MRNA Profiling Involved in Triggering of STAT1 with Regulatory Involvement of IRF7, PTPRF, and miR-145p in Patients Suffering from Gall Bladder Carcinoma

Background. Gall Bladder Cancer (GBC) is a type of extremely malignant tumor, which has high incidences of mortality. There is rare information about its mechanisms of invasion and gene expression regulations. microRNA-155 (miR-155) has mostly been reported to be over expressed in cases of solid tumors and hematopoietic malignancies. In this study, we have investigated the role and clinical significance of miR-155 in a Chinese population suffering from GBC and compared the results with nonneoplastic inflammation. Methods. Tissue specimens were collected on 50 patients of Gall Bladder Carcinoma and 10 patients suffering from nonneoplastic inflammation who have undergone surgeries at the Department of Pathology, Renji Hospital, Shanghai, from January 2019 to January 2020. We performed profiling of miR-155 expression in both nonneoplastic and gall bladder carcinoma tissues by QRT-PCR. Results. Expression levels of miR-155 were found to be extremely high in GBC patients in comparison to the nonneoplastic tissues ( ∗ P < 0.05 ), as high miRNA is correlated with TNM stages. Further results noted were that miR-145-5p expressed genes mimic the gene expression of STAT1, a downregulation of IRF7 was noted in the GBC, and an activation of STAT1 was significantly noted in carcinoma cells of the gallbladder. Downregulation of PTPRF was also noted during the expression of miR-145. Conclusions. As downregulation of IRF7 is linked with low rates of survival, it was found that gall bladder carcinoma patients may face high mortality. The STAT-1 expression of unregulated in GBC patients was also noted.

Clonal hematopoiesis in individuals with ANKRD26 or ETV6 germline mutations

Currently, there are at least a dozen recognized hereditary hematopoietic malignancies (HHMs), some of which phenocopy others. Among these, three HHMs driven by germline mutations in ANKRD26, ETV6, or RUNX1 share a phenotype of thrombocytopenia, qualitative platelet defects, and an increased lifetime risk of hematopoietic malignancies (HMs). Prior work has demonstrated that RUNX1 germline mutation carriers experience an elevated lifetime risk (66%) for developing clonal hematopoiesis (CH) prior to age 50. Germline mutations in ANKRD26 or ETV6 phenocopy RUNX1 germline mutations, but no studies have focused on the risk of CH in individuals with germline mutations in ANKRD26 or ETV6. To determine the prevalence of CH in individuals with germline mutations in ANKRD26 or ETV6, we performed next generation sequencing on hematopoietic tissue from twelve individuals with either germline ANKRD26 or germline ETV6 mutations. Each patient had thrombocytopenia but had not developed HMs. Among the seven individuals with germline ANKRD26 mutations, one patient had a CH clone driven by a somatic SF3B1 mutation (p.Lys700Glu). This mutation increased from a variant allele frequency (VAF) of 9.4% at age 56 to 17.4% at age 60. None of the germline ETV6 mutation carriers had evidence of CH at the limits of detection of the NGS assay (5% VAF). Unlike individuals with germline mutations in RUNX1, no individuals under the age of 50 with germline mutations in ANKRD26 or ETV6 had detectable CH. This work demonstrates that ANKRD26 germline mutation carriers, but not ETV6 mutation carriers, experience elevated risk for CH.

The Role of NKL Homeobox Genes in T-Cell Malignancies

Homeobox genes encode transcription factors controlling basic developmental processes. The homeodomain is encoded by the homeobox and mediates sequence-specific DNA binding and interaction with cofactors, thus operating as a basic regulatory platform. Similarities in their homeobox sequences serve to arrange these genes in classes and subclasses, including NKL homeobox genes. In accordance with their normal functions, deregulated homeobox genes contribute to carcinogenesis along with hematopoietic malignancies. We have recently described the physiological expression of eleven NKL homeobox genes in the course of hematopoiesis and termed this gene expression pattern NKL-code. Due to the developmental impact of NKL homeobox genes these data suggest a key role for their activity in the normal regulation of hematopoietic cell differentiation including T-cells. On the other hand, aberrant overexpression of NKL-code members or ectopical activation of non-code members has been frequently reported in lymphoid and myeloid leukemia/lymphoma, demonstrating their oncogenic impact in the hematopoietic compartment. Here, we provide an overview of the NKL-code in normal hematopoiesis and discuss the oncogenic role of deregulated NKL homeobox genes in T-cell malignancies.

CUX1 Deficiency Potentiates RAS Signaling to Drive Malignancy

-7/del(7q) is prevalent in high-risk myeloid neoplasms and frequently co-occurs with gain-of-function mutations in the RAS pathway. Herein, we identify a genetic interaction between RAS and the 7q-encoded transcription factor, CUX1, that encompasses hematopoietic malignancies and solid-tumors. Mice with both oncogenic NrasG12D and Cux1 knockdown developed accelerated myeloid malignancies with leukemic transformation. Oncogenic RAS imparts increased self-renewal on CUX1-deficient hematopoietic stem/progenitor cells (HSPCs). Reciprocally, CUX1 knockdown amplifies RAS signaling through reduction of negative regulators of RAS/PI3K signaling. Accordingly, NrasG12D;Cux1-knockdown HSPCs have heightened growth factor-sensitivity and downstream RAS pathway activation. Double mutant HSPCs were responsive to PIK3 or MEK inhibition. Similarly, low expression of CUX1 in primary AML samples correlates with sensitivity to the same inhibitors, suggesting a viable therapy for malignancies with CUX1 inactivation. This work demonstrates an unexpected convergence of an oncogene and tumor suppressor gene on the same pathway. Disclosures No relevant conflicts of interest to declare.

Benzyl Isothiocyanate, a Vegetable-Derived Compound, Induces Apoptosis via ROS Accumulation and DNA Damage in Canine Lymphoma and Leukemia Cells

Treatment of neoplastic diseases in companion animals is one of the most important problems of modern veterinary medicine. Given the growing interest in substances of natural origin as potential anti-cancer drugs, our goal was to examine the effectiveness of benzyl isothiocyanate (BITC), a compound found in cruciferous vegetables, against canine lymphoma and leukemia. These are the one of the most common canine cancer types, and chemotherapy is the only treatment option. The study involved established cell lines originating from various hematopoietic malignancies: CLBL-1, GL-1, CLB70 and CNK-89, immortalized noncancerous cell lines: MDCK and NIH-3T3 and canine peripheral blood mononuclear cells (PBMCs). The cytotoxic activity of BITC, apoptosis induction, caspase activity and ROS generation were evaluated by flow cytometry. H2AX phosphorylation was assessed by western blot. The study showed that the compound was especially active against B lymphocyte-derived malignant cells. Their death resulted from caspase-dependent apoptosis. BITC induced ROS accumulation, and glutathione precursor N-acetyl-l-cysteine reversed the effect of the compound, thus proving the role of oxidative stress in BITC activity. In addition, exposure to the compound induced DNA damage in the tested cells. This is the first study that provides information on the activity of BITC in canine hematopoietic malignancies and suggests that the compound may be particularly useful in B-cell neoplasms treatment.

Targeting cholesterol homeostasis in hematopoietic malignancies

Cholesterol is a vital lipid for cellular functions. It is necessary for membrane biogenesis, cell proliferation and differentiation. In addition to maintaining cell integrity and permeability, increasing evidence indicates a strict link between cholesterol homeostasis, inflammation and haematological tumors. This makes cholesterol homeostasis an optimal therapeutic target for hematopoietic malignancies. Manipulating cholesterol homeostasis either interfering with its synthesis or activating the reverse cholesterol transport via the engagement of liver X receptors (LXRs), affects the integrity of tumor cells both in vitro and in vivo. Cholesterol homeostasis has also been manipulated to restore antitumor immune responses in preclinical models. These observations have prompted clinical trials in acute myeloid leukemia (AML) to test the combination of chemotherapy with drugs interfering with cholesterol synthesis, i.e. statins. We review the role of cholesterol homeostasis in hematopoietic malignancies, as well as in cells of the tumor microenvironment, and discuss the potential use of lipid modulators for therapeutic purposes.

Immunotherapy in hematological malignancies: recent advances and open questions

Over recent years, tremendous advances in immunotherapy approaches have been observed, generating significant clinical progress. Cancer immunotherapy has been shown, in different types of blood cancers, to improve the overall survival of patients. Immunotherapy treatment of hematopoietic malignancies is a newly growing field that has been accelerating over the past years. Several US FDA approved drugs and cell-based therapies are being exploited in the late stage of clinical trials. This review attempt to highlight and discuss the numerous innovative immunotherapy approaches of hematopoietic malignancy ranging from nonmyeloablative transplantation, T-cell immunotherapy, natural killer cells and immune agonist to monoclonal antibodies and vaccination. In addition, a brief discussion on the future advances and accomplishments required to counterpart the current immunotherapeutic approaches for hematopoietic malignancies were also highlighted.

t(4;12)(q12;p13) ETV6-rearranged AML without eosinophilia does not involve PDGFRA: relevance for imatinib insensitivity

Acute myeloid leukemia (AML) with t(4;12)(q12;p13) translocation is rare, and often associated with an aggressive clinical course and poor prognosis. Previous reports based on fluorescence in-situ hybridization (FISH) analysis have suggested that ETV6-PDGFRA fusions are present in these patients despite the absence of eosinophilia, which is typically found in other hematopoietic malignancies with PDGFRA¬-containing fusions. We first detected an ETV6-SCFD2 fusion by targeted RNA sequencing in a patient with t(4;12)(q12;p13) who had previously been diagnosed with an ETV6-PDGFRA fusion by FISH analysis but failed to respond to imatinib. We then retrospectively identified four additional AML patients with t(4;12)(q12;p13) with apparent ETV6-PDGFRA fusions using chromosome and FISH analysis and applied targeted RNA sequencing to archival material. We again detected rearrangements between ETV6 and non-PDGFRA 4q12 genes including SCFD2, CHIC2 and GSX2. None of the three patients who received imatinib based on the incorrect assumption of an ETV6-PDGFRA fusion responded. Our findings highlight the importance of using a sequencing-based assay to confirm the presence of targetable gene fusions, particularly in genomic regions such as 4q12 with many clinically relevant genes that are too close to resolve by chromosome or FISH analysis. Finally, combining our data and review of the literature, we show that sequence-confirmed ETV6-PDGFRA fusions are typically found in eosinophilic disorders (3 of 3 cases), and patients with t(4;12)(q12;p13) without eosinophilia are found to have other 4q12 partners on sequencing (17 of 17 cases).

Pharmacological or genetic inhibition of hypoxia signaling attenuates oncogenic RAS-induced cancer phenotypes

Oncogenic Ras mutations are highly prevalent in hematopoietic malignancies. However, it is difficult to directly target oncogenic RAS proteins for therapeutic intervention. We have developed a Drosophila Acute Myeloid Leukemia (AML) model induced by human KRASG12V, which exhibits a dramatic increase in myeloid-like leukemia cells. We performed both genetic and drug screens using this model. The genetic screen identified 24 candidate genes able to attenuate the oncogenic RAS-induced phenotype, including two key hypoxia pathway genes HIF1A and ARNT (HIF1B). The drug screen revealed echinomycin, an inhibitor of HIF1A, could effectively attenuate the leukemia phenotype caused by KRASG12V. Furthermore, we showed that echinomycin treatment could effectively suppress oncogenic RAS-driven leukemia cell proliferation using both human leukemia cell lines and a mouse xenograft model. These data suggest that inhibiting the hypoxia pathway could be an effective treatment approach for oncogenic RAS-induced cancer phenotype, and that echinomycin is a promising targeted drug to attenuate oncogenic RAS-induced cancer phenotypes.

Targeting the actin nucleation promoting factor WASp provides a therapeutic approach for hematopoietic malignancies

Cancer cells depend on actin cytoskeleton rearrangement to carry out hallmark malignant functions including activation, proliferation, migration and invasiveness. Wiskott–Aldrich Syndrome protein (WASp) is an actin nucleation-promoting factor and is a key regulator of actin polymerization in hematopoietic cells. The involvement of WASp in malignancies is incompletely understood. Since WASp is exclusively expressed in hematopoietic cells, we performed in silico screening to identify small molecule compounds (SMCs) that bind WASp and promote its degradation. We describe here one such identified molecule; this WASp-targeting SMC inhibits key WASp-dependent actin processes in several types of hematopoietic malignancies in vitro and in vivo without affecting naïve healthy cells. This small molecule demonstrates limited toxicity and immunogenic effects, and thus, might serve as an effective strategy to treat specific hematopoietic malignancies in a safe and precisely targeted manner.