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In vivo pilot experiment of NSG mice bearing MG-63 subcutaneous tumours treated intravenously with different concentrations of A5/158-vc-MMAE.

Examples of Endo180 positive and negative staining in subtypes of soft tissue sarcoma tumor cores.

Endo180 protein expression in sarcoma cell lines compared to epithelial cancer cells by immunofluorescence. Additionally, this figure shows the isotype control antibody conjugated to AlexaFluor 488 is not internalized into Endo180 expressing sarcoma cell lines and is not trafficked to the lysosome.

Immunohistochemical staining and quantification of the effect of A5/158, A5/158-vc-MMAE and Isotype-vc-MMAE treatment on the spontaneous metastasis of MG-63 cells to the liver in NSG mice.

Continuation of Figure S5 showing the effect of A5/158-vc-MMAE treatment on the spontaneous metastatic spread of MG-63 cells to the lungs and liver.

Antibodies and their dilutions

Endo180 protein and gene expression in healthy human tissue compared to sarcoma subtypes.

Composition and drug to antibody ratio of A5/158 and the isotype control antibody conjugated to vc-MMAE. This figure additionally shows that that conjugation of A5/158 and the isotype control antibody to vc-MMAE does not induce antibody aggregation or disrupt Endo180 protein recognition.

Although the 5-year survival rates for sarcoma patients have improved, the proportion of patients relapsing after first-line treatment remains high, and the survival of patients with metastatic disease is dismal. Moreover, the extensive molecular heterogeneity of the multiple different sarcoma subtypes poses a substantial challenge to developing more personalized treatment strategies. From the IHC staining of a large set of 625 human soft-tissue sarcomas, we demonstrate strong tumor cell staining of the Endo180 (MRC2) receptor in a high proportion of samples, findings echoed in gene-expression data sets showing a significantly increased expression in both soft-tissue and bone sarcomas compared with normal tissue. Endo180 is a constitutively recycling transmembrane receptor and therefore an ideal target for an antibody–drug conjugate (ADC). An anti-Endo180 monoclonal antibody conjugated to the antimitotic agent, MMAE via a cleavable linker, is rapidly internalized into target cells and trafficked to the lysosome for degradation, causing cell death specifically in Endo180-expressing sarcoma cell lines. In a sarcoma tumor xenograft model, the Endo180-vc-MMAE ADC, but not an isotype-vc-MMAE control or the unconjugated Endo180 antibody, drives on-target cytotoxicity resulting in tumor regression and a significant impairment of metastatic colonization of the lungs, liver and lymph nodes. These data, together with the lack of a phenotype in mice with an Mrc2 genetic deletion, provide preclinical proof-of-principle evidence for the future development of an Endo180-ADC as a therapeutic strategy in a broad range of sarcoma subtypes and, importantly, with potential impact both on the primary tumor and in metastatic disease.

A series of hemicyanine dyes was synthesized starting from the vinyl chloride-based cyanine dye IR-780. The dyes absorbed and emitted in the near-infrared region, while heavy atom (bromo and iodo) substitution promoted the generation of both singlet oxygen (1O2) as well as a range of other reactive oxygen-based species (ROS) upon irradiation at wavelengths greater than 610 nm. One hemicyanine dye displayed an outstanding singlet oxygen quantum efficiency (φΔ = 0.8) and was successfully applied in in vitro studies to mimic photodynamic therapy application.

Taking inspiration from the assembly of so-called peptoids (N-alkylglycine oligomers) we present a new synthetic methodology whereby N-heterocyclic carbene (NHC) based Pd ligands were assembled using a sub-monomer approach and loaded with Pd via solid-phase synthesis. This allowed the rapid generation a library of NHC–palladium catalysts that were readily functionalised to allow bioconjugation. These catalysts were able to rapidly activate a caged fluorophore and ‘switch-on’ an anticancer prodrug in 3D cell culture.