Fenretinide (4-HPR) Targets Caspase-9, ERK 1/2 and the Wnt3a/β-Catenin Pathway in Medulloblastoma Cells and Medulloblastoma Cell Spheroids

Abstract

Medulloblastoma (MB), a neuroectodermal tumor arising in the cerebellum, represents the most frequent childhood brain malignancy. Current treatments for MB combine radiation and chemotherapy and are often associated with relevant side effects; novel therapeutic strategies are urgently needed. N-(4-Hydroxyphenyl) retinamide (4-HPR, fenretinide), a synthetic analogue of all-trans retinoic acid, has emerged as a promising and well-tolerated cancer chemopreventive and chemotherapeutic agent for various neoplasms, from breast cancer to neuroblastoma. Here we investigated the effects of 4-HPR on MB cell lines and identified the mechanism of action for a potential use in therapy of MB. Flow cytometry analysis was performed to evaluate 4-HPR induction of apoptosis and oxygen reactive species (ROS) production, as well as cell cycle effects. Functional analysis to determine 4-HPR ability to interfere with MB cell migration and invasion were performed. Western Blot analysis were used to investigate the crucial molecules involved in selected signaling pathways associated with apoptosis (caspase-9 and PARP-1), cell survival (ERK 1/2) and tumor progression (Wnt3a and β-catenin). We show that 4-HPR induces caspase 9-dependent cell death in DAOY and ONS-76 cells, associated with increased ROS generation, suggesting that free radical intermediates might be directly involved. We observed 4-HPR induction of cell cycle arrest in G1/S phase, inactivated β-catenin, and inhibition of MB cell migration and invasion. We also evaluated the ability of 4-HPR to target MB cancer-stem/cancer-initiating cells, using an MB spheroids model, followed by flow cytometry and quantitative real-time PCR. 4-HPR treatment reduced DAOY and ONS-76 spheroid formation, in term of number and size. Decreased expression of the surface markers CD133+ and ABCG2+ as well as Oct-4 and Sox-2 gene expression were observed on BTICs treated with 4-HPR further reducing BITIC invasive activities. Finally, we analyzed 4-HPR ability to inhibit MB tumor cell growth in vivo in nude mice. Taken together, our data suggest that 4-HPR targets both parental and MB tumor stem/initiating cell-like populations. Since 4-HPR exerts low toxicity, it could represent a valid compound in the treatment of human MB.

UAB30, a novel RXR agonist, decreases tumorigenesis and leptomeningeal disease in group 3 medulloblastoma patient-derived xenografts

“…Cheepala and colleagues showed that ATRA decreased total ERK expression in skin cancer studies [51]. Fenretinide, another retinoid, was found to decrease ERK expression in ONS-76 MB cells [52]. In the current study, total ERK expression was diminished in all 3 MB PDX cell lines following RA or UAB30 treatment.…”

<p>Nanomicellar Lenalidomide–Fenretinide Combination Suppresses Tumor Growth in an <em>MYCN</em> Amplified Neuroblastoma Tumor</p>

“…Fenretinide is active in NB [7][8][9][10] and many other cancer types [11][12][13][14][15][16] by multiple mechanisms, [17][18][19][20][21][22] and it has shown efficacy against cancer stem cells. [23][24][25][26][27][28] Lenalidomide is another alternative antitumor agent currently used in the post-consolidation maintenance therapy of multiple myeloma and other hematological malignances. We selected lenalidomide to combine with fenretinide for its antiangiogenic properties and its acceptable toxicity profile.…”

<p>A Novel Nanomicellar Combination of Fenretinide and Lenalidomide Shows Marked Antitumor Activity in a Neuroblastoma Xenograft Model</p>

“…[5][6][7][8][9][10][11][12][13] Among the AABMs, fenretinide has a superior pharmacological profile due to its ability to induce cytotoxicity by multiple mechanisms [14][15][16][17][18] and to target cancer stem cell-associated molecular targets. [19][20][21][22] Previous clinical Phase I-III evaluations of fenretinide showed minimal systemic toxicity and good tolerability. 9,23,24 However, clinical trials aimed at evaluating the activity of fenretinide in cancer patients yielded disappointing results, likely due to the low bioavailability of the initial oral capsule formulation.…”