Molecular glues, or molecular glue degraders, are currently experiencing a renaissance. These amazing molecules can help us develop therapies for previously untreatable diseases, including cancer. In this article, we’re covering the science of targeted protein degradation and molecular glues, and how our powerful Chemmunity screening platform is helping progress molecular glue compounds towards the clinic for the treatment of cancer and immune disorders.
Targeted Protein Degradation and Molecular Glue
Before we get into the oNKo-innate Chemmunity screening platform, let’s delve into some critical background information about targeted protein degradation and how it compares to traditional protein inhibitors. We’ll then explore a unique type of targeted protein degrader called a ‘molecular glue’ and how it works, how molecular glue compares to another major targeted protein degrader technology called ‘PROTACs’, and the rise in popularity of molecular glues for the treatment of disease.
What is targeted protein degradation?
The use of therapeutic molecules to direct the ubiquitin-proteasome system to mark specific proteins for destruction is known as targeted protein degradation (TPD). Think of a cell’s proteasome as the waste and recycling facility for proteins. The role of an E3 ligase is to tag proteins with a chain of ubiquitin molecules, which tells the proteasome that the protein needs to be destroyed. These proteins can be targets that are degraded during normal cellular function, including unwanted proteins such as those that have been misfolded.
Harnessing this highly effective recycling facility to remove certain disease-causing proteins is therefore an attractive therapeutic modality. As we will discuss below, this is especially true for proteins that have been traditionally considered ‘undruggable’.
Targeted protein degradation with molecular glues or PROTACs allows us to destroy these disease-causing proteins, potentially offering therapeutic options for previously untreatable diseases.
Targeted protein degradation vs. inhibition
Most drugs work by inhibiting the function of a target protein, an effect that is often temporary and limits the drug’s overall therapeutic effectiveness. In contrast, targeted protein degradation has a similar effect to gene knockdown – the pool of the target protein is physically reduced, leading to more robust suppression of its biological function.
For inhibition of a target protein to work, the inhibiting molecule (such as an antibody or traditional small molecule) must generally competitively bind to the target and stay there. This often requires high dose treatments, which can lead to toxicity. However, molecular glues can keep working after they have bound cereblon to tag multiple copies of a target protein for degradation, meaning that it’s possible to use very low doses.
Depending on the half-life of the protein in question, the duration of drug action achieved with targeted protein degradation can often be considerably longer than small molecule drug-mediated inhibition. Targeted protein degradation can also have other complex downstream consequences, reinforcing biological mechanisms or adding new ones, such as creating bioactive peptide fragments from the degraded protein.
These advantages mean that it can be preferable to use molecular glue degraders or PROTACs even when a target protein is druggable via inhibition.
What is a molecular glue and how does it work?
Molecular glues are small molecules that can bind or ‘stick’ two proteins together, typically those that would not normally bind to each other. Specifically, many molecular glues recruit the CUL4 E3 ligase complex through the substrate recognition component cereblon, linking it to target proteins and marking them for degradation by the ubiquitin-proteasome system we mentioned earlier.
They do this by altering the surface structure of cereblon and/or the target protein itself, stabilising a weak interaction, or in some cases, creating entirely new binding interfaces. This allows the CUL4 E3 ligase to interact with new proteins, known as neosubstrates, that it wouldn’t naturally degrade. PROTACs are similarly able to recruit E3 ligases to specific proteins, but they have a different structure and mode of action.
Not all molecular glues work as targeted protein degraders – some can actually do the opposite, stabilizing or activating proteins, or otherwise changing their function.
Molecular glue vs. PROTAC: what’s the difference?
While molecular glues change the surface of the E3 ligase or the target protein to promote binding, PROTAC molecules consist of two ends connected by a linker: one end of the PROTAC binds to the E3 ligase, the other binds to the target protein. While this structure makes PROTACs conceptually simpler to design, it creates challenges in achieving drug-like properties due to their increased size.
Another advantage of molecular glues over PROTACs is that they can alter the ‘shape’ of the interface between cereblon (or other substrate receptors) and the target protein to induce an interaction, whereas PROTACs often depend on pre-existing binding sites on the target protein. Additionally, given their smaller size and general understanding of their structure-activity relationship, it’s more straightforward to build and screen libraries of molecular glues than PROTACs.
Molecular glues in the treatment of disease: a complicated history
The best-known molecular glue is one you’ve almost certainly heard of: the notorious drug known as thalidomide. Developed in the 1950s, thalidomide was originally used to treat morning sickness in pregnant women and was eventually withdrawn from the market after causing significant birth defects.
It was later discovered that thalidomide was a ‘racemic’ mixture which included ‘left- and right-handed’ molecules due to the synthesis not being tightly controlled. While the right-handed version provides a treatment for morning sickness, the left-handed form can cause birth defects.
Further research determined that thalidomide acts as a molecular glue; while the left-handed form can cause birth defects by degrading important proteins during fetal development, the right-handed form and similar chemical structures can also degrade important proteins for cancers. In particular, derivatives of thalidomide known as immunomodulatory drugs (IMiDs) have shown benefit for the treatment of multiple myeloma and inflammatory diseases.
In the years since the thalidomide controversy, the potential of molecular glues in the treatment of disease has been investigated to varying levels of success. One of the key problems that has hindered progress in this field is that molecular glues are tricky to identify and design, with advances largely due to serendipitous discovery.
Only in recent years have molecular glues truly come of age, with companies now investing significant resources into understanding the structural and biophysical rules that govern neosubstrate specificity and the creation of large molecular libraries.
Yet there remain obstacles to harnessing the potential of molecular glues, not least due to the fact that the degradation of multiple different substrates often contribute to a desired biological effect. One way to address this challenge is to ‘work backward’ and use high-throughput screening to identify molecular glues that produce a desired phenotype and then reveal their neosubstrate profile.
The New Wave of Phenotypic Screening with Molecular Glues
Now we’ve established the basics of targeted protein degradation and molecular glues, let’s take a look at screening approaches for these compounds.
What is phenotypic screening?
Phenotypic screening involves identifying compounds that produce a desired change in cellular behaviour – for example, restoring a non-pathogenic phenotype to cells. The target and mode of action of the compound are generally unknown when conducting the primary phenotypic screen, these factors are investigated only if the compound can produce the desired effect.
The phenotypic screening approach has been enormously successful in producing many first-in-class drugs that are still used today, including the IMiD molecules we mentioned earlier. However, the molecular biology revolution and sequencing of the human genome has since led to a rise in what is known as target-based drug discovery.
Phenotypic vs target-based screening for drug discovery
In contrast to phenotypic screening approaches, target-based screening involves first identifying a target, forming a hypothesis about its role in disease, and then designing or identifying compounds that act upon that target.
For example, if a disease is known to be caused by a genetic mutation that encodes a faulty protein, researchers will study these molecular mechanisms and attempt to design or find compounds that will reduce the expression of the gene or inhibit the resulting protein.
However, if the goal is to alter a complex cellular phenotype, then there may not be a single obvious target to design a drug against. Instead, a screen that sets the cellular phenotype as an endpoint, then works backwards to understand the mechanism, may be the only approach.
Advantages of phenotypic screening
While target-based screening is immensely useful for drugging well-defined biology, phenotypic screening can reveal molecules that have multiple or unexpected mechanisms of action. This is particularly relevant for molecular glues which, as we discussed earlier, may degrade numerous targets or result in bioactive degradation products.
Another advantage of phenotypic screening is that the chemical ‘hits’ discovered using this approach are often more drug-like. This is because phenotypic screening is typically performed in living cells, meaning that any hit must be active and non-toxic to the cell, whereas target-based screens are often reduced to their minimal biochemical components (e.g. purified proteins).
Phenotypic screening can also produce exceptionally rich datasets; whereas target-based screens often have a single expected readout, a phenotypic screen can quantify many cellular parameters to provide much greater resolution of a compound’s activity.
By layering multiple phenotypic outputs from multiple cell types, it is then possible to build a comprehensive understanding of how drug-like molecules perturb networks of cells such as the immune system.
Phenotypic screening approaches for molecular glues
The recent explosion in popularity of molecular glues, and the relative simplicity of expanding the chemical space for these compounds, means that many companies have developed large libraries in the hope that some of them can be used to treat diseases like cancer. So, how do you identify molecular glues for the treatment of immune-related diseases?
That’s where phenotypic screening comes in. This approach enables the screening of molecular glue compounds at scale to discover their effects on cells, then works backwards to reveal their targets and modes of action.
In the immunology space, this can be a daunting task; the function of pathogenic immune cells or relationships between the immune system and the tumor microenvironment are highly complex, and navigating the effects of novel molecules on this biology requires an exceptional depth and breadth of knowledge of the immune system. Fortunately, that’s exactly the type of expertise we have here at oNKo-innate.
oNKo-innate’s Chemmunity Platform
oNKo-innate has unique expertise in identifying and drugging complex immune pathways for the treatment of cancer and other immune-related disease. Our validated functional genomics platform continues to reveal new therapeutic paradigms and unconventional molecular targets in primary immune cells. We believe that by fusing these data and screening experience with novel compound libraries we will transform how small molecules are developed for immune-related diseases.
With that in mind, we have brought molecular glues into our arsenal of tools for manipulating the biology of immune cells. We’ve adapted our advanced, high-throughput phenotypic screening platform to assess libraries of novel compounds – like molecular glues – in primary immune cells and complex multi-cell type immune assays. Indeed, our deep discovery efforts into the immunomodulatory properties of new molecular glues is truly reinvigorating their historical IMiD name.
Discovering chemicals that perturb immunity.
Our new Chemmunity platform generates highly scalable and detailed phenotypic fingerprints (or ‘phenoprints’) of how molecular glues (or other small molecules) perturb immune cell function. These multi-dimensional parameters include everything from molecular and transcriptional changes through to multi cell type effector function analysis.
Our expertise allows us to design and perform phenotypic screens that probe almost any immune function, not only to destroy cells and proteins, but to manipulate their biology and examine these effects in the context of disease.
The Chemmunity platform is equally powerful in exploring ‘both directions’ of the immune response. Not only can we perform screens that reveal potentiators of an anti-tumor or pro-inflammatory immune response, but we can also identify molecules that have a desired anti-inflammatory effect on certain immune populations. This unique physical screening platform is supported by our highly integrated computational biology capabilities, enabling structured and unstructured learning from these feature-rich, high-dimensional data.
We believe that a diversified screening stack is the key to unlocking novel and effective therapies that target the immune system. Many of our current pipeline assets, including our oNKo-037 cytokine checkpoint program, were identified using target-based screening approaches. Now, the Chemmunity platform allows us to tackle the other angle of drug discovery using phenotypic screening. By employing these two complimentary screening strategies, we’re ensuring that we leave no stone unturned when it comes to finding therapies that transform the treatment of cancer and immune-related disease, fulfilling our mantra: Any target, in any cell type.
Our deep knowledge of immunology and cancer biology, along with our prowess in phenotypic screening, allow us to ask the right questions of the data, screening vast compound libraries with confidence. While we use this platform to seed our own development pipeline, we are also building strategic relationships with key partners, helping to accelerate their clinical development timelines with the power of Chemmunity.
We hope this article got you as excited as we are about the potential of molecular glues in immunology and cancer immunotherapy! Using our Chemmunity platform, we are discovering and developing new molecules that shape the immune system for the treatment of cancer, autoimmune disorders, and other diseases.
Curious to know more about cancer biology, the immune system, screening technologies, and other novel therapies? Stay tuned for more articles like this one and explore our learning centre! You can also follow us on LinkedIn and X for regular updates on what’s happening in the immunology & immuno-oncology space.