Germany-based ProBioGen, a leading specialist for contract development and manufacturing of complex glycoproteins, and Eucodis Bioscience, an expert in enzyme engineering based in Vienna, Austria, jointly announced the signature of an exclusive license agreement involving C-LiNK (CTAT), an innovative, site-specific ADC conjugation technology.

C-LiNK (CTAT™) is an advantageous conjugation system which covalently links, by a specific 1-step enzymatic reaction, toxins, drugs, or other substances to the C-terminus of an antibody or a recombinant protein (see Figure 1.0). The C-LiNK enzyme targets a short recognition sequence of 3-4 amino acids fused to the C-terminus of the to-be-conjugated protein. It then cleaves this specific sequence and covalently links the drug molecule to the antibody by a transamidation reaction.

Figure 1.0: C-LiNK (CTAT™) enables site-specific, controllable and stable conjugation through a single enzymatic reaction under mild conditions with highest efficacy. This allows cost- effective and rapid production of ADCs from small scale screening conditions to large scale GMP production. Source: ENCODES Bioscience; Poster presented at PEGS 2014 [1]. Courtesy: © EUCODIS Bioscience. Used with permission.
This directed conjugation approach is not interfering with the antibody’s antigen binding site, does not alter its integrity, or its manufacturability.

The technology allows adjusting a defined Drug-to-Antibody Ratio (DAR) and, by modifying the linker, attaching more than one drug per site, or inserting a specific cleavage site, if needed.

Advertisement #3 
Figure 2.0: Tumor Volume Kinetics Overview
Figure 2.0: Tumor Volume Kinetics – Overview. Courtesy: © EUCODIS Bioscience. Used with permission.

Superior Efficacy in Xenograft Trial
In earlier trials scientists looked at an anti-Her2 antibody conjugated to the maytansinoid DM1 using the novel CTAT-linker technology (“CTAT-ADC”).

The CTAR-ADC was tested in nude mice with SKOV-3 cancer cell xenografts for maximum tolerated dose and efficacy. Three different dosages of the CTAT-ADC were compared to a commercially available Trastuzumab-DM1-ADC (T-DM1, Kadcyla®; Genentech/Roche), to naked antibody and to the delivery vehicle.

The results showed that CTAT-ADC was well tolerated by the animals in a dosage of up to 70 mg/kg body weight. No animals were lost, and only in the highest dosage group a slight and temporary loss of body weight was observed.

In the xenograft trial, tumor volume kinetics was monitored after two applications of the CTAT-ADC and the control substances. The first dose was applied at study start and a second dose after recurrence of substantial tumor growth in the individual study groups.

During a period of 29 days, the CTAT-ADC at 35mg/kg dosage was the best performing regimen and the only group which showed sustained tumor reduction (see Figure 2.0).

Figure 3.0 Tumor Volume Kinetics CTAT-ADC 35mg
Figure 3.0: Tumor Volume Kinetics – CTAT-ADC 35 mg/kg. Courtesy: © EUCODIS Bioscience. Used with permission.

This group of test animals also showed very homogeneous results compared to the group treated with T-DM1-ADC (see Figure 3.0 and Figure 4.0). The commercial T-DM1-ADC at 10mg/kg and the CTAT-ADC at 70 mg/kg dosage showed good suppression of tumor growth during the initial phase of the treatment but towards the end of the 29 days tumor growth was observed again. The third CTAT-ADC dosage group (10 mg/kg) achieved an initial tumor growth inhibition comparable to T-DM1-ADC, but also in this group the tumor resumed growth closer to the end of the observation period.

In-vitro data of the CTAT-technology were presented earlier. The CTAT-ADC used in the presented xenograft trial, and additional ADCs based on different antibodies and on a Fab- fragment showed efficacy against a variety of cancer cell lines. All of these constructs were generated using the CTAT linker technology for conjugation. The data confirmed a plasma stability and a well-defined drug-antibody-ratio (DAR) of 1 and 2.

Figure 4:0 Tumor Volume Kinetics T-DM1-ADC 10mg
Figure 4.0: Tumor Volume Kinetics – T-DM1-ADC 10 mg/kg. Courtesy: © EUCODIS Bioscience. Used with permission.

“This agreement is an excellent match between Eucodis’ enzyme engineering expertise and ProBioGen’s therapeutic protein- optimizing technologies and we look much forward to position C-LiNK and our contract manufacturing services right in the middle of the hot ADC field,” noted ProBioGen’s CEO, Wieland Wolf, Ph.D.

“We see a huge potential for C-LiNK in Biopharma, especially in [the development of Antibody-drug Conjugates], but our expertise is engineering enzymes,” noted Karl Hübler, Ph.D, Chief Executive Officer of Eucodis Bioscience. “…by teaming up with ProBioGen, our ADC conjugation technology will certainly get a much wider and quicker access to the community of therapeutic protein developing companies and we are convinced that our agreement with ProBioGen will result in fruitful collaborations within the industry,” Hübler continued.

Figure 5.0: CTAT-ADC in-vitro data: cell proliferation test in SKBR-3 cell lines. Courtesy: © EUCODIS Bioscience. Used with permission.

“This innovative ADC technology, which efficiently attaches any payload enzymatically via a pure peptide linker – as short as two amino acids -, blends perfectly into our portfolio of robust cell line/product engineering technologies and manufacturing capabilities. The 1-step C-LiNK coupling technology will further open up the development space for ADCs and represents a competitive alternative to other Antibody Drug Conjugate Technologies,” added Volker Sandig, Ph.D, Chief Scientific Officer at ProBioGen.

“Based on a trypsin variant engineered to enforce formation of a new peptide bond, our ADC conjugation technology links toxins or other molecules to the C-terminus of either antibody chain, without disturbing the native antibody structure. The outcome is a defined Drug Antibody Ratio at high yield with low enzyme requirements and it offers on top the flexibility to add different payloads to separate sites in future ADCs,” said Jan Modregger, Ph.D, Head of Research and Development at Eucodis.

Under the agreement ProBioGen gains exclusive rights to commercialize Eucodis’ enzymatic C-terminal linking technology, and to offer it, together with its antibody development services, royalty-free to ADC-developing parties.

Advertisement #4