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Antibody–drug conjugates (ADCs) that incorporate the exatecan derivative DXd in their payload are showing promising clinical results in solid tumor indications. The payload has an F-ring that also contains a second chiral center, both of which complicate its synthesis and derivatization. Here we report on new camptothecin-ADCs that do not have an F-ring in their payloads yet behave similarly to DXd-bearing conjugates in vitro and in vivo. This simplification allows easier derivatization of camptothecin A and B rings for structure–activity relationship studies and payload optimization. ADCs having different degrees of bystander killing and the ability to release hydroxyl or thiol-bearing metabolites following peptide linker cleavage were investigated.

Dysregulation of ADAM9, a member of the ADAM (a disintegrin and metalloproteinase) family of proteases, has been implicated in tumor progression and metastasis, as well as pathological neovascularization. ADAM9 overexpression correlates with poor prognosis in multiple cancers. We have shown that ADAM9 is overexpressed in multiple solid tumor indications and that anti-ADAM9 antibodies are efficiently internalized and degraded by tumor cell lines making ADAM9 an attractive target for antibody-drug conjugate (ADC) development. Here, we describe IMGC936, the first ADAM9-targeting ADC to enter preclinical development. IMGC936 is comprised of a high-affinity humanized antibody site-specifically conjugated to DM21, a next-generation linker-payload that combines a maytansinoid microtubule-disrupting payload with a stable peptide linker at a drug-antibody ratio of two. To maximize the potential for IMGC936 activity, the M252Y/S254T/T256E (YTE) mutation was introduced into the CH2 domain of the antibody to increase in vivo plasma half-life and exposure. In vitro studies demonstrated targeted cytotoxicity of IMGC936 across a panel of ADAM9-positve tumor cell lines with activity at least 2 logs greater than a non-targeting conjugate. Consistent with the in vitroactivity, an anti-ADAM9-DM21 conjugate displayed compelling anti-tumor activity in multiple xenograft models representing non-small cell lung, gastric and colorectal cancers. For example, in the EBC-1 non-small cell lung cancer subcutaneous xenograft model with only moderate ADAM9 expression (H-score of 130), anti-ADAM9-DM21 not only induced tumor growth delay but produced complete and durable remissions in 6/6 mice following a single intravenous dose of 8.6 mg Ab/kg (100 ug DM21/kg). IMGC936 demonstrated a favorable pharmacokinetic profile with good conjugate stability in non-human primates. Importantly, IMGC936 was well-tolerated following repeat dosing in cynomolgus monkeys with no ADAM9 target-related toxicities identified at doses exceeding the levels required for anti-tumor activity in murine xenograft models. Based on the totality of the preclinical data, IMGC936 represents a promising therapeutic candidate to target a wide range of ADAM9-expressing tumors. Citation Format: Stuart Hicks, Deryk Loo, Kerstin Sinkevicius, Juniper Scribner, Bhaswati Barat, Nicholas Yoder, Christopher Espelin, Marian Themeles, Francine Chen, Jacquelynn Lucas, Jennifer Brown, Bahar Matin, Megan Fuller, Jenny Lee, Paulin Salomon, Juliet Costoplus, Sadiqa Yancey, Gundo Diedrich, Sergey Gorlatov, Thomas Son, Michael Chiechi, Pam Li, Michael Spliedt, Valentina Ciccarone, Jeff Hooley, Nadia Gantt, James Tamura, Kerry Donahue, Paul Moore, Syd Johnson, Thomas Chittenden, Richard Gregory, Ezio Bonvini. IMGC936, a first-in-class ADAM9-targeting antibody-drug conjugate, demonstrates promising anti-tumor activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1533.

DARPin® molecules are small engineered proteins, derived from natural ankyrin repeat proteins that are selected to bind to specific targets with high affinity. Individual DARPin® molecules can be linked together genetically in order to create multi-specific drug molecules. This versatility of DARPin® molecules makes them an attractive alternative to antibodies for the development of drug conjugates. We have developed DARPin® drug conjugates (DDCs) using a model EGFR multi-specific DARPin® molecule, consisting of four DARPin® domains linked together. DARPin® constructs were conjugated to the indolinobenzodiazepine mono-imine DGN549, a potent DNA alkylating payload. Biophysical characterization showed the DDCs to be well behaved in stability and solubility assays. The in vitro binding and cytotoxicity of these DDCs were evaluated. The DARPin® conjugates displayed in vitro potency in direct cytotoxicity assays across a panel of cell lines expressing EGFR. The impact of conjugation on PK parameters and in vivo efficacy is currently under evaluation. DARPin® drug conjugates combine the potency observed with antibody drug conjugates and the modular design of DARPin® molecules to create designer therapeutics. Citation Format: Laura A. Laviolette, Cynthia J. Guidi, Christian Reichen, Qifeng Qiu, Luke Harris, Patricia Schildknecht, Stefanie Fischer, Zita Arany, Tanja Hospodarsch, Anna Skaletskaya, Megan Fuller, Stephen Abbott, Rebecca McCarthy, Jenny Lee, Katherine Francisco, Kerstin Sinkevicius, Sharlene Adams, Christopher Espelin, Emily Reid, Wei Li, Carla Marashio, Kerry Donahue, Stuart Hicks, Dan Snell. Generation of site-specific DARPin® drug conjugates using EGFR as a model system [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 215.