In red blood cells (RBCs) infected with the malaria parasite Plasmodium falciparum, a 19-residue region of the mature parasite-infected erythrocyte surface antigen (MESA) associates with RBC ...cytoskeleton protein 4.1R; an interaction essential for parasite survival. This region in MESA is adjacent to a host targeting motif found in other malaria parasite proteins exported to the membrane skeleton. To demonstrate function of these motifs in vivo, regions of MESA fused to a reporter were expressed in malaria parasites. Immunochemical analyses confirmed the requirement for both motifs in the trafficking and interaction of MESA with the cytoskeleton and demonstrates their function in vivo.
Apicomplexan parasites are characterised by the presence of specialised organelles, such as rhoptries, located at the apical end of invasive forms that play an important role in invasion of the host ...cell and formation of the parasitophorous vacuole. In this study, we have characterised a novel Plasmodium falciparum rhoptry protein, Pf34, encoded by a single exon gene located on chromosome 4 and expressed as a 34kDa protein in mature asexual stage parasites. Pf34 is expressed later in the life cycle than the previously described rhoptry protein, Rhoptry Associated Membrane Antigen (RAMA). Orthologues of Pf34 are present in other Plasmodium species and a potential orthologue has also been identified in Toxoplasma gondii. Indirect immunofluorescence assays show that Pf34 is located at the merozoite apex and localises to the rhoptry neck. Pf34, previously demonstrated to be glycosyl-phosphatidyl-inositol (GPI)-anchored Gilson, P.R., Nebl, T., Vukcevic, D., Moritz, R.L., Sargeant, T., Speed, T.P., Schofield, L., Crabb, B.S. (2006) Identification and stoichiometry of GPI-anchored membrane proteins of the human malaria parasite Plasmodium falciparum. Mol. Cell. Proteomics 5, 1286–1299., is associated with parasite-derived detergent-resistant microdomains (DRMs). Pf34 is carried into the newly invaded ring, consistent with a role for Pf34 in the formation of the parasitophorous vacuole. Pf34 is exposed to the human immune system during infection and is recognised by human immune sera collected from residents of malaria endemic areas of Vietnam and Papua New Guinea.
Chimeric antigen receptor (CAR) T cells have been highly successful in treating hematological malignancies, including acute and chronic lymphoblastic leukemia. However, treatment of solid tumors ...using CAR T cells has been largely unsuccessful to date, partly because of tumor-induced immunosuppressive mechanisms, including adenosine production. Previous studies have shown that adenosine generated by tumor cells potently inhibits endogenous antitumor T cell responses through activation of adenosine 2A receptors (A2ARs). Herein, we have observed that CAR activation resulted in increased A2AR expression and suppression of both murine and human CAR T cells. This was reversible using either A2AR antagonists or genetic targeting of A2AR using shRNA. In 2 syngeneic HER2+ self-antigen tumor models, we found that either genetic or pharmacological targeting of the A2AR profoundly increased CAR T cell efficacy, particularly when combined with PD-1 blockade. Mechanistically, this was associated with increased cytokine production of CD8+ CAR T cells and increased activation of both CD8+ and CD4+ CAR T cells. Given the known clinical relevance of the CD73/adenosine pathway in several solid tumor types, and the initiation of phase I trials for A2AR antagonists in oncology, this approach has high translational potential to enhance CAR T cell efficacy in several cancer types.
The lactate dehydrogenase-A (LDH-A) enzyme catalyzes the interconversion of pyruvate and lactate, is upregulated in human cancers, and is associated with aggressive tumor outcomes. Here we use an ...inducible murine model and demonstrate that inactivation of LDH-A in mouse models of NSCLC driven by oncogenic K-RAS or EGFR leads to decreased tumorigenesis and disease regression in established tumors. We also show that abrogation of LDH-A results in reprogramming of pyruvate metabolism, with decreased lactic fermentation in vitro, in vivo, and ex vivo. This was accompanied by reactivation of mitochondrial function in vitro, but not in vivo or ex vivo. Finally, using a specific small molecule LDH-A inhibitor, we demonstrated that LDH-A is essential for cancer-initiating cell survival and proliferation. Thus, LDH-A can be a viable therapeutic target for NSCLC, including cancer stem cell-dependent drug-resistant tumors.
Display omitted
•An inducible LDH-A mouse model has been developed and characterized•Reduced LDH-A results in regression of established tumors in NSCLC mouse models•Cancer-initiating cells are susceptible to LDH-A inhibition•Pharmacologic inhibition of LDH-A mimics the effects of genetic attenuation
Targeting cancer metabolism offers a unique therapeutic opportunity. Using an elegant mouse model to temporally control the expression of the glycolytic enzyme lactate dehydrogenase-A (LDH-A), Xie et al. show that silencing LDH-A impacts both initiation and maintenance of lung tumors. Cancer stem cells are more susceptible to LDH-A inhibition.
Abstract The rhoptry associated membrane antigen (RAMA) of Plasmodium falciparum has been proposed as a potential candidate for inclusion in a multivalent subunit vaccine against malaria. Previous ...studies have found that the RAMA gene is refractory to genetic deletion in vitro and is conserved in a range of clinical isolates. Importantly, two independent studies demonstrated that antibodies against the C-terminal region of RAMA are associated with immunity in endemic populations of both Asia and Africa. However, there is presently no direct evidence that anti-RAMA immune responses have a demonstrable anti-parasitic effect either in vitro or in vivo . In this study we used an in vitro invasion inhibition assay and the Plasmodium yoelii mouse model of infection to evaluate the potential of RAMA as a vaccine candidate. Our results demonstrate that anti-PfRAMA antibodies have only a weak inhibitory effect on P. falciparum invasion in vitro . Immunisation with recombinant PyRAMA protein did not protect mice against a lethal P. yoelii infection and did not boost the level of protection induced by a known protective antigen, merozoite surface protein 4/5. Taken together, these data do not support RAMA as a priority vaccine candidate.
Stem cell self-renewal and lineage specification are highly dynamic and tightly controlled processes that are essential for normal haematopoiesis and are dysregulated in cancer. The X-linked BCL6 ...Corepressor (BCOR) gene encodes a protein that is widely expressed across adult human tissues and is a component of a non-canonical Polycomb repressive complex 1 (PRC1). The BCOR gene is recurrently mutated in various malignant and non-malignant blood disorders, and we and others have recently provided experimental evidence that BCOR has cell-context dependent functions in regulating the proliferation, differentiation and survival of haematopoietic cells. To comprehensively examine the role of BCOR in haematopoiesis in vivo we used a conditional mouse model that mimics the truncating mutations observed in acute myeloid leukaemia (AML) and myelodysplastic syndrome (MDS). Using stem and progenitor populations isolated ex vivo we comprehensively analysed the role of BCOR in regulating gene expression, modifying chromatin and altering genome architecture. We demonstrate that BCOR has a pivotal role in down-regulating haematopoietic stem cell (HSC) associated transcriptional networks during the transition from multi-potent stem cells to lineage-committed myeloid progenitors. Inactivation of Bcor in HSCs results in expansion of myeloid progenitors and co-operates with oncogenic KrasG12D in the initiation of an aggressive and fully transplantable acute leukaemia. Mechanistically, Bcor regulates a subset of PRC1-target genes including key HSC super-enhancer-linked transcription factors that are normally down-regulated during myeloid differentiation. We used CRISPR/Cas9 to explore the function of Bcor target genes and identified those that are necessary for the proliferation of Bcor mutant leukaemic cells. This study provides a comprehensive mechanistic understanding of how BCOR regulates cell fate decisions and contributes to the development of leukaemia.
No relevant conflicts of interest to declare.
Internal tandem duplication of the FMS-like tyrosine kinase 3 gene (
) occurs in 30% of all acute myeloid leukemias (AML). Limited clinical efficacy of FLT3 inhibitors highlights the need for ...alternative therapeutic modalities in this subset of disease. Using human and murine models of FLT3-ITD-driven AML, we demonstrate that FLT3-ITD promotes serine synthesis and uptake via ATF4-dependent transcriptional regulation of genes in the
serine biosynthesis pathway and neutral amino acid transport. Genetic or pharmacologic inhibition of PHGDH, the rate-limiting enzyme of
serine biosynthesis, selectively inhibited proliferation of FLT3-ITD AMLs
and
. Moreover, pharmacologic inhibition of PHGDH sensitized FLT3-ITD AMLs to the standard-of-care chemotherapeutic cytarabine. Collectively, these data reveal novel insights into FLT3-ITD-induced metabolic reprogramming and reveal a targetable vulnerability in FLT3-ITD AML. SIGNIFICANCE: FLT3-ITD mutations are common in AML and are associated with poor prognosis. We show that FLT3-ITD stimulates serine biosynthesis, thereby rendering FLT3-ITD-driven leukemias dependent upon serine for proliferation and survival. This metabolic dependency can be exploited pharmacologically to sensitize FLT3-ITD-driven AMLs to chemotherapy.
.
Approximately 20% of acute myeloid leukaemia (AML) patients carry mutations in genes that encode the metabolic enzymes isocitrate dehydrogenase (IDH) 1 and -2. Mutant IDH proteins possess a ...neomorphic enzymatic activity and produce the oncometabolite D-2-hydroxyglutarate (2-HG) that modulates numerous pathways implicated in cell fate decisions and cancer. We and others have shown that IDH mutations can contribute to leukaemia initiation and maintenance both in vitro and in vivo. Small molecule inhibitors that block production of 2-HG are starting to enter the clinic, however, the biomarkers of response and mechanisms of de novo and acquired resistance to mutant IDH inhibition remain poorly understood. We have engineered a series of murine AML models driven by inducible expression of mutant IDH proteins. To uncover the molecular mechanisms that underpin response and adaptation to mutant IDH inhibition we integrated bulk and single-cell RNA sequencing with chromatin immuno-precipitation sequencing. We demonstrate that genetic depletion and pharmacological inhibition of mutant IDH has striking effects, altering self-renewal and differentiation programs of leukaemic cells. Importantly, we identify a population of cells that persist during treatment and characterise genetic programs that circumvent effective IDH inhibition, ultimately leading to therapy resistance and disease relapse. Our studies provide detailed mechanistic insight into the oncogenic properties of mutant IDH proteins in maintaining the transformed state of AML cells.
PDF
