Ruthenium is popular as a metal-core for chemotherapeutics, due to versatile molecular coordination. Because new metallodrugs are synthesized at high rates, our studies included assays in zebrafish to expedite the initial evaluation as anti-cancer agents. Here we evaluated novel metallodrugs PMC79 and LCR134), and cisplatin, a widely-used platinum-based chemotherapeutic. We hypothesized that this model could characterize anti-cancer properties and recapitulate previous in vitro results in vivo. Our findings suggest anti-cancer properties of PMC79 and LCR134 were similar with less toxicity than cisplatin. Exposures from 24-72 hrs at or below the LOAELs of PMC79 and LCR134 (3.9 µM and 13.5 µm, respectively), impaired blood vessel development and tailfin regeneration. Blood vessel examination through live-imaging of larvae revealed distinct regional anti-angiogenic impacts. The significant decrease in gene expression of the VEGF-HIF pathway and beta-actin could explain the morphological effects observed in the whole organism following exposure. Tailfin amputation in larvae exposed to PMC79 or LCR134 inhibited tissue regrowth and cell division, but did not impact normal cell proliferation unlike cisplatin. This suggests Ru-drugs may be more selective in targeting cancerous cells than cisplatin. Additionally, in vitro mechanisms were confirmed. PMC79 disrupted cytoskeleton formation in larvae and P-glycoprotein transporters in vivo was inhibited at low doses which could limit off-target effects of chemotherapeutics. Our results demonstrate the value for using the zebrafish in metallodrug research to evaluate mechanisms and off-target effects. In light of the findings reported in this paper, future investigation of PMC79 and LCR134 are warranted in higher vertebrate models.

Biallelic mutations in Protein O-mannosyltransferase 1 (POMT1) are among the most common causes of a severe group of congenital muscular dystrophies (CMDs) known as dystroglycanopathies. POMT1 is a glycosyltransferase responsible for the attachment of a functional glycan mediating interactions between the transmembrane glycoprotein dystroglycan and its binding partners in the extracellular matrix (ECM). Disruptions in these cell-ECM interactions lead to multiple developmental defects causing brain and eye malformations in addition to CMD. RemovingPomt1in the mouse leads to early embryonic death due to the essential role of dystroglycan in embryo implantation in rodents. Here, we characterized and validated a model ofpomt1loss of function in the zebrafish showing that developmental defects found in individuals affected by dystroglycanopathies can be recapitulated in the fish. We also discovered thatpomt1mRNA provided by the mother in the oocyte supports dystroglycan glycosylation during the first few weeks of development. Muscle disease, retinal synapse formation deficits, and axon guidance defects can only be uncovered during the first week post-fertilization by generating knock-out embryos from knock-out mothers. Conversely, maternalpomt1from heterozygous mothers was sufficient to sustain muscle, eye, and brain development only leading to detectable muscle disease and loss of photoreceptor synapses at 30 days post fertilization. Our findings show that it is important to define the contribution of maternal mRNA while developing zebrafish models of dystroglycanopathies and that offspring generated from heterozygous and knock-out mothers can be used to differentiate the role of dystroglycan glycosylation in tissue formation and maintenance.

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