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Thank you for your kind introduction.
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So good afternoon everyone.
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Thank you for your time and interest.
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I'm Tomohiro Fujii, an ADC researcher in Ajinomoto, so I truly appreciate that opportunity to speak at this Oxford Global conference.
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Today, I'd like to talk about Ajicap technology that utilise site-specific conjugation and the stable linker.
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I want to show you the detail of both technologies as well as their applications.
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Before I begin, let me quickly introduce our company Ajinomoto.
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Ajinomoto is Japanese food company that was founded in 1909 and we primarily provide a food seasonings and the frozen food worldwide.
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And also we are expanding CDMO services.
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So we operate under the brand name Ajinomoto Biopharma.
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Ajinomoto Biopharma Services offering the comprehensive CDMO services worldwide with locations in Belgium and the United States, India and Japan.
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We offer the capacity and quality to scale up your process from the clinical development to manufacturing.
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Our services are designed to meet current and the future demand of our client, ensuring the stable scale up and seamless scale up of their promising therapeutics.
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Here are the services we are offering worldwide pharmaceutical and biotechnology companies.
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As we can see, we have original manufacturing platform for various sizes of molecules and last year Forge Biologics adjoined to the company.
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So now we are offering the gene therapy services as well.
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And also we have our original ADC manufacturing platform termed AJICAP that will be discussed in this presentation.
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So as you know, to maximise the function of antibody and the payload in ADCs, site specific conjugation and the stable linker are necessary.
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So because the payload must not be detached inside a blood circulation and normal tissue to reduce toxicity while it has to be released inside a cancer cell to be cytotoxic.
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So, although it seems to be contradictory between being stable and able to disconnect stable conjugation site and the stable linker can achieve this balance.
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From this perspective, we have been developing both technologies AJICAP conjugation and the novel linker technologies.
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The use of both technologies enabled to control their number to precisely 2 and the resulting ADC's with high efficacy and low toxicity.
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From next slide, I want to show you some advantages of our technologies.
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First, I'd like to talk about AJICAP conjugation.
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Traditional ADC's was synthesised through the random conjugation, typically through the lysine or cysteine chemistries.
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However, due to this heterogeneity of the resulting conjugate, traditional ADC showed a wide DL distribution and a high toxicity and a low efficacy.
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To address these issues, site-specific conjugations are gathering the strong interest in this ADC field.
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Site-specific conjugation ADCs not only provide a homogeneous ADCs but also expand the therapeutic window.
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So mainly site-specific conjugation is divided into the four categories.
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First category is the genetic modification and the second is enzymatic modification.
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And also there is a combination of genetic and enzymatic operation as well.
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And there is a chemical modification including a disulfide reduction.
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So these methodologies have some disadvantages in process and the cost while having a lot of benefits.
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So now we can add AJICAP conjugation as a next site-specific conjugation model.
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From next slide I'm going to show you some priorities of our technology.
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First of all, I'd like to talk about the background of AJICAP reagent, that is a key component in this technology.
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AJICAP conjugation was inspired by affinity peptide that has a strong binding affinity with the Fc portion of human IgGs.
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So this non coherent interaction between FC portion and the peptide was confirmed by X-ray analysis and this careful analysis.
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These binding sites were close to Lys248 and 288 respectively.
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So we modified these peptides to create a set of AJICAP reagent possessing the appropriately linker and the reactive group to form covalent bond with a Lys248 or 288.
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We termed this innovative reagent AJICAP reagent.
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So here is the mechanism of AJICAP conjugation.
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AJICAP reagent bind to the FC portion of human IgGs this via non covalent bond subsequently the neighbouring lysine attacks the CYO Ester in this reagent to form this antibody peptide conjugate.
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Since this peptide is not necessary in ADC structure, so this peptide moiety is cleaved by mild chemical reagent hydroxylamine.
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Resulting orthogonal groups such as thiol can react with a various payload not only high potent small molecule but also biomolecule like oligonucleotide protein and the DGS molecule.
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And this technology can control real number precisely to two with a high yield of over 90% and a slight increase in aggregation of less than 1%.
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And also this process required only tangential flow filtration system for purification, not requiring current purification.
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And also this current process does not require redox chemistry.
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So we would like to emphasise the robustness and the simpleness of this technology.
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Next, we applied now conjugation technology to 15 gramme ADC production.
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So as you can see, the target ADC was obtained with a high yield of over 85% and a slight increasing in aggregation of less than 1%.
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And also we confirmed all impurities including AJICAP reagent and the payload linker were completely removed by TFF operation.
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And every analysis supported the high quality of the resulting conjugate.
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And also peptide mapping demonstrated the AJICAP conjugation site could complete to lysine 248
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So as shown here, AJICAP conjugation can provide high productivity and easiness of applying to manufacturing. AJICAP conjugation is a robust process which remained effective even when scaled up.
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So the action proceed with the same quality and yield as in small scale conjugation.
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To date, we have produced the 60 gramme of GMP material achieving the 90% of the target conjugate.
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Yeah, I'd like to conclude the capability of AJICAP conjugation.
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So regarding the antibody, now technology can be utilised on the most of human IgGs except for IgG 3 and another special reagent can work some months under that antibody.
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And also you can choose conjugation site from lysine 248 and the 288 and the both lysine 248 and 288.
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Recently we succeeded developed the site specific conjugation of Fab moiety and also you can choose DAR number from the 1,2,4,10, 12 and also payload ADCs can be obtained using this technology and you can choose any conjugation format like thiol, azide, DBCO chemistry.
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And you can introduce the any payload not only high potent small molecule but also protein, oligo, cargo molecule, even cell.
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And recently we successfully developed the DAR1 conjugation technology.
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So as I mentioned, the conventional AJICAP technology can introduce two attachments in both FC site specifically.
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While this innovative conjugate can introduce single attachment resulting in the asymmetric conjugate. Two affinity peptide in the reagent bind to the both FC sites and allowing for the DAR1 conjugation through the reaction with lysine and the reactive group that was bound to the peptide.
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So typically DAR1 conjugate requires DAR1 separation through column purification.
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However, this technology can achieve the same purpose by just adding chemical reagent.
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This innovative DAR1 conjugation technology can open up the new world of the new modality conjugate such as the oligoconjugate and the high potent DAR1 ADC, DDS molecule and the protein conjugate.
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In last chapter, I'm going to show you some application examples of this DAR1 conjugation technology.
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Let's move on to the next topic, AJICAP linker.
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Let me start by outlining of the potential issues with the traditional PAB linker that is commonly used as a cleavable linker.
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So this linker has two potential issues with Val-Cit-PAB linker.
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First is that it is susceptible to a wide variety of protons enzyme like cathepsin B or neutrophil elastase.
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And the second is it’s hydrophobicity of this linker.
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So to address these issues, we designed a new payload linker from here.
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Here is a very brief structure.
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So in these structures, hydrophilic peptide moiety and the cleavable peptide moiety are located outside.
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So this hydrophilic peptide moiety not only improves the hydrophilicity of the resulting conjugate, but also offer the protection from the various enzymatic attacks.
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On the other hand, this linker can be cleaved by the cathepsin B inside the cancer cell to release the payload properly.
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ADCs with this payload linkers show the promising stability result.
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So we conducted plasma stability studies using the site-specific ADCs with either Val-Cit linker or AJICAP linker.
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So as you can see in every plasma sources now AJICAP linker based ADCs show the promising stability compared to the other ADCs.
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And also please take a look at the light figure showing the HIC HPLC analysis.
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So a drastic retention time shift was observed.
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So this result suggested that now linker can improve the hydrophilicity of the resulting conjugate that contribute to reducing toxicity.
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By using the combination of AJICAP conjugation and the linker technologies, we can generate ADsS with the various DARs such as DAR 1,2,4,8,10.
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And also there are multiple DAR payload ADCs such as 1+1, 2+2, 8 + 2 can be generated by novel technologies.
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So next chapter I'm going to show some application of our technology to the ADC field.
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We applied our technology to ADCs using stochastic DAR4 ADCs generated through reduction with bar seat PAB linker and compared it with AJICAP site-specific DAR2 ADCs with AJICAP linker. AJICAP ADCs showed a better profile compared to stochastic DAR4 ADCs, despite AJICAP ADCs being DAR2 compared to stochastic DAR4 ADCs. The MTD of stochastic DAR4 ADCs was between 10 and 40 mg/kg, while the MTD of AJICAP ADCs was greater than 80 mg/kg, increasing twofold compared to stochastic DAR4 ADCs. This result can be explained by the LAT PK study result. AJICAP ADCs showed low toxicity and expanded the therapeutic window.
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We applied our technology to DAR4 ADCs by utilizing Lys248 and Lys288 modification. We successfully generated site-specific DAR4 ADCs with an average DAR number of 3.7, a slight increase in aggregation around 1%, and high quality of the resulting conjugate. AJICAP site-specific DAR4 ADCs showed promising stability in PK studies. Slight payload detachment was observed in stochastic DAR4 ADCs with AJICAP linker. AJICAP site-specific DAR4 ADCs showed a better profile compared to stochastic DAR4 ADCs even with the same payload linker.
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We applied our DAR4 technology to Exatecan-based ADCs. We compared Enhertu with AJICAP site-specific DAR4 Exatecan-based ADCs. AJICAP ADCs suppressed payload detachment, suggesting potential to reduce toxicity. AJICAP DAR4 ADCs showed comparable efficacy to Enhertu based on the amount of drug loaded.
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Final application is multiple payload ADCs in cancer chemotherapy. Heterogeneity of cancer and drug resistance are critical issues in treatment. ADCs comprised of multiple payloads with distinct MOAs are gathering interest. AJICAP technology can prepare two types of multiple payload ADCs: DAR2+2 generated through the combination of AJICAP Lys248 and Lys288 chemistries, and DAR8+2 ADCs generated through the combination of reduction and AJICAP conjugation. DAR8+2 ADCs showed a better profile compared to DAR8 ADCs.
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AJICAP technology can introduce biorthogonal groups to specific sites, allowing for precise incorporation of 1,2, 4 molecules. This technology can offer optimal intermediates tailored to modalities such as fluorescent conjugate, small molecule conjugate, DDS molecule, oligo, and protein. DAR1 conjugation format is optimal for the AOC field. AJICAP technology can produce DAR1 AOCs with high yield around 70%, while stochastic methods provide AOCs with low yield less than 40% due to the isolation. AJICAP DAR1 AOCs show promising results in every analysis, retaining antigen binding and promising stability in terms of physical properties.
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AJICAP DAR1 AOCs are very stable even after EV1 registration. Incubation results showed that AJICAP DAR1 AOCs retained their structure, making them suitable for AOC development. AJICAP DAR1 conjugation technology can make screening simple and show promising efficacy in T-cell engager models, PK studies, and tumour accumulation studies.
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Our business scheme involves non-confidential discussions to introduce our technologies. If clients are interested, we execute CDA contracts to discuss collaboration in detail. We then execute MTA with SOW, and clients provide antibodies and payloads to us. We generate conjugates and ship them back to clients for evaluation. If clients find drug candidates, we execute license agreements.
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In summary, AJICAP technology offers powerful tools for the development of new modality antibody conjugates. If you are interested in our technologies, please feel free to contact us anytime. Thank you for your time and interest.
