Anti-Acetyl-α-Tubulin monoclonal antibody, clone 7-22C-2

Background Osteosarcoma (OS), the most common bone tumor in children and adolescents, has high rates of metastasis leading to poor survival. Leucine-rich repeat containing 15 (LRRC15), a transmembrane protein whose expression is modulated by TGF beta, was recently shown to be highly expressed on the surface of OS tumor cells. Here, we evaluate a novel antibody-drug conjugate (ADC) targeting LRRC15 in OS human cell lines and murine xenografts. We compare this new ADC, which is conjugated to the anthracycline derivative PNU-159682 (PNU), to a previously studied LRRC15 ADC that is conjugated to the tubulin inhibitor monomethyl auristatin E (MMAE), since anthracyclines are standard of care in OS. Procedure We evaluated LRRC15 expression in OS cells using Western blots and flow cytometry, and analyzed the epigenetic landscape of the LRRC15 locus using chromatin immunoprecipitation. Efficacy of ADCs on cell growth was analyzed by IncuCyte live cell imaging. Intramuscular xenograft tumor growth was assessed by bioluminescence imaging and hematoxylin and eosin staining. Results LRRC15-PNU is more effective at inhibiting growthin vitroandin vivothan an isotype antibody control or the LRRC15-MMAE ADC in two high LRRC15 expressing OS cell lines. Low expressing cell lines are not sensitive to either ADC. Importantly, cells with low LRRC15 expression are amenable to re-expression after TGF beta treatment, suggesting a potential to sensitize insensitive OS cells to LRRC15 ADC treatment.In vivo, LRRC15-PNU had cure rates of 40-100% in OS xenograft models. Conclusions Overall, LRRC15-directed ADCs are a promising new avenue for OS treatment.

The shape of kinetoplastids, such as Trypanosoma brucei, is precisely defined during the stages of the life cycle and governed by a stable subpellicular microtubule cytoskeleton. During the cell cycle and transitions between life cycle stages, this stability has to transiently give way to a dynamic behaviour to enable cell division and morphological rearrangements. How these opposing requirements of the cytoskeleton are regulated is poorly understood. Two possible levels of regulation are activities of cytoskeleton-associated proteins and microtubule post-translational modifications (PTMs). Here, we investigate the functions of two putative tubulin polyglutamylases in T. brucei, TTLL6A and TTLL12B. Depletion of both proteins leads to a reduction in tubulin polyglutamylation in situ and is associated with disintegration of the posterior cell pole, loss of the microtubule plus-end-binding protein EB1 and alterations of microtubule dynamics. We also observe a reduced polyglutamylation of the flagellar axoneme. Quantitative motility analysis reveals that the PTM imbalance correlates with a transition from directional to diffusive cell movement. These data show that microtubule polyglutamylation has an important role in regulating cytoskeletal architecture and motility in the parasite T. brucei. This article has an associated First Person interview with the first author of the paper.