Protein FAQ

Traditional cancer treatments have been largely unfocused in that they indiscriminately kill dividing cells, cancer or otherwise, which often leads to significant toxic side effects in patients. Targeted therapies on the other hand, try to inhibit one or more abnormal proteins that are found in proliferating cancer cells resulting in programmed cell death. One or more mutations in the p53 gene are found in at least 50% of all human cancers. These mutations begin a sequence of events that leads to loss of control of cell growth and proliferation. Due to its central importance in many human cancers, drugs that restore function to mutated p53 proteins have been described as one of the three holy grails of cancer research. Targeting specific abnormal proteins found only in cancer cells could be expected to be associated with less toxicity in normal healthy cells.

We plan to out license COTI-2 for clinical development so that we can continue the preclinical development of a number of other projects behind COTI-2. Our company is built around a computational platform for drug discovery and preclinical drug development. We intend to turn our attention to our acute leukemia program for which we also have patents issued and a growing body of positive preclinical data.

Given the central role of p53 mutations in human cancers, COTI-2 could represent a breakthrough therapy for many cancer patients if clinical trials confirm its’ activity in people. If we consider a specific disease such as ovarian cancer, p53 mutations are found in more than 90% of these tumours. Preclinical animal experiments with a human ovarian cancer known to have a p53 mutation and be resistant to conventional chemotherapy demonstrated that treatment with COTI-2 as a single agent either completely halted tumour growth or lead to dramatic tumour regression depending on the dose. COTI-2 was associated with no observable toxicity in these experiments.

The aim of these experiments was twofold. First, Dr. Mills at MDACC wanted to confirm that COTI-2 was preferentially killing cancer cells with p53 mutations. Secondly he wanted to identify the range of different mutations in the p53 gene that were responsive to COTI-2.

The simplest motifs are composed of multiple secondary structure units. These simple motifs can contain α-helices and β-sheets that are layered adjacent to one another either in the same direction (parallel) or in the opposite direction (anti-parallel). The simplest motif is the formation of a “loop”, known as a β-turn if it is short, while an unstructured connection is termed a “coiled region”.

These α-helices and β-sheets are not only joined by loops and β-turns but can also be joined by a β-sheet or an α-helix. Metal ions can also play a role in the formation of motifs.

Certainly. We're keenly exploring extracellular vesicles as mediums for cells to communicate. These cell made vesicles often carry microRNAs and other non-coding RNAs, presenting a unique avenue to understand how neurons modulate their environment, which is especially interesting in neurological conditions. We are exploring extracellular vesicles as these tools with which cells can communicate and transfer gene expression patterns. And we're looking at, for example, how early life brain tumours such as medulloblastoma can impact neuron development and activity and how this can affect later life pain processing and neurological conditions. This has been possible via funding from the Medical Research Foundation, which supported a big collaboration between the labs of Gareth Hathway, Beth Coyle, Vicky James, Anna Grabowska and myself in Nottingham.

I foresee a shift where RNA will gain more traction, considering recent successes in RNA-based therapies. Given the role sncRNAs play in axons, our research aligns well with this trend. By understanding the axonal RNA biology, we can pave the way for targeted therapies that could revolutionize how we approach various neurological conditions.

As an analogy, when the axon is scaled up to be the arm in a human body, you can think of the axon length in relation to the neuron's cell body as the equivalent distance from Nottingham to Paris. This significant length poses a logistical challenge for the neuron. For proteins to be expressed at the axon terminal, which is distant from the cell body, there's a requirement for translation regulation. In other words, there is a need for descentralized gene expression. Here, small non-coding RNAs play a pivotal role. Our work on microRNAs has shown their importance in spatial-temporal control of gene expression. This research aligns with the current interest in RNA biology and offers potential avenues for drug discovery, especially given the advances in RNA technologies post-covid.

At its core, pain involves sensory neurons that extend axons from their cell bodies into peripheral tissues and the spinal cord. Understanding these sensory terminals, especially their responses under various conditions such as inflammation, is pivotal. Our Axon-RNA-centric approach allows us to detect RNA changes and leverage them to alter sensitized states, providing a more accessible avenue for drug targeting.

Events like ELRIG 2023 are incredibly insightful. Although I'm aware of the academia-industry interface through collaborations, for instance with Eli Lilly, attending such events on a larger scale opens up a new perspective. The goal is to bridge the gap between fundamental science academics and the broader investment realm without losing focus on the core mechanisms. By attending, I hope to identify more opportunities for collaboration and application.

A significant collaboration is with the Versus Arthritis Pain Center in Nottingham, in particular with Vicky Chapman and Gareth Hathway. Initially, my work wasn't focused on pain research, but joining forces with experts from this center led half of my lab's work to be directed towards pain-related projects. Another meaningful collaboration involves studying motor neuron diseases alongside Prof. Rob Layfield and Dr. Dan Scott. This collaboration allows us to leverage our expertise in RNA and neuron development and apply it to conditions like motor neuron disease.

In a typical neuron, the cell body comprises only about 10-20% of the cytoplasm; the rest lies in the axon and dendrites. Thus, understanding neurons requires acknowledging the axon's role. We advocate for the research focus to consider this polarization and complex structure. By highlighting different transcriptomics within axons versus cell bodies and targeting their regulation, we can influence the entire neuron's functioning, potentially transforming treatment approaches.

My lab primarily focuses on neuroscience, but we approach problems from the perspective of axon biology. We're deeply interested in how neurons develop, undergo polarization and establish/maintain connectivity. This understanding helps not only in the development of the nervous system but also sheds light on age-related degeneration linked to the loss of axon connectivity. We use various cellular models, such as compartmentalised microfluidic neuronal cultures, to study how axons face different environments compared to cell bodies. Within this context, our research is focused on RNA biology, specifically the role of non-coding RNAs or microRNAs in regulating local axon protein synthesis. This knowledge is especially valuable when exploring different diseases and conditions like pain or neurodegeneration.

Success in research demands passion. It's crucial to find a subject that genuinely interests you and dedicate yourself to it. While there may be long periods where the efforts don't seem to yield desired or meaningful outcomes, perseverance is key. When success does come, it's a deeply rewarding experience. Hold onto that feeling.

While we've initiated dialogues on integrating AI, it hasn't transformed our work yet. However, I see it as a significant tool, especially for handling and processing big data. AI can help bridge the expertise gap in understanding both bioinformatics and biological terms, aiding in the efficient mining of information.

When the pH is exactly at the pKa value, a buffer arises in which the deprotonated and protonated amino acids exist in equilibrium. For example, when the pH = 2.34 (pKa of glycine), the solution comprises of 50% neutral molecules in which the carboxyl is deprotonated, and 50% positive molecules where the carboxyl is protonated. This pH produces the carboxyl buffer zone. If the pH s increased to that of the pKa of the amino group (9.60), another buffer is produced where there is an equilibration between the protonated neutral zwitterion and the deprotonated negative amino acid. The isoelectric point can, therefore, be approximated by averaging the two pKa values. More complex amino acids have more than two pKa values due to the presence of additional pKa values for their side chains.

The pKa for an amino acid is an acid dissociation constant that refers to the equilibrium between the protonated and deprotonated forms of the backbone amino group, backbone carboxyl group, and any potential acid/base component of the variable group. The net charge of the protein is determined by summing the charge of individual amino acids across the protein. pKa relates to the equilibrium constant and defines the transition between two structural forms of an amino acid–protonated and deprotonated. Amino acids possess distinct pKas for the carbonyl group, amino group, and any functional groups on the side chain that may be protonated or deprotonated. There is always one more structure than the number of pKa values for amino acids. For example, if there were two pKa values, three structures can be discerned. This is illustrated below using glycine, which has two pKa values – pKa 1 represents the (de)protonation of the carboxyl group and pKa2 represents the (de)protonation of the amine group: When the pH< pKa protonation of the amino and carboxyl groups occurs, resulting in a net +1 positive charge for glycine. When the pH is increased to a value between the two pKas(pKa2>pH> pKa1), the carboxyl group is deprotonated while the amino group remains protonated. This represents the zwitterionic form of the amino acid as it simultaneously possesses a positive and negative charge. When the pH> pKa2, the amino group is deprotonated and loses its charge. The amino acid now carries a net negative charge of -1. The zwitterionic form of the amino acid can exist at any pH value between pKa1 and pKa2.

These peptides, like the parent compound AC253, acted as antagonists at the AMY receptor. They were also resistant to protein breakdown, and crossed the blood-brain barrier easily when injected into the abdominal cavity, to localize in the hippocampus, which is crucial in memory. These peptides protected the brain against beta-amyloid injury, and normalized the AD-associated impairment of the memory-associated long-term potentiation of nerve impulses in the hippocampus. They improved memory testing results, and reduced the level of inflammation in the brain. These effects appear to be mediated via the blockade of AMY receptors. For instance, inhibition of microglial AMY receptors reduce the activation of the inflammasome NLRP3. This reduces the secretion of inflammatory chemicals in the surrounding brain tissue, which offers another mechanism for lower amyloid production. In addition, these peptides increase the rate of outflow of amyloid beta from the brain, which also contributes to a lower level of amyloid after treatment. These marked changes all occurred within a relatively short span of treatment. A very important additional finding was that treatment with these peptides brought about improvement in mice which were showing signs of well-established AD in the brain as well as in their behavior. This is unique in that most therapies fail to affect the progress of AD once it has begun to manifest clinically. Peptides also have fewer off-target effects. Small molecules are easy to administer, inexpensive to make and cross the blood-brain barrier more rapidly. For this reason, the team resorted to computational tools and artificial intelligence to come up with a new small molecular drug based on these peptides. This can be taken orally, and is similar in size and structure to the medications used for medical conditions like high blood pressure. An optimized version is being developed to enable human trials to be conducted. The work so far has taken about two decades, building step upon painstaking step to come up with the right solution. However, says Jhamandas, “Occasionally you come across a discovery that has the potential to change the game in a very fundamental way, like hitting a home run, and I'm very excited that we are really on to something here.” Short amylin receptor antagonist peptides improve memory deficits in Alzheimer’s disease mouse model. Rania Soudy, Ryoichi Kimura, Aarti Patel, Wen Fu, Kamaljit Kaur, David Westaway, Jing Yang & Jack Jhamandas. Scientific Reports, volume 9, Article number: 10942 (2019). https://doi.org/10.1038/s41598-019-47255-9. https://www.nature.com/articles/s41598-019-47255-9

In the study, published in Scientific Reports, the researchers built on their earlier discovery of the peptide called AC253. This compound was tested in mice with AD. It was found to block the attachment of beta-amyloid to a brain cell receptor called the amylin receptor, and thus inhibit its toxic effects, as shown by an improvement in spatial memory. However, it is difficult to administer this compound because it doesn’t cross the blood-brain barrier in large amounts, and is quickly broken down in the blood. The dosage must therefore be massively increased, pushing up the amounts required for efficacy and increasing the difficulty of administration, besides enhancing the chances of an immune reaction. One way out is to convert the formulation into a pill rather than an injectable form. The complex structure of AC253 makes this difficult as well. Instead, the team devised an ingenious solution. They cleaved the compound into smaller amylin peptides, or chains of 12-14 amino acids, and tested each for its anti-amyloid activity in old mice which showed signs of AD. In this way, they found two short peptides that had the same effects as the larger compound. In particular, the researchers identified a segment that was common to both peptides, namely, SQELHRLQTY.