A chemogenomic approach to identify personalized therapy for patients with relapse or refractory acute myeloid leukemia: results of a prospective feasibility study.

Targeted next-generation sequencing (tNGS) and ex vivo drug sensitivity/resistance profiling (DSRP) have laid foundations defining the functional genomic landscape of acute myeloid leukemia (AML) and premises of personalized medicine to guide treatment options for patients with aggressive and/or chemorefractory hematological malignancies. Here, we have assessed the feasibility of a tailored treatment strategy (TTS) guided by systematic parallel ex vivo DSRP and tNGS for patients with relapsed/refractory AML (number NCT02619071). A TTS issued by an institutional personalized committee could be achieved for 47/55 included patients (85%), 5 based on tNGS only, 6 on DSRP only, while 36 could be proposed on the basis of both, yielding more options and a better rationale. The TSS was available in <21 days for 28 patients (58.3%). On average, 3 to 4 potentially active drugs were selected per patient with only five patient samples being resistant to the entire drug panel. Seventeen patients received a TTS-guided treatment, resulting in four complete remissions, one partial remission, and five decreased peripheral blast counts. Our results show that chemogenomic combining tNGS with DSRP to determine a TTS is a promising approach to propose patient-specific treatment options within 21 days.

Anti-leukemia activity of chaetocin via death receptor-dependent apoptosis and dual modulation of the histone methyl-transferase SUV39H1.

Epigenetic deregulation is involved in acute myeloid leukemia (AML) pathogenesis and epigenetic targeting drugs are in clinical trial. Since the first results with histone-deacetylase inhibitors in AML are controversial, novel single and combined treatments need to be explored. It is tempting to combine chromatin-targeting drugs. SUV39H1, the main methyl-transferase for lysine 9 tri-methylation on histone H3, interacts with oncogenes involved in AML and acts as a transcriptional repressor for hematopoietic differentiation and immortalization. We report here that pharmacological inhibition of SUV39H1 by chaetocin induces apoptosis in leukemia cell lines in vitro and primary AML cells ex vivo, and that it interferes with leukemia growth in vivo. Chaetocin treatment upregulates reactive oxygen species (ROS) production as well as the transcription of death-receptor-related genes, in a ROS-dependent manner, leading to death receptor-dependent apoptosis. In addition to its direct inhibition by chaetocin, SUV39H1 is indirectly modulated by chaetocin-induced ROS. Accordingly, chaetocin potentiates other anti-AML drugs, in a ROS-dependent manner. The decryption of a dual mechanism of action against AML involving both direct and indirect SUV39H1 modulation represents an innovative read-out for the anticancer activity of chaetocin and for its synergy with other anti-AML drugs, suggesting new therapeutic combination strategies in AML.

Loss of AF6/afadin, a marker of poor outcome in breast cancer, induces cell migration, invasiveness and tumor growth.

Afadin/AF6, an F-actin-binding protein, is ubiquitously expressed in epithelia and has a key role during development, through its regulatory role in cell-cell junction organization. Afadin loss of expression in 15% of breast carcinoma is associated with adverse prognosis and increased risk of metastatic relapse. To determine the role of afadin in breast cancer, we studied the functional consequences of afadin protein extinction using in vitro and in vivo models. Three different breast cancer cell lines representative of the major molecular subtypes were stably repressed for afadin expression (knockdown of afadin (afadin KD)) using RNA interference. Collective and individual migrations as well as Matrigel invasion were markedly increased in afadin KD cells. Heregulin-ß1 (HRG-ß1)-induced migration and invasion were increased by twofold in afadin KD cells. Conversely, ectopic expression of afadin in the afadin-negative T47D cell line inhibited spontaneous and HRG-ß1-induced migrations. RAS/MAPK and SRC kinase pathways were activated in afadin KD cells. Activation levels positively correlated with migration and invasion strength. Use of MEK1/2 (U0126) and SRC kinases (SU6656) inhibitors reduced afadin-dependent migration and invasion. Afadin extinction in the SK-BR-3 cell line markedly accelerated tumor growth development in mouse mammary gland and lung metastasis formation. These results may explain why the loss of afadin expression in tumors correlates with high tumor size and poor metastasis-free survival in patients.

Evidences for ubiquitination and intracellular trafficking of LAT, the linker of activated T cells.

Current evidences indicate that T cells use protein sorting and degradation to control duration and specificity of T cell receptor (TcR) signalling, including the CD3zeta chain which is ubiquitinated upon TcR triggering. In a previous study, we showed that the Linker of activated T cells (LAT) is present at the plasma membrane and in transferrin-labelled intracellular compartments also containing the CD3zeta chain. Here we show that LAT protein levels are tightly regulated in Jurkat lymphoid T cells likely involving proteasome-dependent degradation, recycling through trans-Golgi/endosome compartments and clathrin-dependent internalisation. We further identify a novel post-translational modification of LAT by ubiquitination that is likely to influence LAT protein stability, intracellular localisation and/or recycling. Our results provide novel molecular and regulatory insights into the function of LAT adapter protein in T cell signalling.