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Paul Ehrlich ( ) Ehrlich reasoned that if a compound could be made that selectively targeted a disease-causing organism, then a toxin for that.

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Presentation on theme: "Paul Ehrlich ( ) Ehrlich reasoned that if a compound could be made that selectively targeted a disease-causing organism, then a toxin for that."— Presentation transcript:

1 Introduction to Antibody Drug Conjugates (ADC) John Milne PhD, NIBRT 20th June 2014

2 Paul Ehrlich ( ) Ehrlich reasoned that if a compound could be made that selectively targeted a disease-causing organism, then a toxin for that organism could be delivered along with the agent of selectivity. Hence, in 1913 he coined the phrase "magic bullet" (magische Kugel), his term for an ideal therapeutic agent that could be created that killed only the organism targeted. The concept of a "magic bullet" was to some extent realized by the invention of monoclonal antibodies as they provide a very specific binding affinity. Principle of ADC technology – to selectively deliver payloads through an antibody to enhance antibody activity and reduce systemic toxicity.

3 Antibody Drug Conjugates
Antibody Drug Conjugates (ADC) aim to take advantage of the specificity of monoclonal antibodies (mAbs) to deliver potent cytotoxic drugs selectively to antigen-expressing tumour cells. The antibody (or fragment) is linked via a stable chemical linker, with a labile bond to a biologically active cytotoxic (anti-cancer) drug.

4 Approved ADC’s Gemtuzumab ozogamicin (Mylotarg®): Approved 2000.
An anti-CD33 antibody conjugated to the cytotoxin calicheamicin: withdrawn from market in 2010. Brentuximab vedotin (Adcetris®): Approved 2012; Takeda An anti-CD30 antibody conjugated via cleavable linker to cytotoxic monoethyl auristatin (MMAE) - Hodgkins lymphona and anaplastic large cell lymphona’s. Ado-trastuzumab emtansine (Kadcyla®) Approved 2013; Genentech An anti-Her2 antibody conjugated via non-cleavable linker to cytotoxic “DM-1” (derivative of maytansine) - Her2 metastatic breast cancer.

5 Licensed ADC Products Emerging classes of armed antibody therapeutics against cancer Hess et al., Med. Chem. Commun., 2014, Advanced Article

6 Growth Potential Market for ADC’s predicted to grow significantly
18 of Top 20 Pharma involved in ADC Development. 50+ ADC molecules in clinical trials. Molecules in Phase I account for 70% of ADCs in clinical development. Global sales of ADC - $500M by 2016. The complexity of the ADC molecules adds to the uncertainty in the market. There are opportunities for pharmaceutical companies to replenish their diminishing pipelines and gain first mover advantage in the market. There have been several technological developments in the recent past with more stable linkers and potent cytotoxins allowing the development of next generation ADCs with improved safety/efficacy profile. Adapted from “Antibody Drug Conjugates Market ”, Roots Analysis, Business Research and Consulting.

7 The ADC space: selected deals of interest
Partnership Details of deal (US$ millions)* Date of deal Licensee (buyer) Licenser (seller) Upfront payment Milestone payments (potential) Celgene Sutro Biopharma $500 Not applicable Dec 2012 Abbott Seattle Genetics $25 $220 Oct 2012 Merck Ambrx $15 $288 Jun 2012 Eli Lilly ImmunoGen $20 $200 Dec 2011 Pfizer $8 Jan 2011 Novartis $45 Oct 2010 Deals typically also include an undisclosed single-digit percent of sales that is paid to the licenser for any ADC from the collaboration that makes it to market. Data taken Zolot et al., (2013)., Nature Reviews Drug Discovery 12,

8 Antibody Drug Conjugates
The concept of ADCs appears simple, however the development of efficacious products is a considerable challenge! Many parameters have to be optimised for a successful ADC product; ADC stability (the drug should not be released before the ADC reaches its in vivo target) Pharmacokinetics (the drug should not confer an undesired accumulation in a non-target organ) Retention of immunoreactivity Efficient release of the active form of the drug at the site of disease

9 ADC Toolkit ADC’s are constructed from three distinct components;
A mAb that is specific to a tumour antigen A highly potent cytotoxic agent A linker species that enables covalent attachment of the cytotoxin to the mAb through either the protein or glycan Pay Load can be; Chemotherapeutic Radioisotope Cytokine Protein based toxin

10 Adcetris® Brentuximab vedotin (Adcetris; Seattle Genetics)Approved by the FDA for the treatment of Hodgkin lymphoma (HL) and systemic anaplastic large cell lymphoma (ALCL) in August 2011, brentuximab is the first new drug for the treatment of HL in more than 30 years. It consists of an anti-CD30 antibody conjugated to the antimitotic agent monomethyl auristatin E (MMAE). CD30 is a cell membrane protein of the tumor necrosis factor receptor family. It was granted accelerated approval based on two single-arm multicenter clinical trials of patients with CD30-positive HL after failure of autologous stem cell transplant and patients with CD30-positive systemic ALCL who had previously received chemotherapy. The objective response rates (ORR) were 73% and 86%, respectively, while the complete remission (CR) rates were 32% and 57%, respectively, and the partial remission (PR) rates were 40% and 29%,

11 ADC Design ADC success depends on optimisation of each building block;
Antibody Linker Drug The antibody should target a well characterised antigen with high expression at the tumour site and low expression on normal tissue to maximise efficacy. Bifunctional linkers with attachment sites for both the antibody and drug are used to join the two components. Current linker attachment protocols rely on the modification of solvent accessible lysine or cysteine residues on the antibody, giving rise to ADC populations with variable DAR’s.

12 ADC Design As low drug loading reduces potency and high drug loading can adversely affect pharmacokinetics, DAR can significantly affect ADC efficacy. Crucially the selected linker must be stable in the circulation to minimise toxicity but also be cleaved rapidly after the ADC finds its target antigen. After binding to the target antigen the ADC receptor complex is internalised and once inside the cell the drug is released; Hydrolysis of the linker Enzymatic cleavage of the linker Degradation of the antibody

13 Drug Antibody (DAR) The average number of drugs that are conjugated to the antibody is the most important quality attributes of an ADC. The DAR determines the amount of “payload” that can be delivered to the tumour cell. This can directly affect both safety and efficacy.

14 The Linker The identity and stability of the linker is crucial to the success of an ADC. The linker must be stable to allow the ADC to circulate in the bloodstream before reaching the tumour site without prematurely releasing the free “cytotoxic” drug and damaging normal tissue. The linker should also be labile enough to release the free drug efficiently.

15 The Linker Two main classes of ADC drug linker that use different mechanisms for release of the drug from the antibody Cleavable linker Non-cleavable linker Class 1: Cleavable Linkers Lysosomal protease sensitive linkers Acid sensitive linkers Gluthathione sensitive linkers Class 2: Non-cleavable Linkers Ref: Teicher B A , and Chari R V Clin Cancer Res 2011;17:

16 Cleavable Linkers Lysosomal protease sensitive linkers
This strategy uses lysosomal proteases such as Cathepsin B that recognise and cleave a dipeptide bond to release the free drug from the antibody. Valine-citrulline dipeptide linkers are common in this class e.g. Adcetris (Seattle Genetics/Millennium). Acid sensitive linkers This strategy takes advantage of the low pH (pH ~5) within lysosomes to trigger hydrolysis of an acid labile group (hydrazone) within the linker to release the drug. Glutathione sensitive linkers This strategy exploits higher concentration of thiols such as glutathione within cells relative to the bloodstream. Disulphide bonds within the linker are stable in circulation but are reduced by intracellular glutathione to release the free drug.

17 Non-cleavable Linkers
This strategy depends on complete degradation of the antibody after internalisation of the ADC. The free drug is released with the linker attached to an amino acid from the mAb. Non-cleavable linker strategies are best applied to payloads that are capable of exerting their anti-tumour effect despite being chemically modified e.g. Kadcyla (trastuzumab-MCC-DM1) (Genentech/Immunogen). One advantage of non-cleavable linkers is their greater stability in circulation compared with cleavable linkers.

18 The Drug Payload Two main classes of ADC payload undergoing clinical evaluation. Drugs that disrupt microtubule assembly and affect mitosis Dolastatin 10-based auristatin analogs (Adcetris) Maytansinoids (Kadcyla) Drugs that target DNA structure, bind the minor groove of DNA causing DNA double strand cleavage Calicheamicin analogs (Mylotarg) These cytotoxic agents demonstrate at least fold greater potency in in vitro proliferation assays against a broad range of tumour cell lines compared with conventional chemotherapeutic agents. High potency is crucial because only an estimated 1-2% of the ADC dose will reach the tumour site, resulting in low intracellular drug concentrations!!!

19 Schematic illustrating the several steps from administration of the antibody conjugate to the patient to release of the toxic agent in the tumor cells. If the efficiency of each step is 50%, only 1.56% of the administered dose will reach the intracellular target. If the efficiency of each step is 50%, only 1.56% of the administered dose will reach the intracellular target. Ref: Teicher B A , and Chari R V Clin Cancer Res 2011;17:

20 Summary: Efficacy of ADC
Critical to the efficacy of an ADC are; the target site-specificity and binding properties of the antibody, the in vitro and in vivo stability of the linker and drug species, the potency of the drug both the distribution and average number of drug species on the antibody (DAR) These factors highlight the importance of; understanding the physiochemical properties of ADC’s and choosing the appropriate bioanalytical techniques to assess and monitor them during manufacturing and subsequent storage.

21 Conjugation Conjugation typically involves a controlled chemical reaction with specific amino acid residues exposed on the antibody surface. Selection of the appropriate drug-conjugation strategy significantly affects efficacy, Pharmacokinetics and tolerability. Two types of conjugation Chemical conjugation Site-specific conjugation

22 Chemical Conjugation A reactive moiety on the linker molecule is joined covalently to the antibody via an amino acid side chain, commonly the e-amino group of lysine. Alternatively in a two-step process the surface lysines on the antibody are first modified to introduce a reactive group, such as a maleimide and then conjugated to the drug linker containing a reactive handle (e.g a thiol group), e.g. Kadcyla. Alternatively a controlled reduction of existing disulphide bonds may liberate free cysteine residues on the antibody, which then react with a maleimide attached to the drug linker (e.g. Adcetris)

23 Conjugation The random conjugation processes produce heterogeneous mixtures of conjugated species with variable DAR’s. When lysines are used for conjugation, heterogeneity in overall charge can impact solubility, stability and pharmacokinetics. Thus the clinical success of an ADC produced by random conjugation depends on robust manufacturing processes that provide the ability to monitor, control and purify heterogeneous products

24 Site-Specific Conjugation
Next generation ADC’s are focused on homogeneous products derived via site-specific conjugation. Three differing strategies; Insertion of cysteine residues in the antibody sequence by mutation. Insertion of an “unnatural” amino acid with a bio-orthogonal reactive handle Enzymatic conjugation (utilising specificity of enzymes)

25 Summary: General Process Overview

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