20 people with Alzheimer’s disease and matched healthy controls. This allowed the team to confirm that their results from the lab held true in actual human brain tissue.

They found that senescent neurons are a source of the late-life brain inflammation observed in Alzheimer’s disease. As the neurons deteriorate, they release inflammatory factors that trigger a cascade of brain inflammation and cause other brain cells to run haywire. Additionally, the gene: KRAS, which is commonly involved in cancer, could activate the senescent response.

The authors note that the consequences of even a small number of senescent neurons in the aging brain could have a significant impact on brain function. This is because a single neuron can make more than 1,000 connections with other neurons, affecting the brain’s communication system.

In addition to these findings, the researchers also administered a therapeutic cocktail of Dasatinib and Quercetin to the patient neurons, in a dish. Both drugs are used to remove senescent cells in the body in conditions such as osteoarthritis, so the authors wanted to see if they were effective in senescent cells in the central nervous system as well.

They found that the drug cocktail reduced the number of senescent neurons to normal levels. Targeting senescent cells could thus be a useful approach for slowing neuroinflammation and neurodegeneration in Alzheimer’s disease.

Yet, the therapeutic cocktail in this study cannot normally enter the brain. However, there are known medications that can cross the blood-brain barrier that likely act in a similar manner and could, possibly, be used as a treatment option in the future.  

In the future, the authors plan to test some of the drugs that can enter the brain to see how they affect senescent neurons. They will also explore the driving mechanisms of senescence and see if certain brain regions are more prone to this deterioration than others.

Penn Medicine researchers are interested in how cardiac cells use DNA to establish and maintain their specification.

Scientists have long been fascinated by the intricate process of cell development and specialisation. But…how do cells transform into different types, such as heart, liver, or skin cells?

While researchers have focused on studying specific proteins, the broader understanding of how these proteins influence the activity of hundreds of genes to facilitate the transformation from one cell type to another remains elusive. The mystery surrounding this process has prompted researchers from the Perelman School of Medicine at the University of Pennsylvania, US, to embark on a new study looking at DNA in cardiac cells.

Thanks to a generous $6 million, seven-year grant from the National Heart, Lung, and Blood Institute of the National Institutes of Health (NIH), the Perelman School of Medicine, US, is set to launch an ambitious research project aimed at unravelling the relationship between DNA and the development and maintenance of heart cells. The insights gained from this study could pave the way for future breakthroughs in cardiac disease treatment.

The researchers hypothesise that nuclear architecture, which governs the accessibility of genes within a cell, plays a critical role in determining the identity of a cell. Specifically, they plan to investigate how the three-dimensional packaging and organisation of DNA impact cell development. By delving into the intricacies of how DNA folds and twists within the confined space of a cell nucleus, the scientists hope to shed light on the enigmatic process by which heart cells, such as heart muscle cells, endothelial cells, smooth muscle cells, and cardiac fibroblasts, acquire their unique identities.

“This research has the potential to significantly advance our understanding of how cardiac cells arise and maintain their identity throughout a lifetime,” said lead author Dr Rajan Jain, Assistant professor of Medicine and Cell and Developmental Biology at the Perelman School of Medicine. “By examining congenital heart disease and other cardiac conditions through the lens of DNA organisation, we may uncover therapeutic opportunities that have thus far remained untapped.”

The nucleus exerts control over the genes that determine cell identity. Jain’s previous work suggests that the way DNA is folded and arranged within the nucleus influences the identity of the cell. The researchers aim is to unravel the role of genome folding in governing cell behaviour, particularly in heart cells, and identify the key processes involved in this regulation. Additionally, they will explore how the spatial positioning of DNA influences gene activity during heart cell development, providing insights into the maintenance of heart cell identity.

An accurate and comprehensive understanding of heart cell development is crucial for maintaining overall cardiovascular health. Any deviations or abnormalities in this process can contribute to conditions such as congenital heart disease or cardiomyopathy. Therefore, by shedding light on the mechanisms that dictate heart cell identity, the researchers hope to pave the way for more effective therapeutic interventions.

Jain expressed optimism about the potential impact of this research, stating, “Traditionally, therapies targeting specific proteins in the nucleus were deemed implausible. Drawing inspiration from previous work, I envision that this research will ultimately enable us to design new medications that directly target the organisation of DNA.”