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1.
ESMO Open ; 7(4): 100540, 2022 08.
Article in English | MEDLINE | ID: mdl-35849877

ABSTRACT

BACKGROUND: Next-generation sequencing is used in cancer research to identify somatic and germline mutations, which can predict sensitivity or resistance to therapies, and may be a useful tool to reveal drug repurposing opportunities between tumour types. Multigene panels are used in clinical practice for detecting targetable mutations. However, the value of clinical whole-exome sequencing (WES) and whole-genome sequencing (WGS) for cancer care is less defined, specifically as the majority of variants found using these technologies are of uncertain significance. PATIENTS AND METHODS: We used the Cancer Genome Interpreter and WGS in 726 tumours spanning 10 cancer types to identify drug repurposing opportunities. We compare the ability of WGS to detect actionable variants, tumour mutation burden (TMB) and microsatellite instability (MSI) by using in silico down-sampled data to mimic WES, a comprehensive sequencing panel and a hotspot mutation panel. RESULTS: We reveal drug repurposing opportunities as numerous biomarkers are shared across many solid tumour types. Comprehensive panels identify the majority of approved actionable mutations, with WGS detecting more candidate actionable mutations for biomarkers currently in clinical trials. Moreover, estimated values for TMB and MSI vary when calculated from WGS, WES and panel data, and are dependent on whether all mutations or only non-synonymous mutations were used. Our results suggest that TMB and MSI thresholds should not only be tumour-dependent, but also be sequencing platform-dependent. CONCLUSIONS: There is a large opportunity to repurpose cancer drugs, and these data suggest that comprehensive sequencing is an invaluable source of information to guide clinical decisions by facilitating precision medicine and may provide a wealth of information for future studies. Furthermore, the sequencing and analysis approach used to estimate TMB may have clinical implications if a hard threshold is used to indicate which patients may respond to immunotherapy.


Subject(s)
Exome , Neoplasms , Biomarkers, Tumor , High-Throughput Nucleotide Sequencing , Humans , Microsatellite Instability , Mutation , Exome Sequencing
2.
Cancer Biomark ; 33(1): 159-169, 2022.
Article in English | MEDLINE | ID: mdl-34487023

ABSTRACT

BACKGROUND: Pleural mesothelioma is a deadly asbestos induced cancer. Less than 10% of mesothelioma patients survive 5 years post diagnosis. However survival can range from a few months to a number of years. Accurate prediction of survival is important for patients to plan for their remaining life, and for clinicians to determine appropriate therapy. One unusual feature of mesothelioma is that patients frequently present with tumor-associated pleural effusions early in the course of the disease. OBJECTIVE: To study whether cells and molecules present in pleural effusions provide prognostic information for mesothelioma. METHODS: We profiled the cellular constituents and concentrations of 40 cytokines, chemokines and cellular factors (collectively "soluble factors") involved in inflammatory and immune signalling pathways in pleural effusion samples from 50 mesothelioma patients.Associations with survival were evaluated by Cox proportional hazards regression methods. Results for the two soluble factors most significantly and independently associated with survival were validated in an independent set of samples (n= 51) using a separate assay system. RESULTS: Survival analysis revealed that IL8, IL2Ra (CD25) and PF4 were independent determinants of a more negative prognosis in mesothelioma patients, independent of other known prognostic factors. Lipocalin2 and IL4 were associated with better prognosis. CONCLUSIONS: This study demonstrates that pleural effusions rich in a range of soluble factors are associated with poor prognosis. These findings will enhance our ability to prognosticate outcomes in mesothelioma patients.


Subject(s)
Lung Neoplasms , Mesothelioma , Pleural Effusion, Malignant , Pleural Effusion , Pleural Neoplasms , Biomarkers, Tumor/metabolism , Humans , Lung Neoplasms/metabolism , Mesothelioma/diagnosis , Mesothelioma/metabolism , Pleural Effusion/diagnosis , Pleural Effusion, Malignant/diagnosis , Pleural Neoplasms/diagnosis , Prognosis
3.
Sci Rep ; 9(1): 14640, 2019 10 10.
Article in English | MEDLINE | ID: mdl-31601975

ABSTRACT

Non-small cell lung cancer (NSCLC) causes 19% of all Australian cancer deaths, with a 5-year survival post-resection of around 60%. Post-operative recurrence is due to metastases that were undetectable pre-operatively, or growth of microscopic locoregional residual disease. However, post-operative imaging modalities typically only detect more advanced tumours; where PET-CT has a detection limit of 6-7 mm. Detection of small deposits of lung metastatic disease is of importance in order to facilitate early and potentially more effective treatment. In this study, in a murine model of lung metastatic disease, we explore whether neo-antigen specific T cells are a sensitive marker for the detection of lung cancer after primary tumour resection. We determine lung metastatic disease by histology, and then compare detection by PET-CT and neo-antigen specific T cell frequency. Detection of lung metastatic disease within the histology positive group by PET-CT and neo-antigen specific T cell frequency were 22.9% and 92.2%, respectively. Notably, neo-antigen specific T cells in the lung draining lymph node were indicative of metastatic disease (82.8 ± 12.9 spots/105 cells; mean ± SE), compared to healthy lung control (28.5 ± 8.6 spots/105 cells; mean ± SE). Potentially, monitoring tumour neo-antigen specific T cell profiles is a highly sensitive method for determining disease recurrence.


Subject(s)
Carcinoma, Non-Small-Cell Lung/diagnosis , Lung Neoplasms/pathology , Lymphatic Metastasis/diagnosis , Lymphocytes, Tumor-Infiltrating/immunology , T-Lymphocytes/immunology , Aged , Animals , Antigens, Neoplasm/immunology , Carcinoma, Non-Small-Cell Lung/secondary , Carcinoma, Non-Small-Cell Lung/surgery , Cell Line, Tumor/transplantation , Disease Models, Animal , Enzyme-Linked Immunospot Assay , Female , Humans , Lung/diagnostic imaging , Lung/pathology , Lung/surgery , Lung Neoplasms/diagnosis , Lung Neoplasms/surgery , Lymph Nodes/cytology , Lymph Nodes/diagnostic imaging , Lymph Nodes/pathology , Lymphatic Metastasis/pathology , Mice , Middle Aged , Pneumonectomy , Positron Emission Tomography Computed Tomography , Treatment Outcome
4.
Ann Oncol ; 26(12): 2483-90, 2015 Dec.
Article in English | MEDLINE | ID: mdl-26386124

ABSTRACT

BACKGROUND: Data from murine models suggest that CD40 activation may synergize with cytotoxic chemotherapy. We aimed to determine the maximum tolerated dose (MTD) and toxicity profile and to explore immunological biomarkers of the CD40-activating antibody CP-870,893 with cisplatin and pemetrexed in patients with malignant pleural mesothelioma (MPM). PATIENTS AND METHODS: Eligible patients had confirmed MPM, ECOG performance status 0-1, and measurable disease. Patients received cisplatin 75 mg/m(2) and pemetrexed 500 mg/m(2) on day 1 and CP-870,893 on day 8 of a 21-day cycle for maximum 6 cycles with up to 6 subsequent cycles single-agent CP-870,893. Immune cell subset changes were examined weekly by flow cytometry. RESULTS: Fifteen patients were treated at three dose levels. The MTD of CP-870,893 was 0.15 mg/kg, and was exceeded at 0.2 mg/kg with one grade 4 splenic infarction and one grade 3 confusion and hyponatraemia. Cytokine release syndrome (CRS) occurred in most patients (80%) following CP-870,893. Haematological toxicities were consistent with cisplatin and pemetrexed chemotherapy. Six partial responses (40%) and 9 stable disease (53%) as best response were observed. The median overall survival was 16.5 months; the median progression-free survival was 6.3 months. Three patients survived beyond 30 months. CD19+ B cells decreased over 6 cycles of chemoimmunotherapy (P < 0.001) with a concomitant increase in the proportion of CD27+ memory B cells (P < 0.001) and activated CD86+CD27+ memory B cells (P < 0.001), as an immunopharmacodynamic marker of CD40 activation. CONCLUSIONS: CP-870,893 with cisplatin and pemetrexed is safe and tolerable at 0.15 mg/kg, although most patients experience CRS. While objective response rates are similar to chemotherapy alone, three patients achieved long-term survival. AUSTRALIA NEW ZEALAND CLINICAL TRIALS REGISTRY NUMBER: ACTRN12609000294257.


Subject(s)
Antibodies, Monoclonal/administration & dosage , Antineoplastic Combined Chemotherapy Protocols/administration & dosage , CD40 Antigens/metabolism , Cisplatin/administration & dosage , Lung Neoplasms/drug therapy , Mesothelioma/drug therapy , Pemetrexed/administration & dosage , Pleural Neoplasms/drug therapy , Adult , Aged , Antibodies, Monoclonal, Humanized , CD40 Antigens/agonists , Cohort Studies , Female , Follow-Up Studies , Humans , Lung Neoplasms/diagnosis , Lung Neoplasms/metabolism , Male , Mesothelioma/diagnosis , Mesothelioma/metabolism , Mesothelioma, Malignant , Middle Aged , Pleural Neoplasms/diagnosis , Pleural Neoplasms/metabolism , Prospective Studies
5.
Cytopathology ; 26(3): 142-56, 2015 Jun.
Article in English | MEDLINE | ID: mdl-26052757

ABSTRACT

OBJECTIVE: To provide practical guidelines for the cytopathologic diagnosis of malignant mesothelioma. DATA SOURCES: Cytopathologists with an interest in the field involved in the International Mesothelioma Interest Group (IMIG) and the International Academy of Cytology (IAC) contributed to this update. Reference material includes peer-reviewed publications and textbooks. RATIONALE: This article is the result of discussions during and after the IMIG 2012 conference in Boston, followed by thorough discussions during the 2013 IAC meeting in Paris. Additional contributions have been obtained from cytopathologists and scientists who could not attend these meetings, with final discussions and input during the IMIG 2014 conference in Cape Town.


Subject(s)
Mesothelioma/diagnosis , Cytodiagnosis , Humans
7.
Br J Cancer ; 109(7): 1813-20, 2013 Oct 01.
Article in English | MEDLINE | ID: mdl-23982605

ABSTRACT

BACKGROUND: Recent studies proposed neutrophil-to-lymphocyte ratio (NLR) as a prognostic biomarker in malignant pleural mesothelioma (MPM). We examined baseline prognostic variables including NLR and the EORTC and CALGB models as predictors of overall survival (OS) in MPM. METHODS: In this retrospective study, 274 consecutive eligible, newly presenting patients with MPM were included. Of these, 159 received chemotherapy, 10 had tri-modality therapy, 2 underwent surgery only and 103 received supportive care alone. Univariate analyses and multivariate Cox models were calculated for OS. RESULTS: In univariate analysis, poor prognostic factors were: age ≥65 years, nonepithelioid histology, stage III-IV, poor performance status (PS), weight loss, chest pain, low haemoglobin and high platelet count. A baseline NLR≥ 5 did not predict worse OS (hazard ratio (HR) 1.25; P=0.122). On multivariate analysis, age, histology, PS, weight loss, chest pain and platelet count remained significant. The EORTC and CALGB prognostic groups were validated as predictive for OS (HR 1.62; P<0.001 and HR 1.65; P<0.001, respectively). CONCLUSION: Our findings validate standard prognostic variables and the existing EORTC and CALGB models, but not NLR, at initial diagnosis of MPM. In guiding patient management at diagnosis, it is important to consider multiple baseline variables that jointly predict survival.


Subject(s)
Biomarkers, Tumor , Lung Neoplasms/mortality , Lymphocytes/cytology , Mesothelioma/mortality , Neutrophils/cytology , Adult , Aged , Aged, 80 and over , Female , Humans , Leukocyte Count , Lung Neoplasms/drug therapy , Lymphocyte Count , Male , Mesothelioma/drug therapy , Mesothelioma, Malignant , Middle Aged , Prognosis , Retrospective Studies , Survival Rate
8.
Br J Cancer ; 98(9): 1562-9, 2008 May 06.
Article in English | MEDLINE | ID: mdl-18454162

ABSTRACT

Current interest in the MUC1/EMA mucin relates to its role in malignancy, and its potential as a therapeutic target. MUC1/EMA expression has been observed in the majority of epithelioid mesotheliomas. However, little is known of the characteristics of MUC1/EMA in mesothelioma. Herein, we studied the cell surface and soluble expression of the MUC1/EMA glycoprotein, and determined the mRNA and genomic expression profiles in mesothelioma. We found that the anti-MUC1 antibody, E29, was the most diagnostically useful of seven antibody clones examined with a sensitivity of 84% (16 out of 19 cases) and no false positive results. MUC1 mRNA expression was significantly higher in mesothelioma samples than in benign mesothelial cells. No amplification of the MUC1 gene was observed by FISH. Seven of 9 mesothelioma samples expressed MUC1-secreted mRNA isoform in addition to the archetypal MUC1/transmembrane form. CA15.3 (soluble MUC1) levels were significantly higher in the serum of mesothelioma patients than in healthy controls but were not significantly different to levels in patients with benign asbestos-related disease. CA15-3 in effusions could differentiate malignant from benign effusions but were not specific for mesothelioma. Thus, as in other cancers, alterations in MUC1 biology occur in mesothelioma and these results suggest that specific MUC1 characteristics may be useful for mesothelioma diagnosis and should also be investigated as a potential therapeutic target.


Subject(s)
Biomarkers, Tumor/metabolism , Mesothelioma/diagnosis , Mesothelioma/metabolism , Mucin-1/metabolism , Pleural Effusion, Malignant/diagnosis , Pleural Effusion, Malignant/metabolism , Aged , Aged, 80 and over , Alternative Splicing , Biomarkers, Tumor/analysis , Biomarkers, Tumor/blood , Female , Gene Expression Regulation, Neoplastic , Glycosylation , Humans , Immunohistochemistry , In Situ Hybridization, Fluorescence , Male , Mesothelioma/blood , Mesothelioma/chemistry , Middle Aged , Mucin-1/analysis , Mucin-1/blood , Mucin-1/genetics , Pleural Effusion, Malignant/blood , Pleural Effusion, Malignant/chemistry , Polymerase Chain Reaction , RNA, Messenger/metabolism , Sensitivity and Specificity , Up-Regulation
9.
J Biol Chem ; 276(32): 30031-5, 2001 Aug 10.
Article in English | MEDLINE | ID: mdl-11325959

ABSTRACT

Bloom's syndrome (BS) is a rare autosomal recessive disorder characterized by pre- and postnatal growth deficiency, immunodeficiency, and a tremendous predisposition to a wide variety of cancers. Cells from BS individuals are characterized by a high incidence of chromosomal gaps and breaks, elevated sister chromatid exchange, quadriradial formations, and locus-specific mutations. BS is the consequence of mutations that lead to loss of function of BLM, a gene encoding a helicase with homology to the RecQ helicase family. To delineate the role of BLM in DNA replication, recombination, and repair we used a yeast two-hybrid screen to identify potential protein partners of the BLM helicase. The C terminus of BLM interacts directly with MLH1 in the yeast-two hybrid assay; far Western analysis and co-immunoprecipitations confirmed the interaction. Cell extracts deficient in BLM were competent for DNA mismatch repair. These data suggest that the BLM helicase and MLH1 function together in replication, recombination, or DNA repair events independent of single base mismatch repair.


Subject(s)
Adenosine Triphosphatases/metabolism , Base Pair Mismatch , DNA Helicases/metabolism , DNA Repair , Neoplasm Proteins/metabolism , Adaptor Proteins, Signal Transducing , Adenosine Triphosphatases/chemistry , Blotting, Western , Carrier Proteins , Cell Line , Cell Nucleus/metabolism , Cells, Cultured , Child , DNA Helicases/chemistry , DNA Replication , HeLa Cells , Humans , K562 Cells , Male , MutL Protein Homolog 1 , Mutation , Nuclear Proteins , Precipitin Tests , Protein Binding , RecQ Helicases , Recombination, Genetic , Tumor Cells, Cultured , Two-Hybrid System Techniques
10.
Br J Cancer ; 84(1): 52-6, 2001 Jan 05.
Article in English | MEDLINE | ID: mdl-11139313

ABSTRACT

Malignant mesothelioma (MM) generally occurs as a pleural tumour, related to the inhalation of asbestos fibres. It is highly aggressive and largely unresponsive to treatment. The incidence of MM is particularly high in Western Australia because of the extensive blue asbestos mining operations that occurred in the north of the state until 1966. MM is unusual in that mutations in the tumour suppressor gene p53 are rarely observed, whilst over-expression of p53 protein is common. As the level of antibodies directed against p53 is thought to be of prognostic value in some cancers and as MM is known to be immunogenic, we studied a cohort of Western Australian patients to determine the prevalence of anti-p53 antibodies and their value as diagnostic markers or prognostic indicators. 6/88 (7%) of patients had high titres (>2 SD above the mean of controls) of anti-p53 antibodies. There was no correlation between antibody titre and survival. Although 3/38 (8%) of sera obtained from patients exposed to asbestos but prior to a diagnosis of MM contained antibodies, the same proportion of sera obtained from patients exposed to asbestos but who remained disease free also contained antibodies (2/40; 8%). Sera collected sequentially demonstrated a profound temporal stability in the titre of anti-p53 antibodies in patients with MM throughout the course of their illness. These results show that anti-p53 antibodies are observed only at a low frequency in the sera of MM patients and where they do occur, their elicitation is an early event that may be unrelated to antigen load. The occurrence of anti-p53 antibodies does not serve as either a useful prognostic or diagnostic indicator in MM.


Subject(s)
Antibodies, Neoplasm/blood , Autoantibodies/blood , Mesothelioma/immunology , Pleural Neoplasms/immunology , Tumor Suppressor Protein p53/immunology , Adult , Aged , Aged, 80 and over , Blotting, Western , Case-Control Studies , Enzyme-Linked Immunosorbent Assay , Female , HT29 Cells/immunology , Humans , Male , Mesothelioma/blood , Middle Aged , Pleural Neoplasms/blood
11.
Infect Immun ; 67(2): 989-93, 1999 Feb.
Article in English | MEDLINE | ID: mdl-9916122

ABSTRACT

In this study, interleukin-5 (IL-5) transgenic mice with lifelong eosinophilia were assessed for resistance to primary infections with two tissue-invading nematodes, Nippostrongylus brasiliensis and Toxocara canis. Relative to nontransgenic littermates, three lines of IL-5 transgenic mice with varying degrees of eosinophilia all displayed enhanced resistance to N. brasiliensis. Although the timing of final worm expulsion was similar in transgenic and nontransgenic hosts, intestinal worms in transgenic mice were fewer in number throughout infection, failed to increase in size over the course of the infection, and were much less fecund. In contrast, T. canis larvae were recovered in similar numbers from tissues of transgenic mice with "low" or "high" eosinophilia and from nontransgenic mice. These results and other data suggest that eosinophils can contribute to host resistance to some parasite species. Parasite transit time through the host may correlate with relative sensitivity to eosinophils.


Subject(s)
Interleukin-5/immunology , Nippostrongylus/immunology , Strongylida Infections/immunology , Toxocara canis/immunology , Toxocariasis/immunology , Animals , Eosinophilia/physiopathology , Female , Immunity, Innate/immunology , Interleukin-5/genetics , Intestine, Small/parasitology , Intestine, Small/pathology , Male , Mice , Mice, Inbred CBA , Mice, Transgenic , Nippostrongylus/physiology , Ovum , Strongylida Infections/parasitology , Strongylida Infections/pathology , Toxocara canis/physiology , Toxocariasis/parasitology , Toxocariasis/pathology
12.
Exp Parasitol ; 83(2): 202-15, 1996 Jul.
Article in English | MEDLINE | ID: mdl-8682189

ABSTRACT

Irradiation has been successful in the attenuation of infective stage parasites for use as vaccines against a number of parasites including Fasciola spp. The mechanisms of action of irradiation-attenuated vaccines, however, are not clearly understood. In this study, we examined the effect of 3, 10, and 40 krad of gamma-irradiation on the expression of carbohydrates and cathepsin-B by newly excysted juvenile Fasciola hepatica (NEJ). Following irradiation of metacercariae, the expression of concanavalin A (ConA)-specific sugars was decreased on the surface of NEJ and the expression of wheat germ agglutinin (WGA)-specific sugars was increased in the gut and reduced on the surface of NEJ. Cathepsin proteases are a major component of liver fluke excretory/secretory material (ES) and can cleave host immunoglobulin (Ig). Cathepsin-B protease was localized in nonirradiated NEJ to the gut lumen and to secretory granules within the gut epithelia. Irradiation of fluke with 3, 10, and 40 krad of gamma-rays significantly reduced the tissue expression of cathepsin-B at 8 hr postirradiation in an apparently dose-dependent manner. After a further 24 hr culture tissue expression of cathepsin-B was significantly reduced in 10- and 40-krad-irradiated NEJ. Protease activity of ES samples collected over a 24-hr period from irradiated and nonirradiated NEJ cultured in vitro were tested using a rabbit Ig cleavage assay. The proteolytic activity of ES from 10- and 40-krad-irradiated NEJ was reduced during the initial 6 hr in culture and between 12 and 24 hr when compared to ES from nonirradiated controls. Biosynthetic labeling experiments using [35S]methionine and [35S]cysteine indicated that ES material was actively synthesised during 48 hr in vitro culture. Therefore, from this study, we conclude that gamma-irradiation of NEJ alters expression of cathepsin-B protease and WGA- and ConA-specific sugars which may be detrimental to parasite invasion and contribute to the protective immune responses generated in the host by irradiation-attenuated metacercariae of Fasciola spp.


Subject(s)
Carbohydrates/biosynthesis , Cathepsin B/biosynthesis , Fasciola hepatica/radiation effects , Animals , Concanavalin A/metabolism , Fasciola hepatica/enzymology , Fasciola hepatica/metabolism , Helminth Proteins/biosynthesis , Immunoglobulins/metabolism , Immunohistochemistry , Rabbits , Wheat Germ Agglutinins/metabolism
13.
Mol Biochem Parasitol ; 74(1): 19-29, 1995 Oct.
Article in English | MEDLINE | ID: mdl-8719242

ABSTRACT

A low molecular mass monomeric protein termed Fh-KTM (Fasciola hepatica Kunitz-type molecule) was isolated from the trematode Fasciola hepatica. Fh-KTM is a single polypeptide of 58 amino acids and a Mr of 6751. The complete amino acid sequence of Fh-KTM was determined and revealed significant similarity to the Kunitz-type (BPTI) family of proteinase inhibitors. Several polymorphisms were observed suggesting that more than one Fh-KTM molecule may be expressed by this parasite. Modified proline residues were shown to occur at all four positions in this protein as 3-hydroxy derivatives. This is the first report of 3-hydroxyproline residues in a Kunitz-type molecule. Indirect immunofluorescence and immunogold labelling revealed that Fh-KTM is an abundant molecule within the parasite localised to the gut, the parenchymal tissue and the tegument of adult F. hepatica. Serine protease inhibition assays revealed that Fh-KTM exhibited little or no inhibition against chymotrypsin, kallikrein, urokinase or key serine proteases of the blood coagulation pathways. However, Fh-KTM was able to inhibit trypsin even though the P1 reactive amino acid of Fh-KTM was a leucine residue.


Subject(s)
Fasciola hepatica/genetics , Helminth Proteins/genetics , Amino Acid Sequence , Animals , Aprotinin/chemistry , Aprotinin/genetics , Aprotinin/isolation & purification , Fasciola hepatica/chemistry , Helminth Proteins/chemistry , Helminth Proteins/isolation & purification , Immunohistochemistry , Molecular Sequence Data , Molecular Weight , Sequence Homology, Amino Acid , Serine Proteinase Inhibitors/chemistry , Serine Proteinase Inhibitors/genetics , Serine Proteinase Inhibitors/isolation & purification , Trypsin Inhibitors/chemistry , Trypsin Inhibitors/genetics
14.
Exp Parasitol ; 81(1): 106-16, 1995 Aug.
Article in English | MEDLINE | ID: mdl-7628558

ABSTRACT

Four cDNA clones (GST-1, -7, -47, and -51) encoding isoenzymes of the detoxification enzyme glutathione S-transferase (GST) have previously been identified and characterised from Fasciola hepatica. In the present study, antisera were generated to synthetic peptides of regions unique to each of the four GST proteins predicted by the cDNAs. The antisera were characterised, and two were found to distinguish GST-1 from GST-7, GST-47, and GST-51 as a group. These two antisera were used to localise different GSTs in adult and newly excysted juvenile F. hepatica. The antiserum to GST-1 was specific and localised GST-1 to the parenchyma of adult fluke but not to the lamellae of the intestinal caeca. The antiserum to a GST-51 peptide, which cross-reacted with GST-7 and GST-47 but not GST-1, localised the other GSTs not only to the parenchyma but also to the intestinal lamellae of adult fluke. This appears to be the first evidence of tissue-specific expression of GST isoenzymes in trematodes. In contrast to adult fluke, immunolocalisation of the GSTs in juvenile F. hepatica revealed the binding of both the GST-1 and GST-51 antisera to the parenchymal cytoplasm, to cytoplasmic extensions of the parenchyma cells in the subtegumental area, as well as the excretory ducts. No labeling was observed in the intestinal epithelium of the juvenile fluke. These results demonstrate that adult F. hepatica, in contrast to juvenile flukes, contain a GST, which is not GST-1, associated with the lamellae of the gut and suggest that GSTs in adult fluke may play a role in the absorptive function of the adult gut.


Subject(s)
Aging/metabolism , Fasciola hepatica/enzymology , Glutathione Transferase/metabolism , Isoenzymes/metabolism , Animals , Antibody Specificity , Blotting, Western , Fasciola hepatica/growth & development , Fasciola hepatica/ultrastructure , Fluorescent Antibody Technique , Glutathione Transferase/analysis , Immune Sera , Isoenzymes/analysis , Microscopy, Immunoelectron , Peptide Fragments/chemical synthesis , Peptide Fragments/immunology , Peptides/chemical synthesis , Peptides/immunology , Recombinant Proteins/analysis , Sensitivity and Specificity
15.
Int J Parasitol ; 25(7): 853-6, 1995 Jul.
Article in English | MEDLINE | ID: mdl-7558572

ABSTRACT

The potential of gamma-irradiated Fasciola hepatica metacercariae to vaccinate sheep against fascioliasis was examined. The effect of the size of the inocula of irradiated metacercariae and the level of gamma-irradiation on the recovery of non-irradiated fluke was assessed following homologous challenge. Groups of Merino wethers were vaccinated with a single infection of either 500 or 2000 metacercariae, previously exposed to either 30, 100 or 400 Gy of gamma-irradiation. No significant reduction of fluke burdens were observed in any group, although a nonsignificant 20% reduction was observed in sheep vaccinated with 2000 metacercariae irradiated with 100 Gy. A second trial was conducted in which groups of sheep were vaccinated with 2 doses, given 4 weeks apart, of 2000 metacercariae, previously irradiated at either 70, 100 or 150 Gy. In both trials parasite viability was severely affected by doses of gamma-irradiation of 30 Gy or greater and no mature flukes were recovered from control sheep given metacercariae attenuated with 70 Gy or greater. A strong humoral immune response to somatic F. hepatica antigens was observed in all sheep. Only sera from sheep receiving 70 Gy irradiated metacercariae recognised the 2 candidate liver fluke vaccine molecules, F. hepatica glutathione S-transferase and cathepsin-L proteases. No reduction was observed in either the number of flukes or the production of fluke eggs in any vaccinated group. Vaccination appeared to affect the development of the challenge fluke population, resulting in reduced hepatic damage during migration, as measured by levels of serum glutamate dehydrogenase, and an increase in mean fluke weight.


Subject(s)
Fasciola hepatica/immunology , Fascioliasis/veterinary , Immunization/veterinary , Sheep Diseases/prevention & control , Animals , Antibodies, Helminth/biosynthesis , Antigens, Helminth/administration & dosage , Fasciola hepatica/growth & development , Fasciola hepatica/radiation effects , Fascioliasis/immunology , Fascioliasis/prevention & control , Gamma Rays , Male , Sheep , Sheep Diseases/immunology , Sheep Diseases/parasitology
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