Abstract 376: Sonoporated size selected microbubblesand liposomal doxorubicinadditively induce apoptosis in neuroblastoma xenografts
Authors: Hernandez Sonia L. , Sundland Rachael , Ballan Donia , Bellary Aditi , Kandel Jessica J. , Feshitan Jameel , Sirsi Shashank 
Published: June 2022

Background: Neuroblastoma (NB) is the second most common malignancy diagnosed in infants, accounting for 15% of pediatric tumor deaths. Half of children with NB receive an intensive regimen including high-dose chemotherapy with 50% survival, resulting in acute and long term toxicities. One of the challenges of chemotherapy is irregular tumor vasculature. Thus, increasing targeted drug delivery without increasing drug dosage can result in enhanced drug efficacy and improved patient outcomes. We have shown that sonoporation (focused ultrasound-guided gas-filled microbubbles) increases high dose liposomal doxorubicin (L-DOX) uptake in NB xenografts by increasing tumor perfusion. However, these studies used polydisperse microbubbles (PMB), which were developed for imaging purposes. We hypothesized that MB size restriction would control their response to ultrasound pressure, yielding a higher L-DOX payload despite using using lower L-DOX dosages.

Methods: Nude mice received 1x10^6 NGP cells (NB cells) intrarenally. When tumors reached 1 gram, NB xenografts received an intravenous polydisperse (PMB) or 4-5uM (SIMB) microbubble infusion with or without 1mg/kg liposomal doxorubicin (L-DOX) under focused ultrasound. Tumors were measured over 7 days with calipers, others sacrificed 24 hours after treatment for histology and immunohistochemistry. We assessed endomucin and isolectin-B4 (endothelium), Zona occludens-1 (ZO-1) (tight junction), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL, apoptosis).

Results: Tumors receiving low dose L-DOX alone or PMB sonoporation with L-DOX were not different from untreated controls after 7 days. SIMB alone resulted in a slower tumor growth than control tumors (20 vs 100% p&lt;0.05); L-DOX coupled with SIMB resulted in further tumor growth restriction 7 days after treatment (0 vs 100% p&lt;0.05). SIMB increased tumor apoptosis (TUNEL staining) in the absence of L-DOX compared to controls (7 vs 58% of area p=0.003) as well as in the presence of L-DOX (7 vs 78% of area p&lt;0.001). PMBs did not change TUNEL levels regardless of L-DOX. Tumor vascular lumens (widest axis within the endothelial marker endomucin) confirmed PMB duplicates lumen diameter compared to controls (p&lt;0.05), and revealed SIMB triplicates lumen diameter (p&lt;0.01) regardless of L-DOX. SIMB resulted in loss of tight junction protein ZO-1 both in vasculature and tumor cells and widespread L-DOX uptake. Together, our data shows SIMB sonoporation increases tumor blood volume and vascular permeability leading to higher chemotherapy uptake and apoptosis.

Conclusions: Together, our data shows SIMB sonoporation enables chemotherapy uptake in poorly perfused NB xenografts by increasing perfusion and permeability, potentiating apoptotic effects. SIMB sonoporation could reduce acute and long term toxicities.

Citation Format: Sonia L. Hernandez, Rachael Sundland, Donia Ballan, Aditi Bellary, Jessica J. Kandel, Jameel Feshitan, Shashank Sirsi. Sonoporated size selected microbubblesand liposomal doxorubicinadditively induce apoptosis in neuroblastoma xenografts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 376.