A. J. Lazenby1, A. P. Williams1, L. L. Stafman1, J. Aye1, V. R. Atigadda2, J. Stewart1, D. D. Muccio4, C. Grubbs3, E. A. Beierle1 2University Of Alabama at Birmingham,Dermatology,Birmingham, Alabama, USA 3University Of Alabama at Birmingham,Surgery,Birmingham, Alabama, USA 4University Of Alabama at Birmingham,Chemistry,Birmingham, Alabama, USA 5University Of Alabama at Birmingham,Pharmacology And Toxicology,Birmingham, Alabama, USA 6University Of Alabama at Birmingham,Pediatrics,Birmingham, Alabama, USA 1University Of Alabama at Birmingham,Pediatric Surgery,Birmingham, Alabama, USA
Introduction: Neuroblastoma (NB), a tumor derived from neural crest cells, is the most common extracranial solid tumor in children. 13-cis-retinoic acid (RA) is a differentiating agent currently utilized in therapy for high risk NB, but its use is limited by toxicities related to cholesterol and lipid metabolism. A synthetic rexinoid, 9-cis-UAB30 (UAB30), has been developed which has a favorable toxicity profile, demonstrating no significant toxicities in animal or human studies. Our lab has shown that UAB30 decreased NB cell proliferation and tumor growth in murine NB models. While UAB30 is promising, a reduction in drug dosage would be ideal to decrease the risk of potential side effects. Therefore, the aim of this project was to initiate pre-clinical evaluation of 9 new, synthetic rexinoid compounds (UAB111, UAB113, UAB114, UAB115, UAB116, UAB125, UAB126, 5-Me-UAB30 and 7-Me-UAB30) designed to have greater potency than that of UAB30 or RA. We hypothesized that these compounds would affect NB survival and differentiation in a manner comparable to RA and UAB30.
Methods: Four NB cell lines were utilized: 2 MYCN non-amplified (SK-N-AS, SH-EP) and 2 MYCN amplified [SK-N-BE(2), WAC2]. The effect of the rexinoids on cell viability and differentiation were evaluated using alamarBlue® assay and assessment of neurite outgrowth, respectively. Cells were treated for 72 hours at increasing concentrations ranging from 0 to 100 µM. Data were compared using Student’s t-test or ANOVA as appropriate and reported as mean ± SEM with p≤0.05 considered statistically significant.
Results: The 9 new rexinoids were tested against compounds previously studied, RA and UAB30, in all four NB cell lines. The SK-N-AS cell line was the most sensitive to the novel compounds, showing significantly decreased viability after treatment with 5 of them (UAB111, UAB113, UAB114, UAB116, 7-Me-UAB30) compared to RA or UAB30 (Figure, upper panel). The remaining cell lines, SK-N-BE(2), SH-EP and WAC2 demonstrated results similar to each other and had the most significant changes in viability with UAB116 and 7-Me-UAB30 (Figure, lower panel), so these 2 rexinoids were utilized for further studies. Neurite outgrowths are a measure of NB cell differentiation. Treatment with UAB116 or 7-Me-UAB30, resulted in a statistically significant increase in neurite outgrowths in both the SH-EP and SK-N-AS cell lines, indicating differentiation.
Conclusion: Treatment with these synthetic rexinoids, particularly UAB116 and 7-Me-UAB30, decreased NB cell viability significantly more than RA or UAB30. In addition, they led to NB cell differentiation. These data indicate that UAB116 and 7-Me-UAB30 should be further evaluated as potential novel therapeutics for NB.
