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Objective: to develop a multiple-endpoint monitoring system in order to assess and minimize long term risks in hospital nurses exposed to antiblastic drugs. Design: molecular epidemiology study. Setting: S. Orsola-Malpighi Hospital in Bologna, Italy nurses exposed to antiblastic drugs. Participants: 50 exposed subjects (8 males and 42 females) and 50 unexposed individuals (8 males and 42 females) matched for age and smoking habits. Main outcome measures: urinary markers of exposure; Heat Shock Proteins (HSPs) 27, 70, 90, 110; immunological biomarkers in peripheral blood lymphocytes; apoptosis, cell-cycle analysis G1-S-G, typization of Natural Killer cells (NK) and receptors for IL-2 in peripheral blood lymphocytes; micronuclei: frequency in peripheral blood lymphocytes and in exfoliated buccal mucosa cells; activation of specific oncogenes (bax, bcl2, b-actin). Results: 19/50 subjects showed urinary antiblastic drugs levels (3 subjects MTX, 11 subjects CP, 5 subjects MTX and CP). No statistically significant differences were observed in all the considered biomarkers between the exposed and control groups. Conclusion: this biomonitoring study doesn’t evidence any early significant effect associated to the exposure to antiblastic drugs.

OBJECTIVE: To develop a multiple-endpoint monitoring system in order to assess and minimize long term risks in hospital nurses exposed to antiblastic drugs. DESIGN: Molecular epidemiology study. SETTING: S. Orsola-Malpighi Hospital in Bologna, Italy: nurses exposed to antiblastic drugs. PARTICIPANTS: 50 exposed subjects (8 males and 42 females) and 50 unexposed individuals (8 males and 42 females) matched for age and smoking habits. MAIN OUTCOME MEASURES: Urinary markers of exposure, Heat Shock Proteins (HSPs) 27, 70, 90, 110, immunologic biomarkers in peripheral blood lymphocytes: apoptosis, cell-cycle analysis G1-S-G, typization of Natural Killer cells (NK) and receptors micronuclei; frequency in peripheral blood lymphocytes and in exfoliated buccal mucosa cells; activation ofspecific oncogenes (bax, bcl2). RESULTS: 19/50 subjects showed urinary antiblastic drug levels (3 subjects MTX, 11 subjects CP, 5 subjects MTX and CP). No statistically significant differences were observed in all the considered biomarkers between the exposed and control groups. CONCLUSION: This biomonitoring study doesn't evidence any early significant effect associated to the exposure to antiblastic drugs.

Chemotherapy, as generally available, is of a limited value in curing malignant brain tumors (gliomas), which often develop resistance to drugs, becoming completely unresponsive to any standard therapeutic approach. Camptothecins, a family of topoisomerase I inhibitor drugs, represent a new promising treatment strategy and are currently under evaluation for testing the clinical efficacy. We selected a CPT-resistant sub-line (U87CPT-R) from U87-MG grade III-IV astrocytoma cells, and compared the expression profile of the two cell lines by cDNA-microarray, as a preliminary screening of the molecular mechanisms involved in the acquisition of CPT resistance in glioma cells. The relevant role of IL-1 beta overproduction as well as a generalised up-regulation of genes implicated in angiogenesis and inflammatory response are discussed in details.

The present study aims at verifying the suitability of in vitro cell models as useful tools to profile the toxicological behaviour of environment-occurring endocrine active compounds (EACs), to discriminate among different mixtures, to define endpoints that can be easily employed to assess the risk associated to the exposure as well as used as markers for prevention studies. A modified protocol of BALB/c 3T3 cell transformation was used to predict the toxicity and carcinogenic potential of some environmental EAC mixtures and to discriminate their behaviour from that of Aroclors. The role of environmental PCBs in the multistep carcinogenesis and the mechanism of action was also investigated by the gap junction cell communication (GJIC) assay and by evaluating the ability to induce apoptosis. Because of the estrogenic activity of these compounds, we also used a panel of in vitro mammary models to assess endpoints related to the endocrine disrupting activity. Two cell lines, MCF-7 human breast cancer cells and HuMi-TTu-2, a non commercial tumorigenic cell line, were chosen as eligible targets. The cell proliferation assay and the soft-agar assay (clonogenic assay) were elected as the endpoints to test commercial Aroclors and two phytoestrogens, genistein and ferutinin, which are regarded as promising chemopreventive agents able to counteract the developing of hormonal-dependent cancers. The results gave evidence that the proposed endpoints were very sensitive to the action of the tested mixtures and compounds. We were able to evaluate the dose-related toxicity and transforming activity of environmental EACs, their role in the multistep carcinogenesis and their estrogen-like effects in target cells. We also could highlight different toxicological behavior in commercial and environmental EACs and evaluate dose-response relationships, that are perhaps the most controversial issue regarding EACs. By the means of in vitro assays, we also demonstrated that ferutinin show the same hormetic dose-effect relationship, as previously described for genistein.

Nell’intento di individuare marcatori precoci di danno biologico, l’attività di ricerca si è focalizzata sull’analisi dell’espressione di Heat Shock Proteins ( HSP ) 27, 70, 90 e 110 in linfociti periferici di soggetti professionalmente esposti ad antiblastici. A tale scopo la modulazione dell’espressione di HSP 27, 70, 90 e 110 è stata valutata in Western Blot e quantificata tramite analisi densitometrica delle bande corrispondenti alle proteine in esame. I risultati ottenuti hanno dimostrato l’assenza di differenze statisticamente significative nell’espressione di HSP nei due gruppi a confronto, anche se però la scomposizione del gruppo dei casi in soggetti con monitoraggio biologico (mb) positivo e negativo ci ha consentito di evidenziare una diminuzione di HSP27 nel gruppo mb+ rispetto all’insieme mb- (p=0.029). L’analisi statistica secondaria ha inoltre evidenziato una correlazione positiva tra HSP27/bmi, suggerendo pertanto la possibilità di modulazione di parametri molecolari di stress da parte di un aumento di massa corporea. Lo studio dell’espressione di HSPs in risposta a Metotrexate (MTX) e Ciclofosfamide (CP) è stato approfondito in colture murine dove si è dimostrata una spiccata inibizione del rate proliferativo delle cellule BALB/c3T3 ad opera della sola MTX associata a una diminuzione di espressione di HSP 27, mentre non si sono osservate modulazioni dei marcatori in esame da parte CP a nessuno dei tempi valutati. I risultati ottenuti suggeriscono quindi, in accordo con quanto rilevato nei casi con monitoraggio biologico positivo, una correlazione inversa tra espressione di HSP27 ed esposizione ad agenti antiblastici evidenziando un possibile ruolo dei marcatori heat-shock nella risposta ad antiblastici.

Resveratrol (trans-3,4’,5-trihydroxystibene, Res), a phytoalexin found in grapes, elicits antioxidant and anti-inflammatory activity and has been shown to inhibit the multistage carcinogenesis process. The present study aims at verifying the potential chemopreventive activity of Res by a pattern of in vitro tests. The modified BALB/c 3T3 cell transformation assay was performed in order to evaluate the ability of resveratrol to inhibit the 3-MCA induced cell transformation, by acting as a suppressing agent. Aiming of analyzing possible cytotoxic effects, Res was previously tested in a wide range of concentrations (10-9-10-4 M) to assess their ability to affect the plating efficiency and the proliferation rate of BALB/c 3T3 clone A31 cells. In the anti-transformation assay, carcinogen-treated BALB/c-3T3 cells were chronically exposed to various doses of Res (10-7- 10-5 M) throughout the duration of the experiment. The chronic treatment significantly reduced the transformation frequency of 3-MCA–treated BALB/c 3T3 cells and the inhibition of foci occurrence was related to the assayed dose. The effects of Res on the proliferation rates of malignant cells were also evaluated. Because of the estrogenic activity of Res, human breast (MCF-7 and HuMi-TTu-2) and colon carcinoma (HT29 and Caco-2) cell lines, differing for the estrogen receptor expression, were chosen as eligible targets. Based on the cell viability assay, Res induced a dose-dependent inhibitory effect on the growth of both mammary cell lines with a GI50 of ~ 7.5 x 10-5 µM, whereas the GI50 calculated for the colon cell lines was ~ 5 x 10-5 µM. The toxicological profile, evaluated in both cytotoxicity and proliferation assay, was strongly suggestive of apoptosis induction. Anchorage independent growth has been widely used as a marker of neoplastic transformation and is closely related to cell tumorigenicity. Res induced a dose-related suppression of the clonogenicity of the assayed cell lines. 10-5 µM Res, the effective dose in the anti-transformation assay, was also able to exert a significant inhibition of the cell ability to colonize soft agar. Results give evidence for the feasibility of the proposed in vitro tests battery in vitro screening of candidate chemopreventive agents and for qualifying these compounds to further evaluation in clinical studies.