With the approval of meanwhile five ADCs and more than 80 ADCs in clinical trials, the ADC landscape has developed rapidly during the last decade. However, as indicated also by the large number of ADCs which failed in the clinic, it remains challenging to achieve a sufficiently large therapeutic window in cancer patients. Furthermore, the identification of ADC payload classes with a novel mode of action would increase therapeutic options and potentially help to overcome resistance. Inhibitors of kinesin spindle protein (KSP/Eg5) have raised great interest because of their high antitumor potency which, however, could not be transferred into highly efficient clinical regimens due to dose limiting side effects such as neutropenia and mucositis.
H.-G. Lerchen et al. at Bayer AG investigated a novel pyrrole subclass of KSP inhibitors (KSPis) showing high potency in a broad panel of tumor cell lines for their utility as a novel payload class in ADCs. Different attachment sites for linkers have been identified in the KSPi molecule and were found compatible with cleavable and/or non-cleavable linkers. Highly potent ADCs against different targets, e.g. the tumor-associated antigens TWEAKR and B7H3 were obtained. 
Aiming at an improved therapeutic window, the group investigated a differential ADC activation in tumors versus healthy tissues by the tumor-associated protease legumain (LGMN) to introduce an additional safety level. A specific lysosomal ADC activation mediated by the asparaginyl endopeptidase LGMN was considered promising because of its broad tumor expression, its unique cleavage sequence and a higher LGMN activity in tumors versus healthy organs. 
Starting from highly potent KSPi ADCs with stable linker, a novel class of antibody-prodrug conjugates (APDCs) was designed, in which a key amino group in the KSPi molecule was blocked with a LGMN-cleavable peptide cap. To form active metabolites, in addition to the antibody degradation, this cap in the APDC has to be cleaved by LGMN.
The peptide sequence susceptible to LGMN cleavage has been optimized for further discrimination of the formation of active metabolites within tumor cells versus healthy tissues leveraging different tissue-specific legumain activities. Furthermore, the unique LGMN substrate sequence allowed for high cleavage specificity and discrimination versus other proteases such as elastase, which has been associated with an undesired extracellular cleavage of ADCs resulting in neutropenia. Preclinically, optimized APDCs with slow LGMN-mediated conversion reduced the levels of active metabolite in healthy organs while retaining high activity against different TWEAKR or B7H3 expressing mouse xenograft tumor models. 
Subsequently, the group investigated the utility of ADCs with LGMN-cleavable linkers for the treatment of hematological malignancies.  As an attractive target for the treatment of non-Hodgkin lymphoma the chemokine receptor CXCR5 was selected due to its high expression in all subtypes. IL3RA (CD123) was selected as a target for KSPi-ADCs to treat e.g. acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). Antibodies against both targets were developed and conjugated with KSPi. The optimization of both, LGMN-cleavable linker and KSPi payload to match the KSPi mode of action resulted in stable and highly potent ADCs targeting CXCR5 and IL3RA.
In conclusion, KSP inhibitors are a versatile new payload class for the generation of highly potent and selective ADCs against different targets and metabolism under legumain control is a versatile strategy to provide KSPi-ADCs and KSPi-APDCs with high potency, in vivo anti-tumor activity and an improved safety profile.
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 Lerchen HG, Stelte-Ludwig B, Sommer A, Berndt S, AS Rebstock, Johannes S, Marx L, Mahlert C, Greven S, Dietz L, Joerissen H, Weinmann H. KSPi-ADCs: ADCs with novel kinesin spindle protein inhibitor payloads and a tailor-made linker chemistry [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 228.
 Johannes S, Hammer S, Maersch S, Lerchen HG, Stelte-Ludwig B, Joerissen H, Von Ahsen O, Schatz C, Greven S, Mahlert C, Mumberg D, Lejeune P. Preclinical characterization of BAY-924, a first in class ADC targeting CXCR5-positive B-cell malignancies, with a KSP inhibitor as novel payload [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 4825.
 Sommer A, Kirchhoff D, Wengner AM, Stelte-Ludwig B, Lerchen HG, Rebstock AS, Von Ahsen O, Dietz L, Buchmann P, Johanssen S, Mumberg D, Kreft B. Anti-tumor activity of BAY-943, an anti-IL3RA ADC with a novel KSP inhibitor payload, in CDX and PDX AML models [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 4828.
How to cite:
By: Lerchen HG1, Stelte-Ludwig B1, Sommer A2, S. Berndt2, Kirchhoff D2, Lejeune P2, Rebstock AS3, Johannes S1, Wittrock S2, Marx L4, Mahlert C1, Greven S1, Dietz L1, Jörißen H1, Märsch S1, Zierz R2, Johanssen S2. Antibody-drug Conjugates with Novel Kinesin Spindle Protein Inhibitor Payloads and a Tailor-made Linker Chemistry – J. ADC. November 12, 2019. DOI: 10.14229/jadc.2019.11.12.002.
Affiliation: Bayer AG, Pharmaceuticals R&D
1 Wuppertal, Germany
2 Berlin, Germany
3 CropScience, Lyon, France
4 Debiopharm, Martigny, Switzerland
Last Editorial Review: November 11, 2019
- Original Manuscript Received: November 4, 2019
- Review results received: November 7, 2019
- Manuscript accepted for publication: November 11, 2019
- First Published Online: November 12, 2019.