Non-canonical activation of ß-catenin by PRL-3 phosphatase in acute myeloid leukemia.

Aberrant activation of Wnt/ß-catenin signaling pathway is essential for the development of AML; however, the mechanistic basis for this dysregulation is unclear. PRL-3 is an oncogenic phosphatase implicated in the development of LSCs. Here, we identified Leo1 as a direct and specific substrate of PRL-3. Serine-dephosphorylated form of Leo1 binds directly to ß-catenin, promoting the nuclear accumulation of ß-catenin and transactivation of TCF/LEF downstream target genes such as cyclin D1 and c-myc. Importantly, overexpression of PRL-3 in AML cells displayed enhanced sensitivity towards ß-catenin inhibition in vitro and in vivo, suggesting that these cells are addicted to ß-catenin signaling. Altogether, our study revealed a novel regulatory role of PRL-3 in the sustenance of aberrant ß-catenin signaling in AML. PRL-3 may serve as a biomarker to select for the subset of AML patients who are likely to benefit from treatment with ß-catenin inhibitors. Our study presents a new avenue of cancer inhibition driven by PRL-3 overexpression or ß-catenin hyperactivation.

A loss-of-function genetic screening reveals synergistic targeting of AKT/mTOR and WTN/ß-catenin pathways for treatment of AML with high PRL-3 phosphatase.

BACKGROUND: Protein tyrosine phosphatase of regenerating liver 3 (PRL-3) is overexpressed in a subset of AML patients with inferior prognosis, representing an attractive therapeutic target. However, due to relatively shallow pocket of the catalytic site of PRL-3, it is difficult to develop selective small molecule inhibitor. METHODS: In this study, we performed whole-genome lentiviral shRNA library screening to discover synthetic lethal target to PRL-3 in AML. We used specific small molecule inhibitors to validate the synthetic lethality in human PRL-3 high vs PRL-3 low human AML cell lines and primary bone marrow cells from AML patients. AML mouse xenograft model was used to examine the in vivo synergism. RESULTS: The list of genes depleted in TF1-hPRL3 cells was particularly enriched for members involved in WNT/ß-catenin pathway and AKT/mTOR signaling. These findings prompted us to explore the impact of AKT/mTOR signaling inhibition in PRL-3 high AML cells in combination with WNT/ß-catenin inhibitor. VS-5584, a novel, highly selective dual PI3K/mTOR inhibitor, and ICG-001, a WNT inhibitor, were used as a combination therapy. A synthetic lethal interaction between mTOR/AKT pathway inhibition and WNT/ß-catenin was validated by a variety of cellular assays. Notably, we found that treatment with these two drugs significantly reduced leukemic burden and prolonged survival of mice transplanted with human PRL-3 high AML cells, but not with PRL-3 low AML cells. CONCLUSIONS: In summary, our results support the existence of cooperative signaling networks between AKT/mTOR and WNT/ß-catenin pathways in PRL-3 high AML cells. Simultaneous inhibition of these two pathways could achieve robust clinical efficacy for this subtype of AML patient with high PRL-3 expression and warrant further clinical investigation.

Identification of novel fusion transcripts in multiple myeloma.

AIMS: Multiple myeloma (MM) is a heterogeneous disease characterised by genetically complex abnormalities. The classical mutational spectrum includes recurrent chromosomal aberrations and gene-level mutations. Recurrent translocations involving the IGH gene such as t(11;14), t(4;14) and t(14;16) are well known. However, the presence of complex genetic abnormalities raises the possibility that fusions other than the recurrent IGH translocations exist. We therefore employed a targeted RNA-sequencing panel to identify novel putative fusions in a local cohort of MM. METHODS: Targeted RNA-sequencing was performed on 21 patient samples using the Illumina TruSight RNA Pan-Cancer Panel (comprising 1385 genes). Fusion calls were generated from the Illumina RNA-Sequencing Alignment software (V.1.0.0). These samples had conventional cytogenetic and fluorescence in situ hybridisation data for the common recurrent chromosomal abnormalities (t(11;14), t(4;14), t(14;16) and 17p13 deletion). The MMRF CoMMpass dataset was analysed using the TopHat-fusion pipeline. RESULTS: A total of 10 novel fusions were identified by the TruSight RNA Pan-Cancer Panel. Two of these fusions, HGF/CACNA2D1 and SMC3/MXI1, were validated by reverse transcription PCR and Sanger sequencing as they involve genes that may have biological relevance in MM genesis. Four of these (MAP2K4/MAP2K4P1) are likely to be spurious secondary to misalignment of reads to a pseudogene. One record of the HGF/CACNA2D1 fusion was identified from the MMRF CoMMpass dataset. CONCLUSIONS: The identification of novel fusions offers insights into the biology of MM and might have clinical relevance. Further functional studies are required to determine the biological and clinical relevance of these novel fusions.

Inhibition of LIN28B impairs leukemia cell growth and metabolism in acute myeloid leukemia.

BACKGROUND: Current conventional chemotherapy for acute myeloid leukemia (AML) can achieve remission in over 70% of patients, but a majority of them will relapse within 5 years despite continued treatment. The relapse is postulated to be due to leukemia stem cells (LSCs), which are different from normal hematopoietic stem cells (HSCs). LIN28B is microRNA regulator and stem cell reprogramming factor. Overexpression of LIN28B has been associated with advance human malignancies and cancer stem cells (CSCs), including AML. However, the molecular mechanism by which LIN28B contributes to the development of AML remains largely elusive. METHODS: We modulated LIN28B expression in AML and non-leukemic cells and investigated functional consequences in cell proliferation, cell cycle, and colony-forming assays. We performed a microarray-based analysis for LIN28B-silencing cells and interrogated gene expression data with different bioinformatic tools. AML mouse xenograft model was used to examine the in vivo function of LIN28B. RESULTS: We demonstrated that targeting LIN28B in AML cells resulted in cell cycle arrest, inhibition of cell proliferation and colony formation, which was induced by de-repression of let-7a miRNA. On the other hand, overexpression of LIN28B promoted cell proliferation. Data point to a mechanism where that inhibition of LIN28B induces metabolic changes in AML cells. IGF2BP1 was confirmed to be a novel downstream target of LIN28B via let-7 miRNA in AML. Notably, ectopic expression of LIN28B increased tumorigenicity, while silencing LIN28B led to slow tumor growth in vivo. CONCLUSIONS: In sum, these results uncover a novel mechanism of an important regulatory signaling, LIN28B/let-7/IGF2BP1, in leukemogenesis and provide a rationale to target this pathway as effective therapeutic strategy.

LIN28B Activation by PRL-3 Promotes Leukemogenesis and a Stem Cell-like Transcriptional Program in AML.

PRL-3 (PTP4A3), a metastasis-associated phosphatase, is also upregulated in patients with acute myeloid leukemia (AML) and is associated with poor prognosis, but the underlying molecular mechanism is unknown. Here, constitutive expression of PRL-3 in human AML cells sustains leukemogenesis in vitro and in vivo Furthermore, PRL-3 phosphatase activity dependently upregulates LIN28B, a stem cell reprogramming factor, which in turn represses the let-7 mRNA family, inducing a stem cell-like transcriptional program. Notably, elevated levels of LIN28B protein independently associate with worse survival in AML patients. Thus, these results establish a novel signaling axis involving PRL-3/LIN28B/let-7, which confers stem cell-like properties to leukemia cells that is important for leukemogenesis.Implications: The current study offers a rationale for targeting PRL-3 as a therapeutic approach for a subset of AML patients with poor prognosis. Mol Cancer Res; 15(3); 294-303. ©2016 AACR.

PRIMA-1met (APR-246) inhibits growth of colorectal cancer cells with different p53 status through distinct mechanisms.

PRIMA-1met (APR-246) is a methylated derivative and structural analog of PRIMA-1 (p53 re-activation and induction of massive apoptosis). PRIMA-1met has been reported to restore both the wild type (wt) structure and function of mutant p53. Here, we show that PRIMA-1met is highly effective at limiting the growth of CRC cells regardless of p53 status. However, PRIMA-1met induces robust apoptosis only in CRC cells with mutant p53. Upregulation of Noxa, a proapoptotic molecule, is crucial for PRIMA-1met mediated activity. In human xenograft model of disease, PRIMA-1met effectively suppresses CRC tumor growth. Our results uncover distinct mechanisms of PRIMA-1met in CRC with different p53 status, thus providing a mechanistic rationale to evaluate the clinical efficacy of PRIMA-1met in CRC patients with different p53 status.