Calcium signaling profiles in patient-paired high-risk neuroblastoma tumors - novel mechanisms driving chemotherapy resistance and targeted therapies

Abstract

Neuroblastoma (NB) is a malignant pediatric tumor that accounts for over 15% of all pediatric cancer-related deaths. High-risk NB tumors occur in almost 50% of all patient cases and long-term outcomes for these children remain poor, with a survival rate of only ~40%, mainly due to disease relapse, which occurs frequently. Calcium has been demonstrated to play an important role in cancer cell proliferation, cell death, and metastasis, and alterations in Ca2+ signaling have been linked to tumor progression in many cancer types. However, the relationship between altered calcium signaling and the development of chemo-resistance in NB remains unknown. It was hypothesized that the acquisition of chemo-resistance in relapse NB is accompanied by calcium signaling profile alterations and that targeting these aberrations can reduce and/or reverse chemo-resistance. In this study, we demonstrate that relapse NB cells undergo significant remodeling of calcium signaling pathways, particularly store-operated calcium entry (SOCE), contributing to therapeutic escape. Through Specific Aim 1, we profiled calcium signaling in different patient pairs of pre-chemotherapy and post-relapse NB tumors and observed elevated SOCE activity in relapse NB cells, independent of MYCN amplification. This remodeling was mediated by altered expression of STIM1/STIM2, SERCA1/SERCA3, p-IP3R1, and PMCA, alongside reduced SARAF expression, particularly in the SK-N-Be2c line. Divergent patterns in STIM and SERCA isoform expression suggest complex regulatory adaptations that contribute to chemo-resistance in relapse NB. In Specific Aim 2, we leveraged these calcium profiles to identify therapeutic vulnerabilities unique to chemo-resistant relapse NB. Pharmacological modulation of SOCE using 2-APB re-sensitized relapse NB cells to chemotherapy, validating the therapeutic potential of targeting altered calcium signaling dynamics. Furthermore, we screened and identified potent anti-NB compounds, including extracts from Juniperus oblonga, synthetic molecules (compounds 248 and 249), and traditionally-prepared Native Hawaiian plant-based therapies. Intriguingly, traditional preparations demonstrated greater efficacy than Western methods, highlighting the potential of indigenous knowledge systems in novel drug discovery. In conclusion, calcium signaling remodeling, particularly SOCE elevation, is a key adaptive mechanism in NB chemo-resistance. In addition, our results validate calcium signaling profiling as a model for studying resistance mechanisms and therapeutic development in high-risk relapse NB, warranting further exploration and compound screening efforts focused on calcium modulation.