Sphingolipids and acid ceramidase as therapeutic targets in cancer therapy.

BACKGROUND: Sphingolipids have been shown to play a key part in cancer cell growth and death and have increasingly become the subject of novel anti-cancer therapies. Acid ceramidase, a sphingolipid enzyme, has an important role in the regulation of apoptosis. In this review we aim to assess the current evidence supporting the role of sphingolipids in cancer and the potential role that acid ceramidase may play in cancer treatment. METHODS: A literature search was performed for published full text articles using the PubMed, Cochrane and Scopus databases using the search criteria string "acid ceramidase", "sphingolipid", "cancer". Additional papers were detected by scanning the references of relevant papers. A summary of the evidence for each cancer subgroup was then formed. Given the nature of the data extracted, no meta-analysis was performed. RESULTS: Over expression of acid ceramidase has been demonstrated in a number of human cancers. In vitro data demonstrate that manipulation of acid ceramidase may present a useful therapeutic target. In the clinical setting, a number of drugs have been investigated with the ability to target acid ceramidase, with the most promising of those being small molecular inhibitors, such as LCL521. CONCLUSION: The role of the sphingolipid pathway in cancer is becoming very clearly established by promoting ceramide accumulation in response to cancer or cellular stress. Acid ceramidase is over expressed in a variety of cancers and has a role as a potential target for inhibition by novel specific inhibitors or off-target effects of traditional anti-cancer agents. Further work is required to develop acid ceramidase inhibitors safe for progression to clinical trials.

Systematic review of treatment intensification using novel agents for chemoradiotherapy in rectal cancer.

BACKGROUND: With the well established shift to neoadjuvant treatment for locally advanced rectal cancer, there is increasing focus on the use of radiosensitizers to improve the efficacy and tolerability of radiotherapy. There currently exist few randomized data exploring novel radiosensitizers to improve response and it is unclear what the clinical endpoints of such trials should be. METHODS: A qualitative systematic review was performed according to the PRISMA guidelines using preset search criteria across the PubMed, Cochrane and Scopus databases from 1990 to 2017. Additional results were generated from the reference lists of included papers. RESULTS: A total of 123 papers were identified, of which 37 were included; a further 60 articles were obtained from additional referencing to give a total of 97 articles. Neoadjuvant radiosensitization for locally advanced rectal cancer using fluoropyrimidine-based chemotherapy remains the standard of treatment. The oral derivative capecitabine has practical advantages over 5-fluorouracil, with equal efficacy, but the addition of a second chemotherapeutic agent has yet to show a consistent significant efficacy benefit in randomized clinical assessment. Preclinical and early-phase trials are progressing with promising novel agents, such as small molecular inhibitors and nanoparticles. CONCLUSION: Despite extensive research and promising preclinical studies, a definite further agent in addition to fluoropyrimidines that consistently improves response rate has yet to be found.

Proteomic profiling of rectal cancer reveals acid ceramidase is implicated in radiation response.

BACKGROUND: Neoadjuvant chemoradiotherapy (CRT) is used in locally advanced rectal cancer when tumours threaten the circumferential resection margin, with varying response to treatment. This experimental study aimed to identify significantly differentially expressed proteins between patients responding and not responding to CRT, and to validate any proteins of interest. METHODS: Mass spectrometry (with isobaric tagging for relative quantification) analysis of rectal cancers pre- and post-CRT, and at resection. Validation of proteins of interest was performed by assessing tissue microarray (TMA) immunohistochemistry expression in a further 111 patients with rectal cancer. RESULTS: Proteomic data are available via ProteomeXchange with identifier PXD008436. Reduced abundance of contributing peptide ions for acid ceramidase (AC) (log fold change -1.526, p = 1.17E-02) was observed in CRT responders. Differential expression of AC was confirmed upon analysis of the TMAs. Cancer site expression of AC in stromal cells from post-CRT resection specimens was observed to be relatively low in pathological complete response (p = 0.003), and relatively high with no response to CRT (p = 0.017). CONCLUSION: AC may be implicated in the response of rectal cancer to CRT. We propose its further assessment as a novel potential biomarker and therapeutic target. SIGNIFICANCE: There is a need for biomarkers to guide the use of chemoradiotherapy in rectal cancer, as none are in routine clinical use. We have determined acid ceramidase may have a role in radiation response, based on novel proteomic profiling and validation in a wider dataset using tissue microarrays. The ability to predict or improve response would positively select those patients who will derive benefit, prevent delays in the local and systemic management of disease in non-responders, and reduce morbidity associated with chemoradiotherapy.

Regulation of DNA repair by ubiquitylation.

Cellular DNA repair is a frontline system that is responsible for maintaining genome integrity and thus preventing premature aging and cancer by repairing DNA lesions and strand breaks caused by endogenous and exogenous mutagens. However, it is also the principal cellular system in cancer cells that counteracts the killing effect of the major cancer treatments, e.g. chemotherapy and ionizing radiation. Although it is clear that an individual's DNA repair capacity varies, the mechanisms involved in the regulation of repair systems that are responsible for such variations are only just emerging. This knowledge gap is impeding the finding of new cancer therapy targets and the development of novel treatment strategies. In recent years the vital role of post-translational modifications of DNA repair proteins, including ubiquitylation and phosphorylation, has been uncovered. This review will cover recent progress in our understanding of the role of ubiquitylation in the regulation of DNA repair.

The multiple influences of glutathione on bromate genotoxicity: implications for the dose-response relationship.

Glutathione displays multiple roles in the oxidative genotoxicity of potassium bromate. On the one hand, reduced glutathione has a demonstrated role in the activation of bromate to species capable of oxidising DNA. However, if this activation should occur within the gut or extracellularly once bromate is absorbed, this may limit the ability of the chemical to oxidise cellular DNA in vivo. Moreover, glutathione may offer protection against the damaging species produced by its interaction with bromate. Finally, if bromate exposure of cells is sufficiently high to deplete glutathione, a secondary oxidative stress and associated DNA damage may occur. These observations would suggest non-linearity in the dose-response to DNA damage in vivo.

Monitoring base excision repair proteins on damaged DNA using human cell extracts.

BER (base excision repair) is a major pathway for the removal of simple lesions in DNA including base damage and base loss (abasic site). We have developed an assay, using formaldehyde cross-linking during repair in human cell extracts, to observe BER proteins involved in the repair of damaged DNA. This approach allows visualization of repair proteins on damaged DNA during BER in human cell extracts and provides a detailed view of the molecular events leading to repair.

The role of glutathione in DNA damage by potassium bromate in vitro.

We have investigated the role of reduced glutathione (GSH) in the genetic toxicity of the rodent renal carcinogen potassium bromate (KBrO(3)). A statistically significant increase in the concentration of 8-oxodeoxyguanosine (8-oxodG) relative to deoxyguanosine was measured following incubation of calf thymus DNA with KBrO(3) and GSH or N-acetylcysteine (NACys). This was dependent on these thiols and was associated with the loss of GSH and production of oxidized glutathione. A short-lived (<6 min) intermediate was apparent which did not react with the spin trap dimethylpyrroline N-oxide. DNA oxidation was not evident when potassium chlorate (KClO(3)) or potassium iodate (KIO(3)) were used instead of KBrO(3), though GSH depletion also occurred with KIO(3), but not with KClO(3). Other reductants and thiols in combination with KBrO(3) did not cause a significant increase in DNA oxidation. DNA strand breakage was also induced by KBrO(3) in human white blood cells (5 mM) and rat kidney epithelial cells (NRK-52E, 1.5 mM). This was associated with an apparent small depletion of thiols in NRK-52E cells at 15 min and with an elevation of 8-oxodG at a delayed time of 24 h. Depletion of intra-cellular GSH by diethylmaleate in human lymphocytes decreased the amount of strand breakage induced by KBrO(3). Extracellular GSH, however, protected against DNA strand breakage by KBrO(3), possibly due to the inability of the reactive product to enter the cell. In contrast, membrane-permeant NACys enhanced KBrO(3)-induced DNA strand breakage in these cells. DNA damage by KBrO(3) is therefore largely dependent on access to intracellular GSH.

Interplanetary crew dose rates for the August 1972 solar particle event.

Parsons, J. L. and Townsend, L. W. Interplanetary Crew Dose Rates for the August 1972 Solar Particle Event. Using the coupled neutron-proton space radiation transport computer code (BRYNTRN), estimates of dose rates of protons in the skin, ocular lens and bone marrow, behind various thicknesses of aluminum shielding, for crews on space missions outside the Earth's magnetosphere, are made for the large solar particle event (SPE) of August 1972. Overall, the August 1972 dose rates are significantly higher than those estimated for any of the events that occurred in August-December 1989. The dose rates in the August 1972 SPE are not low dose rates as specified by the major national and international advisory bodies and committees.

Loss of expression of Dpc4 in pancreatic intraepithelial neoplasia: evidence that DPC4 inactivation occurs late in neoplastic progression.

Infiltrating adenocarcinomas of the pancreas are believed to arise from histologically identifiable intraductal precursors [pancreatic intraepithelial neoplasias (PanINs)] that undergo a series of architectural, cytological, and genetic changes. The role of DPC4 tumor suppressor gene inactivation in this progression has not been defined. Immunohistochemistry for the Dpc4 protein in formalin-fixed, paraffin-embedded tissue is a sensitive and specific marker for DPC4 gene status, providing a tool to examine DPC4 status in these putative precursor lesions. A total of 188 PanINs were identified in 40 pancreata, 38 (95%) of which also contained an infiltrating adenocarcinoma. Sections containing these 188 duct lesions were labeled with a monoclonal antibody to Dpc4. All 82 flat (PanIN-1A), all 54 papillary (PanIN-1B), and all 23 atypical papillary (PanIN-2) intraductal lesions expressed Dpc4. In contrast, 9 of 29 (31%) severely atypical lesions (PanIN-3 lesions, carcinomas in situ) did not. The difference in Dpc4 expression between histologically low-grade (PanIN-1 and -2) and histologically high-grade (PanIN-3) duct lesions was statistically significant (P < 0.0001). In three cases, the pattern of Dpc4 expression in the PanIN-3 lesions did not match the pattern of expression in the associated infiltrating carcinomas, indicating that these high-grade lesions did not simply represent infiltrating carcinoma growing along benign ducts. Loss of Dpc4 expression occurs biologically late in the neoplastic progression that leads to the development of infiltrating pancreatic cancer, at the stage of histologically recognizable carcinoma.

DNA oxidation by potassium bromate; a direct mechanism or linked to lipid peroxidation?

Following incubation of calf thymus DNA with potassium bromate (KBrO3) and glutathione (GSH), a statistically significant increase in the concentration of 8-oxodeoxyguanosine (8-oxodG) relative to deoxyguanosine was measured. This was GSH-dependent and was associated with loss of GSH during incubation. In contrast, 8-oxodG was not found to be elevated significantly in either total tissue DNA or mitochondrial DNA isolated from Sprague-Dawley rat kidney perfused in situ with KBrO3 (5 mM) for 15 min or 1 h. There was also no associated increase in the level of renal lipid peroxidation or reduced or oxidised GSH. Following intraperitoneal administration of KBrO3 to Sprague-Dawley rats, a dose of 100 mg/kg (maximum tolerated) gave evidence for oxidative stress in the kidney at 24 h as indicated by a significant increase in lipid peroxidation (P < 0.05) and oxidised GSH (P < 0.05). This was associated with a greater than 2-fold, significant (P < 0.01) increase in the level of 8-oxodG in kidney total DNA and a 57% (not statistically significant) increase in kidney mitochondrial 8-oxodG. Pretreatment of rats with diethylmaleate (DEM) to deplete GSH, elevated the toxicity of 100 mg/kg KBrO3. However, at a dose of 20 mg/kg, no change in any of the parameters indicative of kidney oxidative stress (including indicators of oxidative DNA damage; 8-oxodG or etheno-DNA adducts, which can be produced by lipid peroxides) was seen either with or without DEM pretreatment with the exception of a small but statistically significant (P < 0.05) increase in mitochondrial 8-oxodG when KBrO3 was given following DEM pretreatment. DNA oxidation in the kidney is therefore not inhibited by GSH depletion (contrasting with in vitro findings) and requires a sustained exposure at a near-toxic concentration of KBrO3 which is associated with lipid peroxidation and GSH oxidation. The results do not support a role, in rat kidney, of a direct, GSH-mediated mechanism for KBrO3-induced DNA oxidation as seen in vitro.

Temporal trends in antibody production in captive grey, harp and hooded seals to a single administration immunocontraceptive vaccine.

The temporal production of antibody to a single-administration immunocontraceptive vaccine, known to be immunocontraceptive in free-ranging female grey seals (Halichoerus grypus), was studied in captive grey seals, harp seals (Phoca groenlandica) and hooded seals (Cystophora cristata). The vaccine is based on liposome delivery of porcine zona pellucida antigens. When measured by antigen capture, the response of hooded and harp seals to the vaccine was similar to the response of grey seals. Determination of antibody production by ELISA with protein A, ELISA with rabbit anti-seal immunoglobulin sera and SDS-PAGE after affinity chromatography confirmed the similarity in response to the vaccine by grey and harp seals, but suggested lower titers in hooded seals. The vaccine produced titers in captive, juvenile grey and harp seals known to be immunocontraceptive in wild, adult grey seals.

Evidence for a long-lasting single administration contraceptive vaccine in wild grey seals.

A single-administration birth control vaccine based on liposome delivery of porcine zona pellucida antigens reduced pup production in grey seals (Halichoerus grypus) by about 90%. Anti-porcine zona pellucida titers of individual seals with two or more recaptures were variable but without a diminishing trend during the 5 year post-immunization period. Seals that produced at least one or more pups during the 2-5 year post-immunization period when the vaccine is fully effective, had an average anti-porcine zona pellucida titer of 5% of the reference serum. In contrast, the subset of seals that did not reproduce but were recaptured during the breeding season had an average titer of 31% of the reference serum. As measured by antibody titers and pup production, there were no differences in efficacy of the vaccine in 14-, 20- and 21-year-old female grey seals.

Multiple cutaneous granular cell tumors in a child with possible neurofibromatosis.

A 4-year-old girl with possible neurofibromatosis I had multiple subcutaneous nodules. On histopathologic examination, these nodules were diagnosed as multiple cutaneous granular cell tumors. An association between these tumors and neurofibromatosis I has been suggested because of their common neuroectodermal origin. We review cases of multiple cutaneous granular cell tumors in association with neurofibromatosis I in childhood.

Use of electroejaculation to collect semen samples from wild seals.

Semen samples were obtained from 7 adult male gray seals (Halichoerus grypus) at a colony on Sable Island, Nova Scotia, Canada, using a portable electroejaculation system. Males were anesthetized with tiletamine-zolazepam. A probe was inserted in the rectum, and a progressive series of 10 to 15 stimulations (1 to 9 V, 25 to 750 mA) was applied. Males ejaculated after 15 to 29 minutes of stimulation and produced ejaculates with volumes of 4 to 17 ml that had vigorous, motile spermatozoa. Although trials were performed in winter conditions (0 to -15 C), simple storage methods (semen samples collected into plastic bags and placed in a warm pocket or an insulated cooler) kept spermatozoa motile for up to 4 hours after ejaculation. The procedure described here provides a means for collection of semen samples for genetic analyses and tests of reproductive competence in seals.

Neurologic disease in a child with hepatoerythropoietic porphyria.

Hepatoerythropoietic porphyria (HEP) is a rare, autosomal recessive disorder due to deficient uroporphyrinogen decarboxylase enzyme activity. Patients exhibit photosensitivity, red urine, hypertrichosis, and characteristic serum and urine porphyrin profiles. Two siblings had the classic clinical and biochemical findings of HEP. The older patient developed a left-sided hemiparesis accompanied by an abnormal brain magnetic resonance imaging study. Although central nervous system abnormalities are a common feature of other hepatic porphyrias, they have not been previously documented in association with HEP.

Synthesis of 3-deazaneplanocin A, a powerful inhibitor of S-adenosylhomocysteine hydrolase with potent and selective in vitro and in vivo antiviral activities.

The neplanocin A analogue 3-deazaneplanocin A (2b) has been synthesized. A direct SN2 displacement on the cyclopentenyl mesylate 3 by the sodium salt of 6-chloro-3-deazapurine afforded the desired regioisomer 4 as the major product. After deprotection, this material was converted to 3-deazaneplanocin A in two steps. X-ray crystallographic analysis confirmed the assigned structure. Consistent with its potent inhibition of S-adenosylhomocysteine hydrolase, 3-deazaneplanocin A displayed excellent antiviral activity in cell culture against vesicular stomatitis, parainfluenza type 3, yellow fever, and vaccinia viruses. Antiviral activity was also displayed in vivo against vaccinia virus by using a mouse tailpox assay. The significantly lower cytotoxicity of 3-deazaneplanocin A, relative to its parent compound neplanocin A, may be due to its lack of conversion to 5'-triphosphate and S-adenosylmethionine metabolites.

Cytotoxic activity of enriched large granular lymphocyte populations in rheumatoid arthritis.

Enriched large granular lymphocyte (LGL) populations and mononuclear cell populations from peripheral blood of rheumatoid arthritis (RA) patients and control individuals were compared for natural killer (NK) cell activity. The NK activity of the two study groups was very similar when mean values for both cell populations were analyzed. When comparing the difference between the NK activity of the mononuclear cell and LGL populations, 44% of the RA patients demonstrated less of an increase than all of the control individuals although the percentages of LGL, Leu 11+, and Leu 7+ cells were equivalent.

A multicenter trial of sulindac in osteoarthritis of the hip.

Sulindac (cis-5-fluoro-2-methyl-l-[(p-methyl sulfinyl)-benzylidene]-indene-3-acetic acid) is a new nonsteroidal antirheumatic drug recently evaluated in a double-blind trial of 91 patients with hip osteoarthritis. Consecutive patients with documented flare following previous drug withdrawal were randomly assigned to one of 3 treatment groups: (1) sulindac given twice daily, (2) sulindac given 4 times daily, and (3) placebo. The dosage of sulindac, 100 to 300 mg daily, was adjusted according to patient global response and tolerance at 3- to 7-day intervals over 3 wk. Of 15 efficacy measurements evalulated, there was no difference between sulindac given 2 or 4 times daily, but differences were disclosed between one or both sulindac treatment groups and placebo in 11 of the 15 efficacy measurements (p less than 0.05, less than 0.01). The frequency of adverse reactions was of the same order for each treatment group. These included gastrointestinal upset, rash, and dizziness, usually transient and mild to moderate in severity. Serial laboratory studies revealed no evidence of renal, hepatic, or hematopoietic toxicity.