The use of allogeneic haematopoietic stem cell transplantation (Allo-HSCT) is a standard treatment option for many patients with haematological malignancies. Historically, patients requiring intensive care unit (ICU) admission for transplant-related toxicities have fared extremely poorly, with high ICU mortality rates. Little is known about the impact of reduced intensity Allo-HSCT conditioning regimens in older patients on the ICU and subsequent long-term outcomes. A retrospective analysis of data collected from 164 consecutive Allo-HSCT recipients admitted to ICU for a total of 213 admissions, at a single centre over an 11·5-year study period was performed. Follow-up was recorded until 31 March 2011. Autologous HSCT recipients were excluded. In this study we report favourable ICU survival following Allo-HSCT and, for the first time, demonstrate significantly better survival for patients who underwent Allo-HSCT with reduced intensity conditioning compared to those treated with myeloablative conditioning regimens. In addition, we identified the need for ventilation (invasive or non-invasive) as an independently significant adverse factor affecting short-term ICU outcome. For patients surviving ICU admission, subsequent long-term overall survival was excellent; 61% and 51% at 1 and 5 years, respectively. Reduced intensity Allo-HSCT patients admitted to ICU with critical illness have improved survival compared to myeloablative Allo-HSCT recipients.
Purpose
To determine whether inhibition of mechanistic target of rapamycin (mTOR) kinase-mediated signaling represents a valid therapeutic approach for chronic lymphocytic leukemia (CLL).
Experimental Design
Stratification of mTOR activity was carried out in primary CLL patient samples and an aggressive CLL-like mouse model. The potency of dual mTOR inhibitor AZD8055 to induce apoptosis in primary CLL cells was assessed in the presence/absence of B cell receptor (BCR) ligation. Furthermore, we addressed the molecular and functional impact of dual mTOR inhibition in combination with BTK inhibitor ibrutinib.
Results
Differential regulation of basal mTORC1 activity was observed in poor prognostic CLL samples, with elevated p4EBP1T37/46 and decreased p70S6 kinase activity, suggesting that dual mTORC1/2 inhibitors may exhibit improved response in poor prognostic CLL compared with rapalogs. AZD8055 treatment of primary CLL cells significantly reduced CLL survival in vitro compared with rapamycin, preferentially targeting poor prognostic subsets and overcoming BCR-mediated survival advantages. Furthermore, AZD8055, and clinical analog AZD2014, significantly reduced CLL tumor load in mice. AKT substrate FOXO1, while overexpressed in CLL cells of poor prognostic patients in LN biopsies, peripheral CLL cells, and mouse-derived CLL-like cells, appeared to be inactive. AZD8055 treatment partially reversed FOXO1 inactivation downstream of BCR crosslinking, significantly inhibiting FOXO1T24 phosphorylation in an mTORC2-AKT-dependent manner, to promote FOXO1 nuclear localization, activity and FOXO1-mediated gene regulation. FOXO1 activity was further significantly enhanced on combining AZD8055 with ibrutinib.
Conclusions
Our studies demonstrate that dual mTOR inhibitors show promise as future CLL therapies, particularly in combination with ibrutinib.
Mechanistic target for rapamycin (mTOR) is a serine/threonine protein kinase that forms two distinct complexes mTORC1 and mTORC2, integrating mitogen and nutrient signals to regulate cell survival and proliferation; processes which are commonly deregulated in human cancers. mTORC1 and mTORC2 have divergent molecular associations and cellular functions: mTORC1 regulates in mRNA translation and protein synthesis, while mTORC2 is involved in the regulation of cellular survival and metabolism. Through AKT phosphorylation/activation, mTORC2 has also been reported to regulate cell migration. Recent attention has focused on the aberrant activation of the PI3K/mTOR pathway in B cell malignancies and there is growing evidence for its involvement in disease pathogenesis, due to its location downstream of other established novel drug targets that intercept B cell receptor (BCR) signals. Shared pharmacological features of BCR signal inhibitors include a striking "lymphocyte redistribution" effect whereby patients experience a sharp increase in lymphocyte count on initiation of therapy followed by a steady decline. Chronic lymphocytic leukemia (CLL) serves as a paradigm for migration studies as lymphocytes are among the most widely travelled cells in the body, a product of their role in immunological surveillance. The subversion of normal lymphocyte movement in CLL is being elucidated; this review aims to describe the migration impairment which occurs as part of the wider context of cancer cell migration defects, with a focus on the role of mTOR in mediating migration effects downstream of BCR ligation and other microenvironmental signals.
MYC and RUNX oncogenes each trigger p53‐mediated failsafe responses when overexpressed in vitro and collaborate with p53 deficiency in vivo. However, together they drive rapid onset lymphoma without mutational loss of p53. This phenomenon was investigated further by transcriptomic analysis of premalignant thymus from RUNX2/MYC transgenic mice. The distinctive contributions of MYC and RUNX to transcriptional control were illustrated by differential enrichment of canonical binding sites and gene ontology analyses. Pathway analysis revealed signatures of MYC, CD3, and CD28 regulation indicative of activation and proliferation, but also strong inhibition of cell death pathways. In silico analysis of discordantly expressed genes revealed Tnfsrf8/CD30, Cish, and Il13 among relevant targets for sustained proliferation and survival. Although TP53 mRNA and protein levels were upregulated, its downstream targets in growth suppression and apoptosis were largely unperturbed. Analysis of genes encoding p53 posttranslational modifiers showed significant upregulation of three genes, Smyd2, Set, and Prmt5. Overexpression of SMYD2 was validated in vivo but the functional analysis was constrained by in vitro loss of p53 in RUNX2/MYC lymphoma cell lines. However, an early role is suggested by the ability of SMYD2 to block senescence‐like growth arrest induced by RUNX overexpression in primary fibroblasts.
B cell antigen receptor (BCR) signalling competence is critical for the pathogenesis of chronic lymphocytic leukaemia (CLL). Defining key proteins that facilitate these networks aid in the identification of targets for therapeutic exploitation. We previously demonstrated that reduced PKCα function in mouse hematopoietic stem/progenitor cells (HPSCs) resulted in PKCβII upregulation and generation of a poor-prognostic CLL-like disease. Here, prkcb knockdown in HSPCs leads to reduced survival of PKCα-KR-expressing CLL-like cells, concurrent with reduced expression of the leukemic markers CD5 and CD23. SP1 promotes elevated expression of prkcb in PKCα-KR expressing cells enabling leukemogenesis. Global gene analysis revealed an upregulation of genes associated with B cell activation in PKCα-KR expressing cells, coincident with upregulation of PKCβII: supported by activation of key signalling hubs proximal to the BCR and elevated proliferation. Ibrutinib (BTK inhibitor) or enzastaurin (PKCβII inhibitor) treatment of PKCα-KR expressing cells and primary CLL cells showed similar patterns of Akt/mTOR pathway inhibition, supporting the role for PKCβII in maintaining proliferative signals in our CLL mouse model. Ibrutinib or enzastaurin treatment also reduced PKCα-KR-CLL cell migration towards CXCL12. Overall, we demonstrate that PKCβ expression facilitates leukemogenesis and identify that BCR-mediated signalling is a key driver of CLL development in the PKCα-KR model.
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