The actin cytoskeleton is an ideal chemotherapeutic target due to its role in numerous biological processes essential for tumor cell growth and survival. We developed a novel class of small molecule drugs that target actin filaments containing the tropomyosin Tpm3.1, upregulated in many cancers. Anti-tropomyosins (ATMs) are effective against neuroblastoma cell lines independent of NMYC status. Combination studies with ATMs show a high degree of synergy with anti-microtubule agents, both vinca alkaloids and taxanes, in neuroblastoma and other solid pediatric cell lines but not in untransformed cells. The combination of ATMs plus vincristine (VCR) resulted in a G2/M cell cycle arrest and apoptosis in neuroblastoma cell lines. More importantly, we found the synergy was maintained in vivo in a human neuroblastoma xenograft model. Animals treated with the combination showed a significant and profound regression of tumor growth compared to control animals including several with complete responses. The mechanism of synergy involves the ability of ATMs to sensitise neuroblastoma to the action of anti-microtubule drugs and results in multiple structural defects in the mitotic spindle. The outcomes of this study have significant therapeutic implications as microtubule inhibitors such as VCR are current standard of care for a wide range of solid tumors, including neuroblastoma. ATMs are currently being developed as adjunct therapy for the treatment of both adult and pediatric cancers.