The potential of molecular imprinting as a biosensing devices for monitoring the CEA cancer biomarker

6th Graduate Student Symposium on Molecular Imprinting

A novel antibody-like material for breast cancer antigen CA15-3, used to track breast cancer by potentiometric transduction

This work presents the development of a low cost sensor device for the diagnosis of breast cancer in point-of-care, made with new synthetic biomimetic materials inside plasticized poly(vinyl chloride), PVC, membranes, for subsequent potentiometric detection. This concept was applied to target a conventional biomarker in breast cancer: Breast Cancer Antigen (CA15-3). The new biomimetic material was obtained by molecularly-imprinted technology. In this, a plastic antibody was obtained by polymerizing around the biomarker that acted as an obstacle to the growth of the polymeric matrix. The imprinted polymer was specifically synthetized by electropolymerization on an FTO conductive glass, by using cyclic voltammetry, including 40 cycles within -0.2 and 1.0 V. The reaction used for the polymerization included monomer (pyrrol, 5.0×10-3 mol/L) and protein (CA15-3, 100U/mL), all prepared in phosphate buffer saline (PBS), with a pH of 7.2 and 1% of ethylene glycol. The biomarker was removed from the imprinted sites by proteolytic action of proteinase K. The biomimetic material was employed in the construction of potentiometric sensors and tested with regard to its affinity and selectivity for binding CA15-3, by checking the analytical performance of the obtained electrodes. For this purpose, the biomimetic material was dispersed in plasticized PVC membranes, including or not a lipophilic ionic additive, and applied on a solid conductive support of graphite. The analytical behaviour was evaluated in buffer and in synthetic serum, with regard to linear range, limit of detection, repeatability, and reproducibility. This antibody-like material was tested in synthetic serum, and good results were obtained. The best devices were able to detect 5 times less CA15-3 than that required in clinical use. Selectivity assays were also performed, showing that the various serum components did not interfere with this biomarker. Overall, the potentiometric-based methods showed several advantages compared to other methods reported in the literature. The analytical process was simple, providing fast responses for a reduced amount of analyte, with low cost and feasible miniaturization. It also allowed the detection of a wide range of concentrations, diminishing the required efforts in previous sample pre-treating stages.

Sol-gel biomimetic material designed to target CEA cancer biomarker

1st ASPIC International Congress

A disposable glass-based immunosensor for monitoring the cancer biomarker CEA in urine

NanoPT 2014 International Conference, in Portugal, on February 12-14. Poster presentation based on topic Nanobio/Nanomedicine

Cytotoxicity Induced by Extracts of Pisolithus tinctorius Spores on Human Cancer and Normal Cell Lines—Evaluation of the Anticancer Potential

Fungi have been considered a potential source of natural anticancer drugs. However, studies on these organisms have mainly focused on compounds present in the sporocarp and mycelium. The aim of this study was to assess the anticancer potential of fungal spores using a bioassay-guided fractionation with cancer and normal cell lines. Crude extracts from spores of the basidiomycetous fungus Pisolithus tinctorius were prepared using five solvents/solvent mixtures in order to select the most effective crude extraction procedure. A dichloromethane/methanol (DCM/MeOH) mixture was found to produce the highest extraction yield, and this extract was fractionated into 11 fractions. Crude extracts and fractions were assayed for cytotoxicity in the human osteocarcinoma cell line MG63, the human breast carcinoma cell line T47D, the human colon adenocarcinoma cell line RKO, and the normal human brain capillary endothelial cell line hCMEC/D3. Cytotoxicity was assessed by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) reduction assay. The results showed a reduction in cancer cell viability of approximately 95% with 4 of 11 fractions without a significant reduction in viability of hCMEC/D3 cells. Data demonstrated that spores of P. tinctorius might serve as an interesting source of compounds with potential anticancer properties.

Psychosocial Morbidity and its Correlates in Cancer Patients of the Mediterranean Area: Findings from the Southern European Psycho-Oncology Study

BACKGROUND: A few and partial data are available on psychosocial morbidity among cancer patients in Mediterranean countries. As a part of a more general investigation (Southern European Psycho-Oncology Study-SEPOS), the rate of psychosocial morbidity and its correlation with clinical and cultural variables were examined in cancer patients in Italy, Portugal and Spain. METHODS: A convenience sample of cancer outpatients with good performance status and no cognitive impairment were approached. The Hospital Anxiety-Depression scale (HAD-S), the Mini-Mental Adjustment to Cancer scale (Mini-MAC), and the Cancer Worries Inventory (CWI) were used to measure psychological morbidity, coping strategies and concerns about illness. RESULTS: Of 277 patients, 34% had pathological scores ("borderline cases" plus "true cases") on HAD-S Anxiety and 24.9% on HAD-S Depression. Total psychiatric "caseness" was 28.5% and 16.6%, according to different HAD cut-offs (14 and 19, respectively). Significant relationships of HAD-S Anxiety, HAD-S Depression, HAD-S Total score, with Mini-MAC Hopeless and Anxious Preoccupation, and CWI score were found. No differences emerged between countries on psychosocial morbidity, while some differences emerged between the countries on coping mechanisms. Furthermore, Fatalism, Avoidance and marginally Hopeless were higher compared to studies carried out in English-speaking countries. LIMITATIONS: The relatively small sample size and the good performance status prevent us to generalize data on patients with different cancer sites and advanced phase of illness. CONCLUSIONS: One-third of the patients presented anxiety and depressive morbidity, with significant differences in characteristics of coping in Mediterranean countries in comparison with English-speaking countries.

Use of Distress and Depression Thermometers to Measure Psychosocial Morbidity Among Southern European Cancer Patients

GOALS OF WORK: Recent literature has indicated the need for rapid evaluation of psychosocial issues secondary to cancer. Because of the problems of routine use of psychometric instruments, short instruments such as visual analogue scales or one-item 0-10 scales have been developed as valid assessment alternatives. PATIENTS AND METHODS: A study was conducted to examine the role of two 0-10 scales in measuring emotional stress (distress thermometer, DT) and depressed mood (mood thermometer, MT), respectively, in a multicenter study carried out in southern European countries (Italy, Portugal, Spain, and Switzerland). A convenience sample of 312 cancer outpatients completed the DT and MT and the Hospital Anxiety Depression Scale (HADS). MAIN RESULTS: DT was more significantly associated HADS anxiety than HADS depression while MT was related both to HADS anxiety and depression. The correlation of MT with HADS was higher than DT. A cutoff point >4 on the DT maximized sensitivity (65%) and specificity (79%) for general psychosocial morbidity while a cutoff >5 identified more severe "caseness" (sensitivity=70%; specificity=73%). On the MT, sensitivity and specificity for general psychosocial morbidity were 85% and 72% by using the cutoff score >3. A score >4 on the MT was associated with a sensitivity of 78% and a specificity of 77% in detecting more severe caseness. CONCLUSIONS: Two simple instruments, the DT and the MT, were found to have acceptable levels of sensitivity and specificity in detecting psychosocial morbidity. Compared to the HADS, however, the mood MT performed better than the DT.

Pulmonary thromboembolism in AIDS patient with chronic venous insufficiency, pulmonary tuberculosis and breast cancer: a case report and pathophysiology review

Recent literature reports thrombotic episodes occurring in patients with HIV infection associated with other abnormalities including neoplasms and infections predisposing to a hypercoagulable state. We report a 47-year-old woman who developed pulmonary thromboembolism in association with HIV infection, pulmonary tuberculosis and breast cancer. She was treated with rifampin, isoniazid, pyrazinamide; heparin, phenprocoumon, zidovudine, lamivudine and efavirenz. Acid fast bacilli were visualized in a sputum smear and three months after, Mycobacterium tuberculosis was isolated from lymph node biopsy during a episode of immune reconstitution. The isolated mycobacteria showed sensitivity to all first-line drugs. HIV infection, breast cancer and pulmonary tuberculosis have several mechanisms that induce hypercoagulable state and can lead to thromboembolic complications. Pulmonary thromboembolism in this patient was a diagnostic challenge because of all the other severe diseases that she experienced at the same time.

Mesotelioma Peritoneal Maligno. Dificuldades Diagnósticas e Terapêuticas

O Mesotelioma Peritoneal Maligno (MPM) é um tumor raro, de apresentação clínica inespecífica, colocando dificuldades diagnósticas particularmente na diferenciação com carcinomatose peritoneal. O diagnóstico tardio e a ineficácia da terapêutica convencional – cirurgia, radioterapia, quimioterapia – conferem-lhe um mau prognóstico com sobrevida média de 6-12 meses. Os autores descrevem um caso de MPM diagnosticado durante investigação de ascite inaugural, através de biópsia peritoneal laparoscópica. Submetido a quimioterapia sistémica, o doente encontra-se em remissão parcial aos 42 meses.

Presence of Leishmania amastigotes in peritoneal fluid of a dog with leishmaniasis from Alagoas, Northeast Brazil

The goal of this short communication is to report the uncommon presence of intracellular amastigotes of Leishmania in peritoneal fluid of a dog with leishmaniasis from Alagoas State, Brazil. Physical examination of an adult male rottweiler suspected to be suffering of leishmaniasis revealed severe loss of weight, ascitis, splenomegaly, moderately enlarged lymph nodes, onychogryphosis, generalized alopecia, skin ulcers on the posterior limbs, and conjunctivitis. Samples of bone marrow, popliteal lymph node, skin ulcer, and peritoneal fluid were collected and smears of each sample were prepared and stained with hematoxylin and eosin. Numerous amastigotes were detected in bone marrow, popliteal lymph node, and skin ulcer smears. Smears of peritoneal fluid revealed the unusual presence of several free and intracellular amastigotes of Leishmania. Future studies are needed to determine whether the cytology of ascitic fluid represents a useful tool for diagnosis Leishmania infection in ascitic dogs, particularly in those living in areas where canine leishmaniasis is enzootic.

Effects of Environmental Organochlorine Pesticides on Human Breast Cancer: Putative Involvement on Invasive Cell Ability

Human exposure to persistent organic pollutants (POPs) is a certainty, even to long banned pesticides like o,p′-dichlorodiphenyltrichloroethane (o,p′-DDT), and its metabolites p,p′-dichlorodiphenyldichloroethylene (p,p′-DDE), and p,p′-dichlorodiphenyldichloroethane (p,p′-DDD). POPs are known to be particularly toxic and have been associated with endocrine-disrupting effects in several mammals, including humans even at very low doses. As environmental estrogens, they could play a critical role in carcinogenesis, such as in breast cancer. With the purpose of evaluating their effect on breast cancer biology, o,p′-DDT, p,p′-DDE, and p,p′-DDD (50–1000 nM) were tested on two human breast adenocarcinoma cell lines: MCF-7 expressing estrogen receptor (ER) α and MDA-MB-231 negative for ERα, regarding cell proliferation and viability in addition to their invasive potential. Cell proliferation and viability were not equally affected by these compounds. In MCF-7 cells, the compounds were able to decrease cell proliferation and viability. On the other hand, no evident response was observed in treated MDA-MB-231 cells. Concerning the invasive potential, the less invasive cell line, MCF-7, had its invasion potential significantly induced, while the more invasive cell line MDA-MB-231, had its invasion potential dramatically reduced in the presence of the tested compounds. Altogether, the results showed that these compounds were able to modulate several cancer-related processes, namely in breast cancer cell lines, and underline the relevance of POP exposure to the risk of cancer development and progression, unraveling distinct pathways of action of these compounds on tumor cell biology.

Cytomegalovirus in colorectal cancer and idiopathic ulcerative colitis

Cytomegalovirus (CMV) is a genus in the family Herpesviridae that has been associated with gastrointestinal syndromes. In this work we looked for a possible association of CMV infection with colorectal cancer and ulcerative colitis (UC). Blood and enteric tissue samples of 14 patients with colorectal cancer and of 21 with UC were subjected to a nested-PCR that amplifies part of the gB gene of CMV and also to immunohistochemistry using a specific monoclonal antibody to IE 76kDa protein of CMV. CMV was detected by nested-PCR in the blood and/or the enteric tissue of nine (64.3%) colorectal cancer and 16 (76.2%) ulcerative colitis patients. In the immunohistochemistry it was observed that 12 (12/21, 57.1%) positive enteric tissue samples of patients with UC and none from patients with colorectal cancer (0/14) were positive to CMV. The positivity of CMV infections in the UC patient group (12/21, 57.1%) showed by both techniques, was significantly higher (p = 0.015) than that observed for colorectal cancer patients (2/14, 14.3%). These results suggest an association of ulcerative colitis with CMV infection of the enteric tissue.

Longitudinal analysis of tumor marker CEA of breast cancer patients from Braga's hospital

Allied to an epidemiological study of population of the Senology Unit of Braga’s Hospital that have been diagnosed with malignant breast cancer, we describe the progression in time of repeated measurements of tumor marker Carcinoembryonic antigen (CEA). Our main purpose is to describe the progression of this tumor marker as a function of possible risk factors and, hence, to understand how these risk factors influences that progression. The response variable, values of CEA, was analyzed making use of longitudinal models, testing for different correlation structures. The same covariates used in a previous survival analysis were considered in the longitudinal model. The reference time used was time from diagnose until death from breast cancer. For diagnostic of the models fitted we have used empirical and theoretical variograms. To evaluate the fixed term of the longitudinal model we have tested for a changing point on the effect of time on the tumor marker progression. A longitudinal model was also fitted only to the subset of patients that died from breast cancer, using the reference time as time from date of death until blood test.