The Dual Role of Autophagy in Crizotinib-Treated ALK ALCL: From the Lymphoma Cells Drug Resistance to Their Demise.

Autophagy has been described as harboring a dual role in cancer development and therapy. Depending on the context, it can exert either pro-survival or pro-death functions. Here, we review what is known about autophagy in crizotinib-treated ALK ALCL.

Crizotinib Resistance Mediated by Autophagy Is Higher in the Stem-Like Cell Subset in ALK-Positive Anaplastic Large Cell Lymphoma, and This Effect Is MYC-Dependent.

Previously it was shown that autophagy contributes to crizotinib resistance in ALK-positive anaplastic large cell lymphoma (ALK + ALCL). We asked if autophagy is equally important in two distinct subsets of ALK + ALCL, namely eporter nresponsive () and eporter esponsive (), of which RR cells display stem-like properties. Autophagic flux was assessed with a fluorescence tagged LC3 reporter and immunoblots to detect endogenous LC3 alongside chloroquine, an autophagy inhibitor.

High Levels of miR-7-5p Potentiate Crizotinib-Induced Cytokilling and Autophagic Flux by Targeting RAF1 in NPM-ALK Positive Lymphoma Cells.

Anaplastic lymphoma kinase positive anaplastic large cell lymphomas (ALK+ ALCL) are an aggressive pediatric disease. The therapeutic options comprise chemotherapy, which is efficient in approximately 70% of patients, and targeted therapies, such as crizotinib (an ALK tyrosine kinase inhibitor (TKI)), used in refractory/relapsed cases. Research efforts have also converged toward the development of combined therapies to improve treatment.

Therapeutic Modulation of Autophagy in Leukaemia and Lymphoma.

Haematopoiesis is a tightly orchestrated process where a pool of hematopoietic stem and progenitor cells (HSPCs) with high self-renewal potential can give rise to both lymphoid and myeloid lineages. The HSPCs pool is reduced with ageing resulting in few HSPC clones maintaining haematopoiesis thereby reducing blood cell diversity, a phenomenon called clonal haematopoiesis. Clonal expansion of HSPCs carrying specific genetic mutations leads to increased risk for haematological malignancies.

Blockade of crizotinib-induced BCL2 elevation in ALK-positive anaplastic large cell lymphoma triggers autophagy associated with cell death.

Anaplastic lymphoma kinase (ALK)-positive anaplastic large cell lymphomas are tumors that carry translocations involving the gene at the 2p23 locus, leading to the expression of ALK tyrosine kinase fusion oncoproteins. Amongst hematologic malignancies, these lymphomas are particular in that they express very low levels of B-cell lymphoma 2 (BCL2), a recognized inhibitor of apoptosis and autophagy, two processes that share complex interconnections. We have previously shown that treatment of ALK-positive anaplastic large cell lymphoma cells with the ALK tyrosine kinase inhibitor crizotinib induces autophagy as a pro-survival response.

Targeting Autophagy in ALK-Associated Cancers.

Autophagy is an evolutionarily conserved catabolic process, which is used by the cells for cytoplasmic quality control. This process is induced following different kinds of stresses e.g.

[The yin and the yang of autophagy in cancer cells].

Autophagy is a self-cannibalism process essential for tissue homeostasis, which can be activated following different environmental stressful conditions. In normal cells, autophagy could act as a brake to prevent tumorigenesis, but cancer cells are able to hijack this process to their own benefit, to promote tumor growth and/or tumor resistance to anti-cancer therapies. Scientists and clinicians attempt to modulate this process to improve therapies, using autophagy inhibitors or activators, some of them being tested currently in clinical trials against several types of tumors.

Targeting autophagy enhances the anti-tumoral action of crizotinib in ALK-positive anaplastic large cell lymphoma.

Anaplastic Lymphoma Kinase-positive Anaplastic Large Cell Lymphomas (ALK+ ALCL) occur predominantly in children and young adults. Their treatment, based on aggressive chemotherapy, is not optimal since ALCL patients can still expect a 30% 2-year relapse rate. Tumor relapses are very aggressive and their underlying mechanisms are unknown.

Reversal of microRNA-150 silencing disadvantages crizotinib-resistant NPM-ALK(+) cell growth.

The regulatory microRNA miR-150 is involved in the development of hemopathies and is downregulated in T-lymphomas, such as anaplastic large-cell lymphoma (ALCL) tumors. ALCL is defined by the presence or absence of translocations that activate the anaplastic lymphoma kinase (ALK), with nucleophosmin-ALK (NPM-ALK) fusions being the most common. Here, we compared samples of primary NPM-ALK(+) and NPM-ALK(-) ALCL to investigate the role of miR-150 downstream of NPM-ALK.

Twenty years of modelling NPM-ALK-induced lymphomagenesis.

Our current understanding of oncogenic Anaplastic Lymphoma Kinase (ALK)-induced lymphomagenesis has relied for over 20 years on multiple and complementary studies performed on various experimental models, encompassing ALK oncogene expressing cells, their grafts into immune-compromised mice, the generation of genetically engineered mouse models (GEMMs) and, when available, the use of patient samples from Anaplastic Large Cell Lymphoma (ALCL) tumour banks. Of note, and to our knowledge, no ALK-positive ALCL 3D culture system has been described so far. In this review, we will first outline how these different cell and mouse models were designed, and what key findings they revealed (or confirmed) towards oncogenic ALK-induced lymphomagenesis.

ALK+ALCLs induce cutaneous, HMGB-1-dependent IL-8/CXCL8 production by keratinocytes through NF-κB activation.

Anaplastic large-cell lymphomas (ALCLs) bearing the t(2;5) translocation (ALK(+)ALCLs) are frequently characterized by skin colonization and associated with a poor prognosis. Using conditional transgenic models of anaplastic lymphoma kinase-positive (ALK(+)) lymphomas and human ALK(+)ALCL cell lines, in the present study, we show that high-mobility-group box-1 (HMGB-1), a proinflammatory cytokine, is released by ALK(+) cells, and demonstrate extracellular HMGB-1-stimulated secretion of the IL-8 chemokine by HaCaT keratinocytes through the involvement of MMP-9, PAR-2, and the NF-κB pathway. Furthermore, we demonstrate that, in vitro, IL-8 is able to induce the invasiveness of ALK(+) cells, which express the IL-8 receptors CXCR1 and CXCR2.

A novel antiangiogenic and vascular normalization therapy targeted against human CD160 receptor.

Angiogenesis plays an essential role in several diseases of the eye and in the growth of solid tumors, but existing antiangiogenic therapies have limited benefits in several cases. We report the antiangiogenic effects of a monoclonal antibody, CL1-R2, in several animal models of neovascularization. CL1-R2 recognizes human CD160, a membrane receptor which is conserved in various mammal species.

Conditional TPM3-ALK and NPM-ALK transgenic mice develop reversible ALK-positive early B-cell lymphoma/leukemia.

NPM-ALK (nucleophosmin-anaplastic lymphoma kinase) and TPM3-ALK (nonmuscular tropomyosin 3-anaplastic lymphoma kinase) are oncogenic tyrosine kinases implicated in the pathogenesis of human ALK-positive lymphoma. We report here the development of novel conditional mouse models for ALK-induced lymphomagenesis, with the use of the tetracycline regulatory system under the control of the EmuSRalpha enhancer/promoter. The expression of either oncogene resulted in the arrest of the differentiation of early B cells and lymphomagenesis.

Combined Inactivation of MYC and K-Ras oncogenes reverses tumorigenesis in lung adenocarcinomas and lymphomas.

Background: Conditional transgenic models have established that tumors require sustained oncogene activation for tumor maintenance, exhibiting the phenomenon known as "oncogene-addiction." However, most cancers are caused by multiple genetic events making it difficult to determine which oncogenes or combination of oncogenes will be the most effective targets for their treatment.

Methodology/principal Findings: To examine how the MYC and K-ras(G12D) oncogenes cooperate for the initiation and maintenance of tumorigenesis, we generated double conditional transgenic tumor models of lung adenocarcinoma and lymphoma.

Development of a conditional bioluminescent transplant model for TPM3-ALK-induced tumorigenesis as a tool to validate ALK-dependent cancer targeted therapy.

Overexpression and activation of TPM3-ALK tyrosine kinase fusion protein is a causal oncogenic event in the development of Anaplastic Large Cell Lymphoma and Inflammatory Myofibroblastic ALK-positive tumors. Thus, the development of ALK specific tyrosine kinase inhibitors is a current therapeutic challenge. Animal models are essential to assess, in vivo, the efficiency of ALK-oncogene inhibitors and to identify new and/or additional therapeutic targets in the ALK tumorigenesis pathway.

Gene-expression profiling of systemic anaplastic large-cell lymphoma reveals differences based on ALK status and two distinct morphologic ALK+ subtypes.

With the use of microarray gene-expression profiling, we analyzed a homogeneous series of 32 patients with systemic anaplastic large-cell lymphoma (ALCL) and 5 ALCL cell lines. Unsupervised analysis classified ALCL in 2 clusters, corresponding essentially to morphologic subgroups (ie, common type vs small cell and "mixed" variants) and clinical variables. Patients with a morphologic variant of ALCL had advanced-stage disease.

Sustained regression of tumors upon MYC inactivation requires p53 or thrombospondin-1 to reverse the angiogenic switch.

The targeted inactivation of oncogenes offers a rational therapeutic approach for the treatment of cancer. However, the therapeutic inactivation of a single oncogene has been associated with tumor recurrence. Therefore, it is necessary to develop strategies to override mechanisms of tumor escape from oncogene dependence.

Global analysis of proliferation and cell cycle gene expression in the regulation of hematopoietic stem and progenitor cell fates.

Knowledge of the molecular networks controlling the proliferation and fate of hematopoietic stem cells (HSC) is essential to understand their function in maintaining blood cell production during normal hematopoiesis and upon clinical transplantation. Using highly purified stem and progenitor cell populations, we define the proliferation index and status of the cell cycle machinery at discrete stages of hematopoietic differentiation and during cytokine-mediated HSC mobilization. We identify distinct sets of cell cycle proteins that specifically associate with differentiation, self-renewal, and maintenance of quiescence in HSC and progenitor cells.

Comparative genomic hybridization on mouse cDNA microarrays and its application to a murine lymphoma model.

Microarray-based formats offer a high-resolution alternative to conventional, chromosome-based comparative genomic hybridization (CGH) methods for assessing DNA copy number alteration (CNA) genome-wide in human cancer. For murine tumors, array CGH should provide even greater advantage, since murine chromosomes are more difficult to individually discern. We report here the adaptation and evaluation of a cDNA microarray-based CGH method for the routine characterization of CNAs in murine tumors, using mouse cDNA microarrays representing approximately 14,000 different genes, thereby providing an average mapping resolution of 109 kb.

Conditional animal models: a strategy to define when oncogenes will be effective targets to treat cancer.

The ability to model cancer in the mouse has provided a robust methodology to dissect the molecular etiology of cancer. These models serve as potentially powerful platforms to preclinically evaluate novel therapeutics. In particular, the recent development of strategies to conditionally induce the or knockout the function of genes in a tissue specific manner has enabled investigators to engineer mice to demonstrate that the targeted inactivation of specific oncogenes can be effective in inducing sustained regression of tumors.

SH2-containing inositol 5-phosphatases 1 and 2 in blood platelets: their interactions and roles in the control of phosphatidylinositol 3,4,5-trisphosphate levels.

Src homology domain 2-containing inositol 5-phosphatases 1 and 2 (SHIP1 and SHIP2) are capable of dephosphorylating the second messenger PtdIns(3,4,5) P3 (phosphatidylinositol 3,4,5-trisphosphate) and interacting with several signalling proteins. SHIP1 is essentially expressed in haematopoietic cells, whereas SHIP2, a closely related enzyme, is ubiquitous. In the present study, we show that SHIP1 and SHIP2 are expressed as functional PtdIns(3,4,5) P3 5-phosphatases in human blood platelets and are capable of interacting when these two lipid phosphatases are co-expressed, either naturally (platelets and A20 B lymphoma cells) or artificially (COS-7 cells).

How cancers escape their oncogene habit.

Many recent reports demonstrate that at least initially, the inactivation of an oncogene can induce sustained regression of even a highly invasive and genetically complex cancer. However, upon prolonged oncogene inactivation, some cancers ultimately relapse, becoming independent of the very oncogene that initiated the process of tumorigenesis. Understanding the specific mechanisms by which cancers can escape dependence upon a particular oncogene will be critical to anticipate mechanisms by which human cancers will evade therapies that target individual oncogenes.

Genomically complex lymphomas undergo sustained tumor regression upon MYC inactivation unless they acquire novel chromosomal translocations.

The targeted inactivation of oncogenes may be a specific and effective treatment for cancer. However, because human cancers are the consequence of multiple genetic changes, the inactivation of one oncogene may not be sufficient to cause sustained tumor regression. Moreover, cancers are genomically unstable and may readily compensate for the inactivation of a single oncogene.

Conversion of PtdIns(4,5)P(2) into PtdIns(5)P by the S.flexneri effector IpgD reorganizes host cell morphology.

Phosphoinositides play a central role in the control of several cellular events including actin cytoskeleton organization. Here we show that, upon infection of epithelial cells with the Gram-negative pathogen Shigella flexneri, the virulence factor IpgD is translocated directly into eukaryotic cells and acts as a potent inositol 4-phosphatase that specifically dephosphorylates phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P(2)] into phosphatidylinositol 5-monophosphate [PtdIns(5)P] that then accumulates. Transfection experiments indicate that the transformation of PtdIns(4,5)P(2) into PtdIns(5)P by IpgD is responsible for dramatic morphological changes of the host cell, leading to a decrease in membrane tether force associated with membrane blebbing and actin filament remodelling.