Site-selective antibody modification by cysteine-to-lysine transfer (CLT)
Lead Research Organisation:
University College London
Department Name: Chemistry
Abstract
With CRUK recently highlighting that 1 in 2 of us will have cancer in our lifetimes, the requirement for progress in developing new medicines and improved diagnostics for oncology is a high priority for our society. The current state of the art in chemotherapeutics still relies heavily on untargeted cytotoxins, leading to severe side-effects which are intolerable in many cases. In contrast, the attachment of cytotoxins to a delivery agent, which targets the 'warhead' specifically to the tumor, offers the enticing possibility of 'magic-bullet' chemotherapies. Antibody-Drug Conjugates (ADCs) represent amongst the most promising class of such drugs in development, with 3 ADCs achieving clinical approval in the last few years. However, it is widely considered that the molecular construction of ADCs currently is still far from optimal, and that new technologies in this area are urgently required to help enable them to achieve their clinical potential. Most notably current approaches employed to attach the cytotoxic drugs to the antibodies lead to a highly complex mixture of products. The result is a drug which contains a vast number of distinct species, each with a different pharmacological profile.
In this project we are proposing to pioneer a new chemical approach for the generation of ADCs, which attaches the drugs at specific locations on the antibody, generating superior homogeneous conjugates. Our strategy, crucially, will not require genetic engineering of the antibodies to incorporate reactive handles, and is thus applicable directly to native 'off-the-shelf' antibodies. This will maximise the accessibility of homogenous ADCs to researchers across the world and ensure that the production yields are maintained as high as possible, ultimately reducing the cost of these relatively complex biopharmaceuticals.
We will achieve goal this by targeting specific lysine amino-acids on the surface of the antibodies. This is a challenging aim as there are numerous lysines present; and as such we are proposing to develop new methodology, in which the conjugation reagent is guided to a specific location by a neighbouring cysteine amino-acid. This unprecedented cysteine-to-lysine transfer (CLT) approach will generate amide linkages between the antibody and the drug which are already extremely well characterised and known to be robustly stable in vivo. This will afford a high-level of confidence which will facilitate rapid uptake in the field.
Overall this CLT platform will represent an optimum approach for producing ADCs and facilitate the wider success of these exciting targeted therapies.
In this project we are proposing to pioneer a new chemical approach for the generation of ADCs, which attaches the drugs at specific locations on the antibody, generating superior homogeneous conjugates. Our strategy, crucially, will not require genetic engineering of the antibodies to incorporate reactive handles, and is thus applicable directly to native 'off-the-shelf' antibodies. This will maximise the accessibility of homogenous ADCs to researchers across the world and ensure that the production yields are maintained as high as possible, ultimately reducing the cost of these relatively complex biopharmaceuticals.
We will achieve goal this by targeting specific lysine amino-acids on the surface of the antibodies. This is a challenging aim as there are numerous lysines present; and as such we are proposing to develop new methodology, in which the conjugation reagent is guided to a specific location by a neighbouring cysteine amino-acid. This unprecedented cysteine-to-lysine transfer (CLT) approach will generate amide linkages between the antibody and the drug which are already extremely well characterised and known to be robustly stable in vivo. This will afford a high-level of confidence which will facilitate rapid uptake in the field.
Overall this CLT platform will represent an optimum approach for producing ADCs and facilitate the wider success of these exciting targeted therapies.
Planned Impact
This project aims to increase our capability to produce targeted cancer therapies, and thus improved patient healthcare is the main, long-term, prospective impact. By overcoming key limitations in the design and synthesis of ADCs this work will contribute to the broader success of these therapeutics going forward. By developing a homogeneous conjugation strategy that can be employed directly on native antibodies, forming clinically validated amide linkages, maximises the chances of this approach being taken-up into clinical products. It also ensures the costs are kept as low as possible, and will even allow existing clinical antibodies to be re-purposed as antibody conjugates.
The technology development described in this proposal will be carried out using Herceptin, which is an antibody used in the treatment of breast, oesophageal and stomach cancers. This will ensure the products we generate are of direct potential for future clinical development in these areas.
The high economic value associated with antibody conjugates, particularly for therapy and imaging applications, also reveals that our approach could have very significant economic impact. For example the ADC field alone is expected to be worth $10 billion by 2024. We will work closely with UCL Business to obtain IP protection at key moments in the project, to establish a proprietary platform which will incentivize the future investment required to pursue final healthcare products.
Throughout this project we will disseminate our findings widely, through publications, posters and seminars, with the aim of facilitating impact throughout the various scientific communities in which antibody conjugates play diverse roles.
The technology development described in this proposal will be carried out using Herceptin, which is an antibody used in the treatment of breast, oesophageal and stomach cancers. This will ensure the products we generate are of direct potential for future clinical development in these areas.
The high economic value associated with antibody conjugates, particularly for therapy and imaging applications, also reveals that our approach could have very significant economic impact. For example the ADC field alone is expected to be worth $10 billion by 2024. We will work closely with UCL Business to obtain IP protection at key moments in the project, to establish a proprietary platform which will incentivize the future investment required to pursue final healthcare products.
Throughout this project we will disseminate our findings widely, through publications, posters and seminars, with the aim of facilitating impact throughout the various scientific communities in which antibody conjugates play diverse roles.
Organisations
Publications
Maneiro MA
(2020)
Antibody-PROTAC Conjugates Enable HER2-Dependent Targeted Protein Degradation of BRD4.
in ACS chemical biology
Farleigh M
(2021)
New Bifunctional Chelators Incorporating Dibromomaleimide Groups for Radiolabeling of Antibodies with Positron Emission Tomography Imaging Radioisotopes.
in Bioconjugate chemistry
Bahou C
(2021)
A Plug-and-Play Platform for the Formation of Trifunctional Cysteine Bioconjugates that also Offers Control over Thiol Cleavability.
in Bioconjugate chemistry
Spears RJ
(2022)
A novel thiol-labile cysteine protecting group for peptide synthesis based on a pyridazinedione (PD) scaffold.
in Chemical communications (Cambridge, England)
Haque M
(2021)
Site-selective lysine conjugation methods and applications towards antibody-drug conjugates.
in Chemical communications (Cambridge, England)
Forte N
(2019)
Cysteine-to-lysine transfer antibody fragment conjugation
in Chemical Science
Wall A
(2020)
One-pot thiol-amine bioconjugation to maleimides: simultaneous stabilisation and dual functionalisation.
in Chemical science
Forte N
(2018)
Homogeneous antibody-drug conjugates via site-selective disulfide bridging.
in Drug discovery today. Technologies
Patel M
(2023)
The Nitrile Bis-Thiol Bioconjugation Reaction
in Journal of the American Chemical Society
Morais M
(2019)
Application of Next-Generation Maleimides (NGMs) to Site-Selective Antibody Conjugation.
in Methods in molecular biology (Clifton, N.J.)
Forte N
(2019)
Cysteine-To-Lysine Transfer Antibody Fragment Conjugation
Description | In this project we have discovered a new approach to the construction of antibody conjugates, by the site-spectific modification of specific lysine residues. This method is referred to as 'Cysteine-to-Lysine' (CLT) transfer, and involved the development of reagents and protocols for carrying out reactions onto thiols (cysteine) which then flip onto nearby amines (lysines) to form robustly stable amide bonds. The method represents a novel approach to the construction of highly defined, homogenous, and stable antibody conjugates - which have broad potential applicability in the fields of chemistry, biology and medicine. Along with this methodology, and the insights it has provided into transfer ligation chemistry more generally, we have also explored a number of applications in the bioconjugation area. For example, we have described the formation of antibody-protac conjugates, as a potential new class of targeted therapeutics. |
Exploitation Route | The new discoveries made in this project will contribute directly to the keenly sought aspiration of having methods available to enable the construction of homogeneous antibody conjugates for therapeutics/diagnostics. Such methods offer the prospect of overcoming the heterogeneity associated with currently employed methods (which can lead to reduced therapeutic windows, poor PK, and challenging reproducibility); for example, as in the 11 antibody-drug conjugates (ADCs) which have made it to the clinic so far. The broader discoveries we have made in this project about chemical transfer ligations on proteins will contribute to the rapidly growing field of site-specific bioconjugations, both in academia and industry. The new antibody conjugates constructed, have also opened up the possibility of the delivery of new drugs to target cells (e.g. PROTACs), which can be further explored in the future. |
Sectors | Chemicals,Healthcare,Pharmaceuticals and Medical Biotechnology |
Description | C-Terminal Selective Ligation to Access Homogeneous Antibody Conjugates |
Amount | £444,295 (GBP) |
Funding ID | EP/T016043/1 |
Organisation | Engineering and Physical Sciences Research Council (EPSRC) |
Sector | Public |
Country | United Kingdom |
Start | 09/2020 |
End | 08/2024 |
Description | Conference talk |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Industry/Business |
Results and Impact | The PI gave a talk at internationally leading conference on antibody drug conjugates - World ADC London 2020, 3rd-4th March 2020. |
Year(s) Of Engagement Activity | 2020 |