Galardy, Paul J.

Abstract The mechanistic target of rapamycin (mTOR) is a central regulator of cellular proliferation and metabolism. Depending on its binding partners, mTOR is at the core of 2 complexes that either promote protein biosynthesis (mTOR complex 1; mTORC1) or provide survival and proliferation signals (mTORC2). Protein biosynthesis downstream of mTORC1 plays an important role in MYC-driven oncogenesis with translation inhibitors garnering increasing therapeutic attention. The germinal center B-cell oncogene UCHL1 encodes a deubiquitinating enzyme that regulates the balance between mTOR complexes by disrupting mTORC1 and promoting mTORC2 assembly. While supporting mTORC2-dependent growth and survival signals may contribute to its role in cancer, the suppression of mTORC1 activity is enigmatic, as its phosphorylation of its substrate 4EBP1 promotes protein biosynthesis. To address this, we used proximity-based proteomics to identify molecular complexes with which UCH-L1 associates in malignant B cells. We identified a novel association of UCH-L1 with the translation initiation complex eIF4F, the target of 4EBP1. UCH-L1 associates with and promotes the assembly of eIF4F and stimulates protein synthesis through a mechanism that requires its catalytic activity. Because of the importance of mTOR in MYC-driven oncogenesis, we used novel mutant Uchl1 transgenic mice and found that catalytic activity is required for its acceleration of lymphoma in the Eμ-myc model. Further, we demonstrate that mice lacking UCH-L1 are resistant to MYC-induced lymphomas. We conclude that UCH-L1 bypasses the need for mTORC1-dependent protein synthesis by directly promoting translation initiation, and that this mechanism may be essential for MYC in B-cell malignancy.

SummaryNon‐Hodgkin lymphoma (NHL) is the third most common malignancy in children, adolescents and young adults (CAYA). NHL is a diverse set of diseases that arise at key regulatory checkpoints during B or T cell development in the bone marrow, germinal centre or thymus. While advances in the use of conventional cytotoxic agents have led to dramatic improvements in survival, these cures are associated with significant acute and long‐term toxicities. Moreover, the prognosis for CAYA patients with relapsed or refractory NHL remains dismal, with the vast majority dying of their disease. Thanks to a large number of candidate‐based biological studies, together with large‐scale sequencing efforts, there has been an explosion of knowledge regarding the molecular pathophysiology of B‐ and T‐NHL. This has ushered development of a flurry of novel therapeutic approaches that may simultaneously provide new hope for relapsed patients and an opportunity to reduce the therapeutic burden in newly diagnosed CAYA. Here we review a selection of the most promising new therapeutic approaches to these diseases. While the vast majority of these agents are untested in children, on‐going work from many cooperative groups will soon explore their use in paediatric disease, in hope of further improving outcomes while maximizing quality of life.

Key Points The neuronal marker UCH-L1 is induced in, and specifically augments the oncogene-induced transformation of, GCB cells. High levels of UCHL1 identify patients with GC DLBCL with an increased risk for poor outcomes.

Abstract BACKGROUND: Burkitt lymphoma (BL) represents approximately 40% of all childhood and adolescent non-Hodgkin lymphoma (Miles/Cairo, BJH, 2012). Children with relapsed or progressive BL develop chemotherapy-resistant disease and can rarely be cured with salvage therapy (Cairo et al, JCO, 2012). Bruton's tyrosine kinase (BTK) is a regulator of normal B-cell development and is activated upon B-cell receptor (BCR) stimulation. Chronic active BCR signaling through BTK activation can be inhibited by the selective and covalent BTK inhibitor, ibrutinib (Young et al, Nat Rev, 2013). Ibrutinib has been highly effective in the treatment of refractory patients with CLL and MCL and has been approved by the FDA for patients with CLL or MCL who have received at least one prior therapy (USPI) (Byrd et al, NEJM, 2013 and Wang et al, NEJM, 2013). BL, however, is associated with tonic or possibly chronic active BCR signaling; while, both CLL and MCL have chronic active BCR signaling. We have previously demonstrated ibrutinib significantly decrease BL proliferation and viability in vitro (Lee/Cairo et al, ASH, 2014) OBJECTIVE: We hypothesize that ibrutinib may be a potential adjuvant agent in the treatment of BL. Therefore, we investigated the in vivo anti-tumor activity of ibrutinib in BL xenografted NOD/SCID (NSG) mice. METHODS: The luciferase expression plasmid (ffluc-Zeo), kindly provided by L. Cooper, MD) was transfected into Raji cells. Cells were subcutaneously injected into NSG mice (6-8wks old, NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ, Jackson lab). Tumor cells engraftment and progression were examined by Bioluminescent Imaging using the Xenogen IVIS-200 (Caliper Life Sciences). Mice were orally gavaged with vehicle control or Ibrutinib (1.25, 12.5 and 25mg/kg/day, generously provided by Janssen R&D, LLC) for 10 days. Statistical probability of survival and curve were determined by Kaplan Meier method. RESULTS: We observed a significant decrease in tumor luminescence intensity following ibrutinib treated BL xenografted NSG mice at day 20 (12.5mg/kg, p<0.001 and 25mg/kg, p<0.05) and at day 25 (12.5mg/kg, p<0.05 and 25mg/kg, p<0.05) compared to vehicle control. Furthermore, ibrutinib (12.5 mg/kg) treated mice significantly prolonged survival compared to vehicle control mice (p<0.05). The median survival of mice following ibrutinib treatment (1.25, 12.5 and 25 mg/kg) were 31.5, 37.5 and 22.5 days, respectively, compared to control (24 days). CONCLUSIONS: Ibrutinib significantly decreased tumor progression and significantly increased the survival in BL xenografted NSG mice following 12.5mg/kg ibrutinib treatment. These results indicate that ibrutinib may be a potential adjuvant agent therapy in the treatment of BL. Future directions will be investigated the efficacy of ibrutinib in combination with dexamethasone in BL xenografted NSG mice. Citation Format: Sanghoon Lee, Changhong Yin, Timmy O'Connell, Matthew Barth, Janet Ayello, Lauren Harrison, Carmella van de Ven, Rodney Miles, Paul Galardy, Stanton C. Goldman, Megan Lim, Michelle Hermiston, Linda McAllister-Lucas, Lisa G. Roth, Sherrie L. Perkins, Mitchell S. Cairo. Ibrutinib significantly improves survival in a human Burkitt lymphoma (BL) xenograft NSG mouse model: Ibrutinib may be a potential adjuvant agent in the treatment of BL. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2608. doi:10.1158/1538-7445.AM2015-2608

SummaryPatient‐specific primers from 10 children/adolescents with Burkitt leukaemia (BL) ± central nervous system disease who were treated with French‐British‐American/Lymphome Malins de Burkitt 96 C1 plus rituximab were developed from diagnostic blood/bone marrow. Minimal residual disease (MRD) was assessed by real‐time polymerase chain reaction at the end of induction (EOI) and consolidation (EOC). Seventy per cent (7/10) and 71% (5/7) were MRD‐positive at EOI and EOC, respectively, with no disease recurrences. MRD after induction and consolidation did not predict relapse and subsequent therapy appeared to eliminate MRD. Thus, assessing MRD at a later time point is warranted in future trials to determine its clinical significance.

SummaryChildren and adolescents with Burkitt Lymphoma (BL) and combined central nervous system (CNS) and bone marrow involvement still have a poor prognosis with chemotherapy alone. We therefore investigated in children and adolescents with bone marrow (≥25% blasts) and/or CNS‐positive Burkitt lymphoma the chemoimmunotherapy combination of rituximab (375 mg/m2) and the standard chemotherapy arm of our previously reported French‐American‐British (FAB) Lymphome Malins de Burkitt (LMB) 96 trial. Central pathological and cytogenetic characterization was also performed. There were 40 evaluable patients with Burkitt histology (25 with leukaemia and 15 with CNS disease ± leukaemia). The chemoimmunotherapy regimen was well tolerated. The incidence of grade III/IV mucositis during induction cycles with combined chemotherapy and rituximab was 31% and 26%, respectively. The 3‐year event‐free survival (EFS)/overall survival (OS) was 90% (95% confidence interval [CI], 76–96%) in the entire cohort and 93% (95% CI, 61–99%) in patients with CNS disease. Based on the results of this trial, an international randomized study of FAB/LMB 96 chemotherapy ± rituximab for high‐risk patients is currently under investigation.