Axial - Inventors #22
Analysis of exciting life sciences inventors and their inventions
More well thought out work can be found at — https://axial.substack.com/
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Inventors #22
A set of ideas and observations on inventions and discoveries in life sciences.
Immunology
The immune system and everything in it.
AIM2 in regulatory T cells restrains autoimmune diseases - https://www.nature.com/articles/s41586-021-03231-w - the Ting Lab at the University of North Carolina at Chapel Hill established a new role for AIM2 in T regulatory cells (Treg):
AIM2 is an inflammasome receptor that recognizes cytosolic double-stranded DNA (dsDNA) and activates the assembly of the inflammasome and production of IL-18 and IL-1β leading caspase-1 activation and pyroptosis
The group had the premise that AIM2 has a role in autoimmunity given the known roles of caspase-1 and the inflammasome in these diseases
Their first observation was that AIM2 is more highly expressed in Tregs, which control immune responses, than other immune cells. This led to the confirmation that TGFβ induces AIM2 expression in human and mouse Tregs. Lastly, the group found that the AIM2 promoter was occupied by transcription factors with established roles in Tregs activity and development: RUNX1, ETS1, BCL11B and CREB
Biochemical studies found that AIM2 reduces AKT phosphorylation by interfering with the RACK-1-PP2A complex, which may lead to AIM2’s secondary effects of promoting lipid metabolism in Tregs
Ultimately, the paper establishes a mechanism by which AIM2 regulations Tregs metabolism through altering the AKT-mTOR pathway
Biochemistry and structural biology
The granddaddy of them all.
Development of Antibody-Based PROTACs for the Degradation of the Cell-Surface Immune Checkpoint Protein PD-L1 - https://pubs.acs.org/doi/abs/10.1021/jacs.0c10008 - out of the Wells Lab at UCSF, the group invented antibody-based proteolysis-targeting chimeras (AbTAC) to selectively degrade cell-surface proteins:
Proteolysis-targeting chimeras (PROTAC) are small molecules that selectively degrade a target protein by recruiting an E3 ligase to promote target ubiquitination
To get to targeted degradation of cell-surface and extracellular proteins, the group used a bispecific antibody to engage a target and E3 ligase to promote degradation
To validate the idea, the group created a model of a cell-surface GFP along with RNF43, a cell-surface E3 ligase, tethered to an anti-GFP fragment. They showed GFP was degraded.
Next, an antibody specific to RNF43 was developed and fused it to an anti-PD-L1 antibody (Tecentriq). Excitedly, they found that the bispecific molecule degrades PD-L1 in 3 cancer cell lines with around 1/2 degradation efficiencies.
LYTACs from the Bertozzi Lab are a similar tool - https://www.nature.com/articles/s41586-020-2545-9
The paper invented a degrader that is a complete biologic, which may imbue higher specificity and lower off-target effects
Congrats Adam
Neuroscience
Roughly 20 years behind but set up to transform the concept of human.
CamKIIα Positive Interneurons Identified via A microRNA Based Viral Gene Targeting Strategy - https://www.jneurosci.org/content/40/50/9576 - the Han Lab at Boston University combined a viral labeling strategy and single cell sequencing to discover that calcium/calmodulin-dependent protein kinase type II subunit alpha positive (CaMKIIα+) neurons are both excitatory and inhibitory:
Viruses are useful labeling tools for neurons. Most existing methods rely on cell-specific promoters; however, the group had the clever idea to use microRNA (miRNA) binding sites as another way to selectively label neurons.
The idea is to incorporate a miRNA binding site into an AAV that will only label cells with low-to-no miRNA expression. The group named the method mAGNET: microRNA-guided neuron tags
As a proof-of-concept, the paper focused on miR-128, which has known roles in motor neurons and designed a viral vector with a CamKIIα-promoter (thought to be specific to excitatory neurons) and a binding site for miR-128
The mAGNET AAVs were injected into mice and discovered a new class of neurons (Lm128C) that are CamKIIα-positive, have low miR-128 expression, and are actually inhibitory neurons
In vivo calcium imaging along with other physiology experiments found that Lm128C neurons resemble interneurons, which connect motor and sensory neurons. A lot more work needs to be done to characterize this new neuron class.
The paper invents a new way to label neurons based on their endogenous miRNA expression profiles
Cell biology
Cell structure and function.
Metabolic regulation of skeletal cell fate and function in physiology and disease - https://www.nature.com/articles/s42255-020-00321-3 - a review out of the Carmeliet Lab at KU Leuven does a great job at characterizing the role of metabolism in skeletal cells:
Skeletal muscle metabolism is regulated by a wide set of signals from hormones and good to stimulus from neurons and immune cells
These signalling pathways are driven by a diverse group of cells from stromal cells to osteoblasts and hematopoietic-lineage cells
In short, it’s still an open field to understand how metabolism and various byproducts influence these cell interactions. Each cell type has a unique metabolism, which the authors suggest might create unique skeletal microenvironments.
Another important line of work is understanding the role of lipids here. They are an important source of energy for skeletal cells and are important to buffer against increases in sugar concentrations after meals, which is pretty important in diabetes and other chronic diseases
The role of skeletal metabolism in aging particularly sarcopenia is interesting as well as the important skeletal metabolites on stem cell differentiation
Genetics, genomics, and developmental biology
Heredity and variation.
Robust RNA editing via recruitment of endogenous ADARs using circular guide RNAs - https://www.biorxiv.org/content/10.1101/2021.01.12.426286v1 - the Mali Lab at UCSD invented a new RNA editing tool:
Endogenous ADARs (adenosine deaminases acting on RNA) can convert adenosines to inosine for RNAs. A gene editing strategy for RNA is to use antisense RNAs to recruit ADARs to specific transcripts.
A big issue in the field is gene editing efficiency, which the group hypothesized was due to the stability of the antisense RNAs
So they invented cadRNAs (circular ADAR recruiting guide RNA) that has higher stability probably due to the fact that the N-terminal is no longer exposed
First, the group integrated domains known to recruit ADARs into an cadRNA - domains were verified for editing at the 3’UTR of RAB7A mRNA transcript in HEK292 cells
Then cadRNAs with a combination of domains were screened to hone in on which ones confer the longest half-life. Moreover, length was increased and circularization was used to make cadRNAs more resistant to exonucleases. To create circular RNA, the group used a clever approach of having ribozymes integrated into the cadRNA to promote ligation by RtcB. This work led to improved stability by 2x-3.5x.
The cadRNAs were delivered as DNA with AAV and as in vitro transcribed (IVT) RNA to edit mPCSK9 transcripts in mice livers (38% RNA editing efficiency) and a mouse model for MPS I-H (12%). Moreover, editing was observed 96 hours post transfection where linear RNA had no activity.
The next steps are to discover new ADAR recruitment domains and understand the innate immune response to cadRNAs especially for IVT produced versions, where based modifications might be needed to subvert the immune system
The paper invents a new genome engineering tool that does not require an effector protein co-delivered that can tune mRNA translations and potentially fix G-to-A point mutations and premature stop codons in human disease
