Pharmacological dynamics and bioanalytical analysis of antibody-coupled drugs (ADCs).

Antibody drug conjugate is a compound that co-couples monoclonal antibodies with small molecule drugs through a linker.
ADC drug molecules contain antibodies, linker, small molecule drugs three structural modules.
typical ADC molecular structure diagram is shown in Figure 1.
1 ADC Molecular Structure Map (Source Wikipedia) German Nobel Laureate Paul Ehrlich first came up with the idea for ADC drugs in the early 20th century.
in 1958, Mathe first used anti-rat leukocyte immunoglobulin and methotrexate in the treatment of leukemia, the beginning of the study of antibody association drugs.
was limited by the anti-institutional preparation technology of the time, the antibody-coupled drugs of rat origin were not clinically successful, and the birth of hybrid tumor technology in 1975 gave birth to the first known ADC molecule.
, the selection of target antigens, antibody humanization technology, ultra-high cytotoxic small molecule drugs, antibody drug load and other issues with the technological progress of the time was solved one by one.
the FDA approved the first ADC drug, Gemtuzumab ozogamicin (market name Mylotarg), in 2000, post-market monitoring found that Mylotarg was ineffective, had significant side effects, and that patients were at greater risk of taking the drug than benefiting from it.
mylotarg pulled out of the market in 2010 because of the dismal performance of the market.
kadcyla became the second ADC drug approved in 2013.
Kadcyla is Genentech in the star product Herceptin based on the development of an ADC, along with Kadcyla's development, the domestic also set up a number of ADC start-ups, including Rongchang Bio (2008), Dongyu Pharmaceuticals (2009), Therese (2010), Hangzhou Doxi (2012) and so on, these early startups have become the domestic ADC drug research and development force.
Wyeth brought the adjusted dose of Mylotarg back to market in 2017 and successfully launched Besponsa the same year.
ADC drugs are truly on the market in 2019, with five ADCs approved so far in 2019.
2 A brief history of drug development at ADC (The Royal Society of Chemistry, 2019) The three phases of drug approval for ADC also truly reflect the three processes of ADC drug development.
technology platform for ADC drugs has also progressed over the course of these three processes.
the technical characteristics of ADCs at all stages, the industry divides ADCs into three generations.
the first generation ADC represented by Mylotarg, the second generation ADC represented by Adcetris and Kadcyla, and the third generation ADC represented by Besponsa, Padcev, Enhertu, etc.
1 ADC drug technology features at different times, nine ADC drugs have been approved for market by the FDA, and China's NMPA has approved two ADC drugs (Kadcyla and Adcetris).
August 2020, Rongchang Bio's Wedixito monoanti (RC48) submitted a listing application and was included in the priority review, if approved, it will be the first domestic ADC drug to be marketed.
2 Approved ADC Drug Note: Data from the antibody society website and the AAPS Journal, 2020) ADC drug development key elements target, antibody, effect molecule (payload), linker, DAR value (DAR, drug-to-antibody ratio, antibody binding drug molecule number is drug-antibody ratio), these five aspects are the ADC drug development needs to pay attention to the focus, but also the key element of ADC drug development.
the DAR values are mainly determined by linker and tagging techniques.
3 ADC Development Structure Elements Diagram ( Polymers for Advanced Technologies, Target 1. The nine ADC drugs approved for market involve a total of 8 targets, of which CD22, CD30, CD33, CD79b, BCMA and other 5 target adaptations are hematomas: HER2, Nectin-4, Trop-2 These three target adaptations are solid tumors, of which 2 targets have two HER approved products.
four ADC drugs approved before 2019 were only Kadcyla as solid tumor adaptations, while three of the five ADC drugs approved after 2019 were solid tumor adaptations.
it is not difficult to find that the hot spot of ADC drug development has shifted from hematoma to solid tumor.
finding suitable drug targets has become a hot spot for ADC research and development companies.
Because the drug erratic effect of ADC drugs mainly depends on small molecule drugs, and the target is only as a marker, as long as the ADC molecule can be recruited, as to whether the target has biological effects is not very important, so compared to monoclonal antibodies, ADC drug target selection is more.
potential targets are either over-expressed on the surface of the tumor or widely distributed in the tumor micro-environment.
Figure 4 ADC Can Develop Solid Tumor Antigen Targets (British Journal of Cancer, 2016) 2. Antibodies are used as ideal cytotoxic drug delivery vectors due to high antibody target specificity, long half-life, etc., ideal antibodies need to meet the target with a high affinity, usually Kd 0.1 to 1 nM, good blood stability, low immunogenicity and low cross-reaction.
IgG is more easily ingested by tumor cells and is often used as an antibody part of the ADC.
as the navigation system of ADC drugs, antibodies are mainly responsible for bringing small molecule drugs to the target cells to play a role, the antibody itself is not very important.
the antibody part of the ADC is different from the simple antibody drug in terms of mediated endo-swallowing and the therapeutic effect of the antibody itself, but the overall antibody selection and development strategy is not much different from the simple antibody drug.
3. The mechanism of effect molecules (Payload, with load/cytotoxic drugs/small molecule drugs) effect molecules determines the effectiveness of ADC and the production of toxicity.
currently, commonly used cytotoxic drug effect molecules are micro-tube inhibitors (e.g. auristatins, maytansinoids), DNA damage agents (e.g. calicheamicin, duocarmycins, anthracyclines, pyrrolobenzodiazepine dimers) and DNA transcription inhibitors (Amatoxin and Quinoline alkaloids) (SN-38).
nine ADC drugs approved for the market use a total of 6 different small molecule drugs, of which 3 ADC drugs use MMAE as a couple of drugs, 2 drugs use Calicheamicin as a couple of drugs, and the successful application of MMAF, DM1, SN-38, Dxd.
small molecule drug results with more applications are shown in Table 4.
most of the ADCs currently approved use heterogeneic connection technologies, namely amine-based lysine conjugation and -based cysteine conjugation.
each antibody has multiple Lys/Cys bits, ADCs that combine small molecule drugs with antibodies are a mixture of ingredients with different DAR values (0-8).
's in vitro effect is increased with the increase of DAR, but in vivo ADC ability does not increase with DAR value increase, may be due to hydrophobic small molecule drugs or / and linker increased to a certain extent, will lead to ADC in the water phase environment non-specific hydrophobic interaction and removal rate increase, in fact, the higher the DAR value toxicity, TI lower.
currently, using site-specificity, The combination method of homogeneous conjugation chemistries can effectively control the DAR value and obtain better ADC stability and aggregation, just as the ADC with a quality (homogeneous) DAR value of 2 has a lower hydrophobic and aggregation effect of ADC with an average DAR value of 3.4-4 than the random binding method, but TI may be limited (e.g. PBD and Aur0101).
Table 4 ADC drug is currently more studied small molecule drug list 4. Linker linker is a combination of co-price to connect the monoantigen and small molecule drug parts, linker stability in the blood flow is very important, generally linker than the ADC half-life is 10 times better.
ideal linker needs to be stable enough in the circulatory system to reduce the toxic reaction caused by off-target and release effect molecules once internalized into the target tissue.
, the linker's association determines many important properties of ADC drugs, such as DAR value distribution, therapeutic index (therapeuticindex, TI), PK/PD, and so on.
of linker and binding points is related to the stability of ADC, considering that small molecule drugs are generally hydrophobic, linker needs to have a certain hydrophobicity for easy connection.
the advantages and disadvantages of different conceded links are shown in Table 5, depending on whether linker is cracked within the cell: cleavable linker and non-cleavable linker.
table 5 The pros and cons of different conceding links (Protein Cell, 2018) Advances in pharmacodynamics The complex structure and mechanism of action make ADC pharmacological research more challenging.
ADC administration is mainly intravenous administration, the drug enters the body, can enter the cell through specific and non-specific ways.
The specific pathway is to bind to the receptors on the surface of the cell, to reach the cell through internalization, to enter the lysosome by swallowing small bodies, the acidic environment of the lysosome and the proteolytic enzyme will cause ADC degradation, the release of the effect molecules then into the cytostyte, and then through DNA insertion or inhibition of micro-tube synthesis and other ways to induce apoptosis;
addition, the ADC can also use the "bystand effects" to play a lethal role, that is, lysed or released cytotoxic drugs can enter adjacent cells, play a lethal role, and of course, will produce resistance.
Figure 5 ADCs' in vivo processes and their tumor immune mechanism schematics (BioDrugs, 2018) The main body of the ADC is antibodies (-98%), in addition to the pharmacodynamic characteristics of antibodies, but also because of the binding of effect molecules, so that it presents a unique PK characteristics.
this section mainly introduces the influence factors, PK characteristics and dynamic models of ADC PK.
1. After PK's influencer ADC enters the body, the effect molecules can be gradually dissogated from the main structure by enzymatic or chemical reaction, and the total antibody (Total antibody, Tab), binding antibody (ADC) and free effect molecule (unconjugated drug) (Figure 6) are obtained, and their changes in concentration in the body are essential for interpreting the pharmacy characteristics of the ADC.
The total antibody is the sum of the binding antibody and the naked antibody (DAR-0), the binding antibody is the antibody binding at least one effect molecule, is the prototype drug of ADC, the free effect molecule is the free effect molecule formed after ADC lysing or decomposed metabolism and/or retaining the effect molecular structure of the associated amino acid residue and/or linker effect molecular metabolite, is the material basis of ADC toxicity evaluation. The specificity of the combination of
and target, the stability of linker, the application of new effect molecules and other research is the key to the success or failure of ADC drugs, by optimizing the structure of ADC, improve its PK characteristics, can better clarify the dose-effect relationship, so as to evaluate safety and effectiveness.
Figure 6 ADC Drug Disposition (The AAPS Journal, 2020) 1.1 Structure 1.1.1 Antibody Structure Itself, Affinity, Specificity, FcRn binding capability, and Fc-effect functions of Fc-mediated antibodies such as ADC or CDC all affect the ADC's PK.
effects of ADC effector function on drug effectiveness and toxicity are not clear.
although Fc can enhance the role of ADC in killing tumors, Fc gamma is the main cause of off-target and dose-limiting toxicity.
ADC, which already has the missing effects of the IgG4 antibody, has been approved for listing (Mylotarg and Besponsa).
high antibody specificity avoids toxicity, and antibodies that lack specificity are eliminated by the circulatory system before reaching tumor tissue.
1.1.2 Effect Molecular Effect Molecule Introduction will make ADC aggregation, the physicophysical properties are unstable, not only because of its quantity and hydrophobic increase makes ADC hydrophobic significantly increased and unstable, but also because this binding effect will cause changes in the secondary and third-level structures of antibodies to reduce the stability of the composition.
, the permeability, metabolism, and whether the P-gp substrate of the effect molecule is bystander e.