Llama antibodies – known as VHHs or nanobodies – could be a gamechanger for precision medicine. Marion Cubitt, Director of Discovery at Isogenica, explains how an accidental discovery opened a fresh pathway for immunotherapy.

Tell us about Isogenica’s work

Isogenica are experts in the discovery of small-format antibodies, with a specialist focus on VHHs – single-domain camelid antibodies that are around a tenth of the size of a traditional monoclonal.

Using our proprietary CIS display technology and our ultra-high diversity synthetic libraries, we can mine much larger libraries than is possible with conventional screening techniques such as phage display, increasing the chances of finding the best binders.

Over the past 20 years, we have discovered novel VHHs as well as peptides and fibronectin-based molecules for big pharma, biotechs, and academic groups.

Using synthetic libraries means we can find new VHHs faster, because we don’t need to wait for an animal to develop an immune response to a target antigen. It also means we have more control over the screening conditions, such as enriching for particular domains, epitopes, or species cross-reactivity.

This gives greater diversity in the hits we get, especially for common targets that have already been tried in animals where it’s harder to find new sequences that haven’t already been discovered.

What is the company’s significance in personalised medicine?

Conventional monoclonal antibody-based drugs are highly specific to their target, which is great for minimizing side effects. However, they have a number of limitations, such as target availability, size, and engineerability. The simplicity and robustness of VHHs means they can be used as simple building blocks in more complex biotherapeutics, such as cell therapies and bi- or multi-specific immune engagers.

This latter modality is particularly well-suited to our rapid discovery capabilities and ‘virtual cancer patient’ collaboration with the University of Leicester – being able to quickly test bi-specific efficacy with a patient’s unique combination of tumour and immune system so they can be treated with the right drug.

The flexibility and engineerability of VHHs also unlocks the possibility of repurposing the same core VHH in different ways according to the principle of ‘theranostics’ – using the same building block as both a diagnostic and as a therapeutic, so you can be certain the target is expressed and that your drug is likely to work before administering an expensive medicine.

A good paradigm for this is in radiopharmaceutical approaches for cancer, where a VHH antibody is conjugated to a radioisotope for imaging followed by a drug-conjugated version of the VHH to treat the tumours. This is something we’ve looked at with Steven Archibald, Professor of Molecular Imaging at King’s College London.

What are VHHs – and what makes yours unique?

VHHs consist of a single protein domain that is equivalent to the variable heavy chain domain at the tip of a monoclonal antibody, rather than the paired heavy and light chains found in conventional Y-shaped monoclonal antibodies. They have all the target-binding activity of a full antibody but having evolved without a light chain means they have some interesting functional properties such as higher solubility and longer variable regions.

Isogenica’s VHHs are a carefully curated balance between llama and human sequences, and our different libraries vary in their degree of humanisation. This takes into account the natural repertoire of camelids and where that overlaps with human antibody repertoires.

 We have also been able to greatly reduce the frequency of manufacturing liabilities present in the libraries through our build methods. This means that we can isolate VHHs directly from the libraries without requiring time-consuming engineering to introduce mutations to support humanisation or manufacturing.

VHHs were discovered by accident – can you tell us briefly how? And why are llamas so important?

Until the late 1980s, antibodies with no light chain were viewed as defective. And in humans, they are. But some research by students using camel blood (human blood was considered far too risky in the early days of the HIV/AIDS crisis), revealed that in some species, the lack of a light chain was actually quite normal.

Further research showed that camelid species such as llamas, camels and alpacas all had these heavy chain-only antibodies. Curiously, sharks also have similar single-domain antibodies, but these are not genetically related to camelid VHHs. This example of convergent evolution demonstrates the advantage of a heavy-chain only molecule and all the special properties they have. Read more about the discovery of VHHs on our blog: isogenica.com/on-the-origin-of-vhhs-student-serendipity-and-coincidental-camels.

How did the partnership with the University of Leicester come about?

One of our former members of staff, Mandeep Sehmi, completed her PhD in immunology in Professor Martin Dyer’s group at the University of Leicester. When Isogenica began working on its own drug pipeline, Mandeep reached out to Martin to come and give a seminar and hear his thoughts about what was really needed in the clinic.

 I vividly remember him speaking so passionately about imagining a drawer he could open as a doctor and have not just one or two ‘best guesses’ for an individual patient, but a whole range from which you could select the medicine that would be most likely to work.

Using our rapid VHH discovery platform together with the expertise in immunology and ‘virtual patient’ approach at Leicester through a Knowledge Transfer Partnership funded by Innovate UK, we’re working towards achieving that vision.

We have a joint iCASE PhD student, Natasha Spena, who has done some brilliant work already in building bi-paratopic VHHs and testing them in in vitro models at Leicester. Initial results are looking good, and we can’t wait to see how her project develops.

What is your virtual patient? Is it proven to work yet?

Despite the immense progress in making mouse models as similar as possible to humans, the differences between human and mice and the lack of an intact human tumour-specific environment and structure are a significant limitation to predicting how a drug will perform in clinical trials.

What Harriet Walters and her team have developed at the University of Leicester is a more rigorous and specific test to allow more accurate prediction of how a drug will work in a particular patient where the tumour is treated in the context of the patient’s own immune system.

The virtual patient is a sophisticated pre-clinical personalised cancer model that retains the cells and structures that are found in the human body. This is a step further from the mouse models that are the current standard in the industry because we are actually testing how a patient’s immune system reacts to the drug in virtually the same tumour environment as if it were in their own body.

While it’s early days for testing drugs in virtual patient models, there are extremely encouraging indications that these are game-changing models in predicting the behaviour of new drugs going into the clinic.

(www.nature.com/articles/s41416-019-0672-6,                  www.pubmed.ncbi.nlm.nih.gov/35364017).

The generous access that patients have granted to their samples at the University of Leicester means that more data can be collected all the time to build the predictive power of the model and validate new drugs and understand where the limits lie for this exciting new technology.

Why has VHHs potential for use in cancer treatment not been fully recognised yet?

VHHs have been a late bloomer in the field of biotherapeutics. Large pharmaceutical companies are notorious for wanting to be first, often without having the guts to take the first leap into the truly unknown. That level of risk-taking is often left to smaller biotechs with fewer resources who end up moving more slowly.

In the VHH field, those risk-takers were Ablynx (now part of Sanofi), who gained FDA approval for the very first VHH drug caplacizumab (Cablivi) for the treatment of blood disorders in 2019. Since that first green light, interest in VHH-based therapeutics has exploded. Timelines are long in drug development, but I’m sure we’ll be seeing more and more VHH-based therapies in the coming years.

What other therapies and targeted drugs could they be used in?

The only limit is your imagination! Bi-specific immune engagers and radiopharma are some of the biggest areas at the moment. There is also strong interest in ADCs and cell and gene therapies, especially since the approval of the VHH-based CAR-T agent ciltacabtagene autoleucel (Carvykti®) in 2022. VHHs are particularly well suited to use as intracellular antibodies (‘intrabodies’) as their high solubility and short gene length (<400 bp per unit) means they can be delivered easily in vivo with conventional viral vectors (e.g. www.nature.com/articles/s41467-022-29703-9).

VHHs are also ideal for high-volume, low-cost uses for antibodies in therapeutics, diagnostics, research reagents and more. Because of their simple structure, they can be manufactured cheaply in microbial systems such as yeast, making them ideal for industrial applications as well as cost-effective for countries with limited financial resources.

What critical drawbacks of current therapeutics are you seeking to address?

Current antibody-based therapies for cancer have produced some incredible success stories, especially immunotherapies, but only for patients who respond, and often after a battle with traumatic side effects such as cytokine release syndrome.

All too often, patients will relapse because the tumour evolves resistance to the treatment, particularly if given as a monotherapy. We are working towards generating a wider selection of safer, more effective therapies to address the needs of a greater number of patients.

How optimistic are you that the new immunotherapies will work. When do you hope to receive meaningful feedback? Could they be a genuine gamechanger?

You have to be optimistic in antibody drug development. But you also have to be scientific and go by the data. The question I always ask myself is “What is the fastest way to get to the most meaningful result?”, and that is getting as close as you can to patients as early as ethically and practically possible.

The virtual cancer patient model could be a practical and fast way to put new therapies to the test, as well as reducing animal use, and we expect to know the outcomes for our lead molecules in 2025.

We hope that the therapeutics we are developing will be the first of many on a theme, building up what’s available ‘in the drawer’ for clinicians like Harriet and Martin to offer patients. That’s where the game will change. By rapidly addressing new and exciting targets using our synthetic VHH libraries, we hope to be able to plug these into our antibody engineering platform to efficiently re-target the same machinery for many patients.

How do you see the sector developing in the next 10-20 years (both realistic and ‘would love to have’ scenarios)

Realistically, I think the next 10 years will be slow and that’s down to money more than anything. Developing drugs is expensive, and right now a lot of the innovation is coming from academic spin-outs or small biotech companies who are struggling to find investment.

On the other side, you have increasingly squeezed healthcare budgets that are less able to pay for big-ticket therapies. I think that leaves the door open for lower-cost drugs, such as VHH therapeutics.

One lesson from Covid that I hope the pharmaceutical industry does not forget is the power of collaboration. We saw fast-tracking and efficient knowledge-sharing to move good research along at record-breaking speed. As we all become more mindful of our climate impact, I would love to see more large-scale, collaborative efficiencies that will become increasingly necessary to create more sustainable and affordable therapies.

Isogenica are leaders in antibody discovery and engineering, developing highly versatile, small-format VHH antibodies used to construct next generation biotherapeutics for the treatment of cancer, inflammation, and other serious diseases.

VHH can be assembled to create multi-specific biotherapeutics or used to achieve targeted drug delivery as components of ADCs and cell therapies including CAR-Ts.