The tight control of wild-type (wt) p53 by mainly MDM2 in normal cells is permanently lost in tumors harboring mutant p53 (mutp53), which exhibit dramatic constitutive p53 hyperstabilization that far exceeds that of wtp53 tumors. Importantly, mutp53 hyperstabilization is critical for mutp53′s oncogenic gain-of function in vivo. Current insight into the mechanism of this dysregulation is fragmentary and largely derived from ectopically constructed cell systems. Importantly, mutp53 knockin mice established that normal mutp53 tissues have sufficient enzymatic reserves in MDM2 and other E3 ligases to maintain full control of mutp53. We find that in human cancer cells endogenous mutant p53, despite its ability to interact with MDM2, suffers from a profound lack of ubiquitination as the root of its degradation defect. In contrast to wtp53, the many mutp53 proteins which are conformationally aberrant are engaged in complexes with the HSP90 chaperone machinery to prevent its aggregation. In contrast to wtp53 cancer cells, we show that in mutp53 cancer cells this HSP90 interaction blocks the endogenous MDM2 and CHIP E3 ligase activity. Interference with HSP90 either by RNAi against HSF1, the transcriptional regulator of the HSP90 pathway, or by direct knockdown of Hsp90 protein or by pharmacological inhibition of Hsp90 activity with 17AAG destroys the complex, liberates mutp53 and reactivates endogenous MDM2 and CHIP to degrade mutp53. Of note, 17AAG induces a stronger viability loss in mutp53 than in wtp53 cancer cells. Our data supports the rationale that suppression of mutp53 levels in vivo in established cancers might achieve clinically significant effects.

Key Points• Disease initiation and maintenance in murine AML models occurs via HIF-1a independent mechanisms. • HIF-1a deficiency in mice accelerates leukemogenesis induced by certain oncogenes.Self-renewal of hematopoietic stem cells (HSCs) and leukemia-initiating cells (LICs) has been proposed to be influenced by low oxygen tension (hypoxia). This signaling, related to the cellular localization inside the bone marrow niche and/or influenced by extrinsic factors, promotes the stabilization of hypoxia-inducible factors (HIFs). Whether HIF-1a can be used as a therapeutic target in the treatment of myeloid malignancies remains unknown. We have used 3 different murine models to investigate the role of HIF-1a in acute myeloid leukemia (AML) initiation/progression and self-renewal of LICs. Unexpectedly, we failed to observe a delay or prevention of disease development from hematopoietic cells lacking Hif-1a. In contrast, deletion of Hif-1a resulted in faster development of the disease and an enhanced leukemia phenotype in some of the investigated models. Our results therefore warrant reconsideration of the role of HIF-1a and, as a consequence, question its generic therapeutic usefulness in AML. (Blood. 2014;124(24):3597-3607)

Relapsed/refractory T-cell acute lymphoblastic leukemia (T-ALL) has a dismal outcome, and no effective targeted immunotherapies for T-ALL exist. The extension of chimeric antigen receptor (CAR) T cells (CARTs) to T-ALL remains challenging because the shared expression of target antigens between CARTs and T-ALL blasts leads to CART fratricide. CD1a is exclusively expressed in cortical T-ALL (coT-ALL), a major subset of T-ALL, and retained at relapse. This article reports that the expression of CD1a is mainly restricted to developing cortical thymocytes, and neither CD34+ progenitors nor T cells express CD1a during ontogeny, confining the risk of on-target/off-tumor toxicity. We thus developed and preclinically validated a CD1a-specific CAR with robust and specific cytotoxicity in vitro and antileukemic activity in vivo in xenograft models of coT-ALL, using both cell lines and coT-ALL patient–derived primary blasts. CD1a-CARTs are fratricide resistant, persist long term in vivo (retaining antileukemic activity in re-challenge experiments), and respond to viral antigens. Our data support the therapeutic and safe use of fratricide-resistant CD1a-CARTs for relapsed/refractory coT-ALL.