abt-199 and Neuroblastoma

The prognosis of relapsed/refractory (R/R) neuroblastoma (NB) is dismal, calling for new therapeutic strategies. Venetoclax (VEN) is a highly selective, potent, orally bioavailable, BCL-2 inhibitor small-molecule that showed a synergistic effect with cyclophosphamide and topotecan (Cy-Topo) in murine NB models. Our aim was to evaluate the feasibility of VEN plus Cy-Topo in children with R/R NB. Four patients, who had previously failed > 3 lines of treatment, were treated with VEN plus Cy-Topo based on a 28-day schedule in an outpatient setting. BCL-2 expression in immunochemistry on tumor samples at relapse and the BCL2 gene status was evaluated in all patients. The main toxicity was hematological, with grade 4 neutropenia and thrombocytopenia occurring in all courses and leading to transient VEN discontinuation. Grade 3 oral mucositis was observed in 1/8 courses. No other grade 2-4 toxicities were observed. BCL-2 was expressed in all tumors, while no molecular abnormalities in the BCL-2 genes were detected. A stable disease was observed in all patients, without any progression during the study period. VEN plus Cy-Topo is well tolerated, with encouraging results that may be improved by testing the schedule in less advanced patients.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Bridged Bicyclo Compounds, Heterocyclic; Child; Chronic Disease; Cyclophosphamide; Humans; Mice; Neoplasm Recurrence, Local; Neoplasms, Second Primary; Neuroblastoma; Proto-Oncogene Proteins c-bcl-2; Topotecan

Venetoclax is a small molecule inhibitor of the prosurvival protein BCL-2 that has gained market approval in BCL-2-dependent hematologic cancers including chronic lymphocytic leukemia and acute myeloid leukemia. Neuroblastoma is a heterogenous pediatric cancer with a 5-year survival rate of less than 50% for high-risk patients, which includes nearly all cases with amplified

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Bridged Bicyclo Compounds, Heterocyclic; Cell Proliferation; Cyclophosphamide; Female; Gene Amplification; Gene Expression Regulation, Neoplastic; Humans; Mice; Mice, Inbred NOD; Mice, SCID; N-Myc Proto-Oncogene Protein; Neuroblastoma; Sulfonamides; Topotecan; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Despite advances in the treatment of neuroblastoma, patients with high-risk disease still have dismal survival prognosis. Neuroblastoma cells display elevated expression of the antiapoptotic BCL-2 proteins, suggesting that BH3-mimetics may be a promising treatment option. Here, we investigated the role of BCL-2, BCL-X. A panel of neuroblastoma cell lines and primary patient-derived cells were exposed to BH3-mimetics targeting BCL-2 (ABT-199), BCL-X. All tested BH3-mimetics were able to induce apoptosis in neuroblastoma cell lines, indicating that not only BCL-2 but also BCL-X. By using selective BH3-mimetics, this study demonstrates that BCL-2, BCL-X

Topics: Apoptosis; bcl-2 Homologous Antagonist-Killer Protein; Bcl-2-Like Protein 11; bcl-X Protein; Biomimetics; Bridged Bicyclo Compounds, Heterocyclic; Cell Line, Tumor; Cell Proliferation; Gene Expression Regulation, Neoplastic; Humans; Myeloid Cell Leukemia Sequence 1 Protein; Neuroblastoma; Peptide Fragments; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Pyrimidines; Sulfonamides; Thiophenes

Recurrent high-risk neuroblastoma is a childhood cancer that often fails to respond to therapy. Fenretinide (4-HPR) is a cytotoxic retinoid with clinical activity in recurrent neuroblastoma and venetoclax (ABT-199) is a selective inhibitor of the antiapoptotic protein B-cell lymphoma-2 (BCL-2). We evaluated activity of 4-HPR + ABT-199 in preclinical models of neuroblastoma. Patient-derived cell lines and xenografts from progressive neuroblastoma were tested. Cytotoxicity was evaluated by DIMSCAN, apoptosis by flow cytometry, and gene expression by RNA sequencing, quantitative RT-PCR, and immunoblotting. 4-HPR + ABT-199 was highly synergistic against high BCL-2-expressing neuroblastoma cell lines and significantly improved event-free survival of mice carrying high BCL-2-expressing patient-derived xenografts (PDX). In 10 matched-pair cell lines [established at diagnosis (DX) and progressive disease (PD) from the same patients], BCL-2 expression in the DX and PD lines was comparable, suggesting that BCL-2 expression at diagnosis may provide a biomarker for neuroblastomas likely to respond to 4-HPR + ABT-199. In a pair of DX (COG-N-603x) and PD (COG-N-623x) PDXs established from the same patient, COG-N-623x was less responsive to cyclophosphamide + topotecan than COG-N-603x, but both DX and PD PDXs were responsive to 4-HPR + ABT-199. Synergy of 4-HPR + ABT-199 was mediated by induction of NOXA via 4-HPR stimulation of reactive oxygen species that induced expression of ATF4 and ATF3, transcription factors for NOXA. Thus, fenretinide + venetoclax is a synergistic combination that warrants clinical testing in high BCL-2-expressing neuroblastoma.

Topics: Animals; Antineoplastic Agents; Bridged Bicyclo Compounds, Heterocyclic; Cell Culture Techniques; Cell Line, Tumor; Disease Models, Animal; Female; Fenretinide; Humans; Mice; Neuroblastoma; Proto-Oncogene Proteins c-bcl-2; Sulfonamides

High-risk neuroblastoma is often distinguished by amplification of

Topics: Animals; Axons; Benzazepines; Bridged Bicyclo Compounds, Heterocyclic; Cell Line, Tumor; Cell Proliferation; Demethylation; Drug Resistance, Neoplasm; Female; Gene Expression Regulation, Neoplastic; Histone Demethylases; Histones; Humans; Lysine; Mice, Nude; Neuroblastoma; Pyrimidines; Risk Factors; Sulfonamides; Tretinoin

Pediatric patients with high-risk neuroblastoma (HR NB) often fail to respond to upfront intensive multimodal therapy. Tumor-acquired suppression of apoptosis contributes to therapy resistance. Many HR NB tumors depend on the anti-apoptotic protein Bcl-2 for survival, through Bcl-2 sequestration and inhibition of the pro-apoptotic protein, Bim. Bcl-2 dependent xenografts derived from aggressive human NB tumors are cured with a combination of cyclophosphamide and ABT-737, a Bcl-2/Bcl-XL/Bcl-w small molecule antagonist. The oral analogue to ABT-737, Navitoclax (ABT-263), clinically causes an immediate drop in peripheral platelet counts as mature platelets depend on Bcl-xL for survival. This led to the creation of a Bcl-2 selective inhibitor, ABT-199 (Venetoclax). A Phase I trial of ABT-199 in CLL showed remarkable antitumor activity and stable patient platelet counts. Given Bcl-XL does not play a role in HR NB survival, we hypothesized that ABT-199 would be equally potent against HR NB.. Cytotoxicity and apoptosis were measured in human derived NB cell lines exposed to ABT-199 combinations. Co-Immunoprecipitation evaluated Bim displacement from Bcl-2, following ABT-199. Murine xenografts of NB cell lines were grown and then exposed to a 14-day course of ABT-199 alone and with cyclophosphamide.. Bcl-2 dependent NB cell lines are exquisitely sensitive to ABT-199 (IC50 1.5-5 nM) in vitro, where Mcl-1 dependent NBs are completely resistant. Treatment with ABT-199 displaces Bim from Bcl-2 in NB to activate caspase 3, confirming the restoration of mitochondrial apoptosis. Murine xenografts of Mcl-1 and Bcl-2 dependent NBs were treated with a two-week course of ABT-199, cyclophosphamide, or ABT-199/cyclophosphamide combination. Mcl-1 dependent tumors did not respond to ABT-199 alone and showed no significant difference in time to tumor progression between chemotherapy alone or ABT-199/cyclophosphamide combination. In contrast, Bcl-2 dependent xenografts responded to ABT-199 alone and had sustained complete remission (CR) to the ABT-199/cyclophosphamide combination, with one recurrent tumor maintaining Bcl-2 dependence and obtaining a second CR after a second course of therapy.. HR NB patients are often thrombocytopenic at relapse, raising concerns for therapies like ABT-263 despite its HR NB tumor targeting potential. Our data confirms that Bcl-2 selective inhibitors like ABT-199 are equally potent in HR NB in vitro and in vivo and given their lack of platelet toxicity, should be translated into the clinic for HR NB.

Topics: Aniline Compounds; Apoptosis; Apoptosis Regulatory Proteins; Bcl-2-Like Protein 11; bcl-X Protein; Bridged Bicyclo Compounds, Heterocyclic; Caspase 3; Cell Line, Tumor; Cyclophosphamide; Drug Resistance, Neoplasm; Humans; Membrane Proteins; Neuroblastoma; Platelet Count; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Risk Assessment; Sulfonamides; Xenograft Model Antitumor Assays

The anti-apoptotic protein B cell lymphoma/leukaemia 2 (BCL-2) is highly expressed in neuroblastoma and plays an important role in oncogenesis. In this study, the selective BCL-2 inhibitor ABT199 was tested in a panel of neuroblastoma cell lines with diverse expression levels of BCL-2 and other BCL-2 family proteins. ABT199 caused apoptosis more potently in neuroblastoma cell lines expressing high BCL-2 and BIM/BCL-2 complex levels than low expressing cell lines. Effects on cell viability correlated with effects on BIM displacement from BCL-2 and cytochrome c release from the mitochondria. ABT199 treatment of mice with neuroblastoma tumors expressing high BCL-2 levels only resulted in growth inhibition, despite maximum BIM displacement from BCL-2 and the induction of a strong apoptotic response. We showed that neuroblastoma cells might survive ABT199 treatment due to its acute upregulation of the anti-apoptotic BCL-2 family protein myeloid cell leukaemia sequence 1 (MCL-1) and BIM sequestration by MCL-1. In vitro inhibition of MCL-1 sensitized neuroblastoma cell lines to ABT199, confirming the pivotal role of MCL-1 in ABT199 resistance. Our findings suggest that neuroblastoma patients with high BCL-2 and BIM/BCL-2 complex levels might benefit from combination treatment with ABT199 and compounds that inhibit MCL-1 expression.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Bcl-2-Like Protein 11; Bridged Bicyclo Compounds, Heterocyclic; Cell Line, Tumor; Cell Survival; Cytochromes c; Drug Evaluation, Preclinical; Female; Humans; Inhibitory Concentration 50; Mice; Mitochondria; Myeloid Cell Leukemia Sequence 1 Protein; Neuroblastoma; Proto-Oncogene Proteins c-bcl-2; Sulfonamides; Treatment Outcome; Xenograft Model Antitumor Assays

MYCN amplification in human cancers predicts poor prognosis and resistance to therapy. However, pharmacological strategies that directly target N-Myc, the protein encoded by MYCN, remain elusive. Here, we identify a molecular mechanism responsible for reciprocal activation between Polo-like kinase-1 (PLK1) and N-Myc. PLK1 specifically binds to the SCF

Topics: Animals; Antineoplastic Agents; Brain Neoplasms; Bridged Bicyclo Compounds, Heterocyclic; Cell Cycle Proteins; Cell Line, Tumor; Drug Synergism; F-Box Proteins; F-Box-WD Repeat-Containing Protein 7; Feedback, Physiological; Gene Expression Regulation, Neoplastic; Humans; Mice, Nude; N-Myc Proto-Oncogene Protein; Neuroblastoma; Neurons; Polo-Like Kinase 1; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Pteridines; RNA, Small Interfering; Signal Transduction; Sulfonamides; Survival Analysis; Transcription, Genetic; Tumor Burden; Ubiquitin-Protein Ligases; Xenograft Model Antitumor Assays