Thomas Loughran - Academia.edu (original) (raw)

Papers by Thomas Loughran

Multiphoton Microscopy in the Biomedical Sciences XIX, 2019

Self-sweeping of laser frequency is relatively new effect in fiber lasers. The effect consists in... more Self-sweeping of laser frequency is relatively new effect in fiber lasers. The effect consists in periodic dynamics of the laser frequency without use of tuning elements and electrical drivers for frequency tuning. Owing to broad sweeping range (up to 23 nm) and simplicity, self-sweeping fiber lasers are attractive sources for applications demanding tunable radiation. Currently the self-sweeping effect in fiber lasers was observed in different spectral regions covering range from 1 to 2.1 μm. However, it is difficult to control spectral dynamics due to self-induced nature of the sweeping effect. In the paper, we demonstrated linearly-polarized Tm-doped fiber laser with lasing near 1.9 μm with manually controlled the spectral dynamics with pump power adjustment. The laser operates in three self-sweeping regimes depending on pump power: 1) with normal scanning direction at high rate (~5 nm/sec) and, 2) with reverse one at low sweeping rate (~0.1 nm/sec) and 3) wavelength stopping. In the case of wavelength stopping, the wavelength can be stopped at arbitrary value in the range from 1912 to 1923 nm depending on prehistory of spectral dynamics of the laser. The wavelength stability in case of wavelength stopping is better than 50 pm within 5 minutes. In the case of linear scanning of laser line, sweeping range exceeds 15 nm.

Modifications in sphingolipid (SL) metabolism and mitochondrial bioenergetics are key factors imp... more Modifications in sphingolipid (SL) metabolism and mitochondrial bioenergetics are key factors implicated in cancer cell response to chemotherapy, including chemotherapy resistance. Vinca alkaloids such as vincristine (VCR), widely used in cancer treatment, are no exception, as their beneficial actions are often supplanted by resistance. In the present work we utilized HL-60 human leukemia cells and a VCR-resistant counterpart, HL-60/VCR, as a model to determine potential interplay between SL metabolism and mitochondrial bioenergetics supportive of multidrug resistance (MDR). Relative to wild-type cells, HL-60/VCR presented with global alterations in SL composition, typified by upregulated expression of sphingosine kinase (SPHK1), which catalyzes formation of sphingosine 1-phosphate (S1P), glucosylceramide synthase (GCS), which catalyzes formation of glucosylceramides (GC), and acid ceramidase, responsible for ceramide hydrolysis. In support of these changes, VCR resistance was also ...

Journal of Lipid Research, 2019

Acute myeloid leukemia (AML) is the most common acute leukemia in adults. More than half of older... more Acute myeloid leukemia (AML) is the most common acute leukemia in adults. More than half of older AML patients fail to respond to cytotoxic chemotherapy, and most responders relapse with drug-resistant disease. Failure to achieve complete remission can be partly attributed to the drug resistance advantage of AML blasts that frequently express P-glycoprotein (P-gp), an ATP-binding cassette transporter. Our previous work showed that elevated acid ceramidase (AC) levels in AML contribute to blast survival. Here, we investigated P-gp expression levels in AML relative to AC. Using parental HL-60 cells and drug-resistant derivatives as our model, we found that P-gp expression and efflux activity were highly upregulated in resistant derivatives. AC overexpression in HL-60 conferred resistance to the AML chemotherapeutic drugs, cytarabine, mitoxantrone, and daunorubicin, and was linked to P-gp upregulation. Furthermore, targeting AC through pharmacologic or genetic approaches decreased P-gp...

Blood, Jan 26, 2018

Large granular lymphocyte (LGL) leukemia results from clonal expansion of CD3 cytotoxic T-lymphoc... more Large granular lymphocyte (LGL) leukemia results from clonal expansion of CD3 cytotoxic T-lymphocytes or CD3 natural killer (NK) cells. Chronic antigen stimulation is postulated to promote long-term survival of LGL leukemia cells through constitutive activation of multiple survival pathways, resulting in global dysregulation of apoptosis and resistance to activation-induced cell death. We reported previously that nuclear factor kappa B (NF-κB) is a central regulator of the survival network for leukemic LGL. However, the mechanisms that trigger constitutive activation of NF-κB in LGL leukemia remain undefined. TNF-related apoptosis-inducing ligand (TRAIL) is known to induce apoptosis in tumor cells, but can also activate NF-κB through interaction with TRAIL receptors 1, 2 and 4 (also known as DR4, DR5 and DcR2 respectively). The role of TRAIL has not been studied in LGL leukemia. In this study, we hypothesized that TRAIL interaction with DcR2 contributes to NF-κB activation in LGL le...

Signal transduction and targeted therapy, 2017

The pathogenesis of chronic lymphocytic leukemia (CLL) is poorly understood and it remains incura... more The pathogenesis of chronic lymphocytic leukemia (CLL) is poorly understood and it remains incurable with current therapies. We have previously shown that nanoliposomal C6-ceramide (CNL) is an effective therapy in anmurine model of CLL. However, the key signaling pathways mediating CNL-induced cell death in CLL remains unknown. We hypothesized that CNL targets STAT3, a critical regulator of hematopoietic biology. We observed that CNL treatment reduced phosphorylated STAT3 at both Y705 and S727 residues in CLL cell lines and patient cells. This, in turn, reduced STAT3 transcriptional activity and expression of critical STAT3-dependent survival factors like Mcl-1 and survivin. The effect of CNL on STAT3 was further confirmedas shown by reduced STAT3 phosphorylation in xenograft tumors obtained from mice treated with CNL. CNL suppressed STAT3 phosphorylation at Y705 and S727 through reduction in BTK activity and MEK1/2 kinase/PKC activities, respectively. Moreover, a synergistic reduct...

Cancer Translational Medicine, 2017

Aim-To further characterize the selectivity, mechanism-of-action and therapeutic efficacy of the ... more Aim-To further characterize the selectivity, mechanism-of-action and therapeutic efficacy of the novel small molecule inhibitor, SKI-178. Methods-Using the state-of-the-art Cellular Thermal Shift Assay (CETSA) technique to detect "direct target engagement" of proteins intact cells, in vitro and in vivo assays, pharmacological assays and multiple mouse models of acute myeloid leukemia (AML). Results-Herein, we demonstrate that SKI-178 directly target engages both Sphingosine Kinase 1 and 2. We also present evidence that, in addition to its actions as a Sphingosine Kinase Inhibitor, SKI-178 functions as a microtubule network disrupting agent both in vitro and in intact cells. Interestingly, we separately demonstrate that simultaneous SphK inhibition and microtubule disruption synergistically induces apoptosis in AML cell lines. Furthermore, we demonstrate that SKI-178 is well tolerated in normal healthy mice. Most importantly, we demonstrate that SKI-178 has therapeutic efficacy in several mouse models of AML. Conclusion-SKI-178 is a multi-targeted agent that functions both as an inhibitor of the SphKs as well as a disruptor of the microtubule network. SKI-178 induced apoptosis arises from a synergistic interaction of these two activities. SKI-178 is safe and effective in mouse models of AML, supporting its further development as a multi-targeted anti-cancer therapeutic agent.

Biochemical pharmacology, Apr 15, 2017

The anticancer properties of ceramide, a sphingolipid with potent tumor-suppressor properties, ca... more The anticancer properties of ceramide, a sphingolipid with potent tumor-suppressor properties, can be dampened via glycosylation, notably in multidrug resistance wherein ceramide glycosylation is characteristically elevated. Earlier works using the ceramide analog, C6-ceramide, demonstrated that the antiestrogen tamoxifen, a first generation P-glycoprotein (P-gp) inhibitor, blocked C6-ceramide glycosylation and magnified apoptotic responses. The present investigation was undertaken with the goal of discovering non-anti-estrogenic alternatives to tamoxifen that could be employed as adjuvants for improving the efficacy of ceramide-centric therapeutics in treatment of cancer. Herein we demonstrate that the tamoxifen metabolites, desmethyltamoxifen and didesmethyltamoxifen, and specific, high-affinity P-gp inhibitors, tariquidar and zosuquidar, synergistically enhanced C6-ceramide cytotoxicity in multidrug resistant HL-60/VCR acute myelogenous leukemia (AML) cells, whereas the selective...

Oncotarget, Jan 4, 2016

There is an urgent unmet need for new therapeutics in acute myeloid leukemia (AML) as standard th... more There is an urgent unmet need for new therapeutics in acute myeloid leukemia (AML) as standard therapy has not changed in the past three decades and outcome remains poor for most patients. Sphingolipid dysregulation through decreased ceramide levels and elevated sphingosine 1-phosphate (S1P) promotes cancer cell growth and survival. Acid ceramidase (AC) catalyzes ceramide breakdown to sphingosine, the precursor for S1P. We report for the first time that AC is required for AML blast survival. Transcriptome analysis and enzymatic assay show that primary AML cells have high levels of AC expression and activity. Treatment of patient samples and cell lines with AC inhibitor LCL204 reduced viability and induced apoptosis. AC overexpression increased the expression of anti-apoptotic Mcl-1, significantly increased S1P and decreased ceramide. Conversely, LCL204 induced ceramide accumulation and decreased Mcl-1 through post-translational mechanisms. LCL204 treatment significantly increased ov...

Journal of Lipid Research, 2016

The bioactive, tumor suppressor sphingolipid ceramide (1-3), which plays a central role in initia... more The bioactive, tumor suppressor sphingolipid ceramide (1-3), which plays a central role in initiating cancer cell death (4, 5), can be of unique utility from a therapeutic standpoint (6). Many drugs used in the treatment of cancer are themselves ceramide generators, a property that contributes in part to their apoptosis-inducing effects (7). Once generated, however, cancer cells can convert ceramide to higher sphingolipids, notably glucosylceramide (GC), blunting ceramide's anticancer benefits (1, 8). As upregulated ceramide glycosylation is linked with multidrug resistance, limiting ceramide glycosylation would appear to be an appropriate intervention, a scenario that has been demonstrated in past works (9, 10). In addition to glycosylation, ceramide metabolism via hydrolysis gives rise to sphingosine 1-phosphate (S1-P), a mitogenic sphingolipid that competes with ceramide's proapoptotic effects (11, 12). Acute myelogenous leukemia (AML) is the most common type of leukemia in adults. Only 25% of patients who experience remission with cytotoxic chemotherapy remain disease free. Therapy for these patients generally consists of cytosine arabinoside plus anthracycline, a regimen that has been in use for more than three decades; thus, there exists a critical need to develop more effective therapies in AML. One novel approach involves the use of C6-ceramide, a ceramide mimic, which like its long-chain, natural counterpart has tumor-suppressor properties. An

Leukemia Research, 2015

Fenretinide, N-(4-hydroxyphenyl)retinamide, (4-HPR), a synthetic retinoid, owes its cancer-toxic ... more Fenretinide, N-(4-hydroxyphenyl)retinamide, (4-HPR), a synthetic retinoid, owes its cancer-toxic effects in part to the generation of ceramide, a potent tumor-suppressing sphingolipid. As such, 4-HPR has garnered considerable interest as a chemotherapeutic. Cancer cells, however, via various metabolic routes, inactivate ceramide, and this can limit 4-HPR efficacy. As relatively little is known regarding 4-HPR-induced ceramide management in acute myelogeneous leukemia (AML), we undertook the present study to evaluate the impact of 4-HPR on ceramide production, metabolism, and cytotoxicity. In KG-1, HL-60, and HL-60/VCR (multidrug resistant) human leukemia cells, 4-HPR induced 15-, 2-, and 20-fold increases in ceramide (measured using [ 3 H]palmitic acid), respectively. By use of specific inhibitors we show that ceramide was produced by sphingomyelinase and de novo pathways in response to 4-HPR exposure. HL-60/VCR cells metabolized ceramide to glucosylceramide (GC). 4-HPR exposure (1.25-10 μM) reduced viability in all cell lines, with approximate IC50's ranging from 1-8.0 μM. Reactive oxygen species (ROS) were generated in response to 4-HPR treatment, and the concomitant cytotoxicity was reversed by addition of vitamin E. 4-HPR was not cytotoxic nor did it elicit ceramide formation in K562, a chronic myeloid leukemia cell line; however, K562 cells were sensitive to a cell-deliverable form of ceramide, C6-ceramide. Treatment of Molt-3, an acute lymphoblastic leukemia cell line, with 4-HPR revealed moderate ceramide production (5-fold over control), robust conversion of ceramide to GC and sphingomyelin, and resistance to 4-HPR and

Cancer biology & therapy, Jan 7, 2015

Sphingolipid metabolism has been identified as a potential therapeutic target in cancer. Sphingos... more Sphingolipid metabolism has been identified as a potential therapeutic target in cancer. Sphingosine-1-phosphate (S1P) is a potent bioactive sphingolipid metabolite produced by sphingosine kinases-1 and -2 (SPHK1 and SPHK2). Elevated SPHK1 has been found in numerous cancer types and been shown to contribute to survival, chemotherapeutic resistance and malignancy. However, its role in large granular Natural Killer (NK)-lymphocyte (LGL) leukemia has not been investigated. Here, we examine SPHK1 as a therapeutic target in LGL leukemia. We found that SPHK1 is overexpressed in peripheral blood mononuclear cells (PBMCs) from LGL leukemia patients which results in elevated S1P in the sera. The use of SPHK1 inhibitors, SKI-II or SKI-178, decreased leukemic NK cell viability and induced caspase-dependent apoptosis. SKI-II and SKI-178 restored the sphingolipid balance by increasing ceramide and decreasing S1P in leukemic NKL cells. SKI-II and SKI-178 also induced apoptosis in primary NK-LGLs ...

Hematology / the Education Program of the American Society of Hematology. American Society of Hematology. Education Program, 2012

Large granular lymphocyte (LGL) leukemia represents a spectrum of rare lymphoproliferative diseas... more Large granular lymphocyte (LGL) leukemia represents a spectrum of rare lymphoproliferative diseases defined by clonal amplification of either CD3(+) cytotoxic T-lymphocytes or CD3(-) natural killer cells. This chapter focuses on the T-cell form of LGL leukemia. Clinical features include neutropenia, anemia, and rheumatoid arthritis. LGL leukemia is thought to arise from chronic antigenic stimulation, with the long-term survival of LGL being promoted by constitutive activation of multiple survival signaling pathways, such as the JAK/STAT3, sphingolipid, and Ras/MEK/ERK pathways. Therefore, these lead to global deregulation of apoptosis and resistance to normal pathways of activation-induced cell death. The majority of LGL leukemia patients eventually need treatment. Treatment of leukemic LGL is based on immunosuppressive therapy, primarily using low doses of methotrexate or cyclophosphamide. However, no standard therapy has been established because of the lack of large, prospective t...

Biochimica et biophysica acta, Jan 10, 2015

The triphenylethylene antiestrogen, tamoxifen, can be an effective inhibitor of sphingolipid meta... more The triphenylethylene antiestrogen, tamoxifen, can be an effective inhibitor of sphingolipid metabolism. This off-target activity makes tamoxifen an interesting ancillary for boosting the apoptosis-inducing properties of ceramide, a sphingolipid with valuable tumor censoring activity. Here we show for the first time that tamoxifen and metabolite, N -desmethyltamoxifen (DMT) block ceramide glycosylation and inhibit ceramide hydrolysis (by acid ceramidase, AC) in human acute myelogenous leukemia (AML) cell lines and in AML cells derived from patients. Tamoxifen (1-10μM) inhibition of AC in AML cells was accompanied by decreases in AC protein expression. Tamoxifen also depressed expression and activity of sphingosine kinase 1 (SphK1), the enzyme catalyzing production of mitogenic sphingosine 1-phosphate (S1-P). Results from mass spectroscopy showed that tamoxifen and DMT, i ) increased the levels of endogenous C16:0- and C24:1 ceramide molecular species, ii) nearly totally halted produ...

Cancer Biology & Therapy, 2014

The Journal of pharmacology and experimental therapeutics, 2015

We previously developed SKI-178 (N'-[(1E)-1-(3,4-dimethoxyphenyl)ethylidene]-3-(4-methoxxyphe... more We previously developed SKI-178 (N'-[(1E)-1-(3,4-dimethoxyphenyl)ethylidene]-3-(4-methoxxyphenyl)-1H-pyrazole-5-carbohydrazide) as a novel sphingosine kinase-1 (SphK1) selective inhibitor and, herein, sought to determine the mechanism-of-action of SKI-178-induced cell death. Using human acute myeloid leukemia (AML) cell lines as a model, we present evidence that SKI-178 induces prolonged mitosis followed by apoptotic cell death through the intrinsic apoptotic cascade. Further examination of the mechanism of action of SKI-178 implicated c-Jun NH2-terminal kinase (JNK) and cyclin-dependent protein kinase 1 (CDK1) as critical factors required for SKI-178-induced apoptosis. In cell cycle synchronized human AML cell lines, we demonstrate that entry into mitosis is required for apoptotic induction by SKI-178 and that CDK1, not JNK, is required for SKI-178-induced apoptosis. We further demonstrate that the sustained activation of CDK1 during prolonged mitosis, mediated by SKI-178, lead...

PLoS ONE, 2013

Ceramide is a sphingolipid metabolite that induces cancer cell death. When C6-ceramide is encapsu... more Ceramide is a sphingolipid metabolite that induces cancer cell death. When C6-ceramide is encapsulated in a nanoliposome bilayer formulation, cell death is selectively induced in tumor models. However, the mechanism underlying this selectivity is unknown. As most tumors exhibit a preferential switch to glycolysis, as described in the "Warburg effect", we hypothesize that ceramide nanoliposomes selectively target this glycolytic pathway in cancer. We utilize chronic lymphocytic leukemia (CLL) as a cancer model, which has an increased dependency on glycolysis. In CLL cells, we demonstrate that C6-ceramide nanoliposomes, but not control nanoliposomes, induce caspase 3/7independent necrotic cell death. Nanoliposomal ceramide inhibits both the RNA and protein expression of GAPDH, an enzyme in the glycolytic pathway, which is overexpressed in CLL. To confirm that ceramide targets GAPDH, we demonstrate that downregulation of GAPDH potentiates the decrease in ATP after ceramide treatment and exogenous pyruvate treatment as well as GAPDH overexpression partially rescues ceramide-induced necrosis. Finally, an in vivo murine model of CLL shows that nanoliposomal C6-ceramide treatment elicits tumor regression, concomitant with GAPDH downregulation. We conclude that selective inhibition of the glycolytic pathway in CLL cells with nanoliposomal C6-ceramide could potentially be an effective therapy for leukemia by targeting the Warburg effect.

Leukemia & Lymphoma, 2012

Natural killer cell leukemia is characterized by clonal expansion of CD3 − NK cells and comprises... more Natural killer cell leukemia is characterized by clonal expansion of CD3 − NK cells and comprises both chronic and aggressive forms. Currently, no effective treatment exists, thus providing a need for identification of novel therapeutics. Lipidomic studies revealed dysregulated sphingolipid metabolism as evidenced by decreased levels of overall ceramide species and increased levels of cerebrosides in leukemic NK cells, concomitant with increased glucosylceramide synthase (GCS) expression. GCS, a key enzyme of this pathway, neutralizes pro-apoptotic ceramide by transfer of a UDP-glucose. Thus, we treated both rat and human leukemic NK cells in combination with: 1) exogenous C 6-ceramide nanoliposomes in order to target mitochondria and increase physiological pro-apoptotic levels of long chain ceramide, and 2) 1-phenyl-2-palmitoylamino-3-morpholino-1propanol (PPMP), an inhibitor of GCS. Co-administration of C 6-ceramide nanoliposomes and PPMP elicited an increase in endogenous long-chain ceramide species, which led to cellular apoptosis in a synergistic manner via the mitochondrial intrinsic cell death pathway in leukemic NK cells.

Leukemia & Lymphoma, 2011

Large granular lymphocyte (LGL) leukemia is a rare disorder of cytotoxic lymphocytes. LGL cells p... more Large granular lymphocyte (LGL) leukemia is a rare disorder of cytotoxic lymphocytes. LGL cells play an integral role in the immune system and are divided into two major lineages of CD3− natural killer (NK) cells and CD3+ T cells that circulate throughout the blood in search of infected cells, in which they will make contact through a receptor ligand and induce cell death. LGLs cells are also programmed to undergo apoptosis after contact with an infected target cell; however they continue to survive in individuals with LGL leukemia. This unchecked proliferation and cytotoxicity of LGLs in patients results in autoimmunity or malignancy. Rheumatoid arthritis is the most common autoimmune condition seen in individuals with LGL leukemia; however, LGL leukemia is associated with a wide spectrum of other autoimmune diseases. Patients may also suffer from other hematological conditions including hemolytic anemia, pure red cell aplasia, and neutropenia which lead to recurrent bacterial infections. Currently, the only established treatment involves a low dose of an immunosuppressive regimen with methotrexate, in which 40-50% of patients are either resistant or do not respond. In order to establish new therapeutics it is important to understand the current state of LGL leukemia both in clinic and in basic research.

Future Oncology, 2012

Large granular lymphocyte (LGL) leukemia is a clonal lymphoproliferative disorder of cytotoxic ly... more Large granular lymphocyte (LGL) leukemia is a clonal lymphoproliferative disorder of cytotoxic lymphocytes characterized by an expansion of CD3+cytotoxic T lymphocytes or CD3-natural killer cells. Patients present with various cytopenias including neutropenia, anemia and thrombocytopenia. In addition, there is an association of T-cell large granular lymphocytic leukemia with rheumatoid arthritis. It is believed that LGL leukemia begins as an antigen-driven immune response with subsequent constitutive activation of cytotoxic T lymphocytes or natural killer cells through PDGF and IL-15 contributing to their survival. Consequently, this leads to a dysregulation of apoptosis and dysfunction of the activation-induced cell death pathway. Treatment of LGL leukemia is based on a low-dose immunosuppressive regimen using methotrexate or cyclophosphamide. However, no standard of therapy has been established, as large prospective trials have not been conducted. In addition, some patients are re...