Cyclophosphamide levels in sites of preparation and administration of antineoplastic drugs

The extensive use of antineoplastic agents in chemotherapy may be at risk to health care workers involved in the preparation and administration of these drugs. In this study cyclophosphamide, a drug classified as a human carcinogen, was quantified by adapting a previous analytical method using gas chromatography coupled to mass spectrometry (GC-MS) after solid phase extraction with diatomaceous earth. The drug was measured by analysis in surfaces (wipe samples) and gloves, collected from four different hospitals, before and after the practice of cleaning procedures, and the use of a closed-system device for the preparation and administration. Validation results were satisfactory and cyclophosphamide levels ranging from below the quantification limit to 141000 ng. Our findings demonstrated that surfaces and materials contamination was found in all hospitals during the traditional open technique for preparation and administration of cyclophosphamide and a significant reduction in contamination when a closed-system device was used. However, some values were considered unexpected, especially those obtained from samples collected after the cleaning surfaces.

Does Cyclophosphamide Play a Protective Role Against Neuronal Loss in Chronic T. cruzi Infection?

In this study, we verified the possible role of cyclophosphamide (CY) in protecting or not against neuronal losses in young and aged male Calomys callosus chronically infected with the MORC-1 strain of Trypanosoma cruzi through numerical quantification of neurons from the myenteric plexus of the colon and quantification of nitric-oxide concentration (NO) during the acute and chronic phase of infection. For this purpose, groups of young C. callosus were infected with the MORC-1 strain of T. cruzi. A group of infected animals received i.p. 0.2 mg/ml genuxal dissolved in distilled water treatment with CY. NO concentration in aged animals displayed reduced levels when compared to those found in young animals. No significant alterations in the number of neurons were observed in young animals, but for aged ones, a protective role of CY in reducing neuron loss was noted, in addition to enhancing the neuronal volume, area, and perimeter. These results suggest that CY administration, depending on the dose and time span, can act as a protective agent against neuronal losses.

Could Cyclophosphamide exert a protective role avoiding esophagic neuron loss in Calomys callosus infected with Trypanosoma cruzi?

The protective role of Cyclophosphamide was studied in this work. Young male Calomys callosus were infected with Trypanosoma cruzi and allowed to age. Cyclophosphamide therapy was administered to animals during acute and late chronic phases of infection. Esophageal neurons were counted, displaying enhanced neuronal loss for the young and treated infected groups. For aged and cyclophosphamide treated animals, a protection was observed through a reduced loss of neurons as compared to the young and infected groups. Enhanced nitric oxide concentrations were observed for young animals as compared to aged counterparts. Splenocyte proliferation was reduced during the acute phase in comparison with those found in the chronic phase. Morphometry of neuronal body displayed a significant reduction concerning the area, perimeter, diameter and volume for aged animals as compared to young groups. These results indicate that the protective effects of cyclophosphamide together with process of neuroplasty of peripheral nervous system could lead to a protection against neuronal loss.

Gene expression profile of residual breast cancer after doxorubicin and cyclophosphamide neoadjuvant chemotherapy

In breast cancer patients, primary chemotherapy is associated with the same survival benefits as adjuvant chemotherapy. Residual tumors represent a clinical challenge, Lis they may be resistant to additional cycles of the same drugs. Our aim was to identify differential transcripts expressed in residual tumors, after neoadjuvant chemotherapy, that might be related with tumor resistance. Hence, 16 patients with paired tumor samples, collected before and after treatment (4 cycles doxorubicin/cyclophosphamide, AC) had their gene expression evaluated on cDNA microarray slides containing 4,608 genes. Three hundred and eighty-nine genes were differentially expressed (paired Student`s t-test, pFDR<0.01) between pre- and post-chemotherapy samples and among the regulated functions were the JNK cascade and cell death. Unsupervised hierarchical clustering identified one branch comprising exclusively, eight pre-chemotherapy samples and another branch, including the former correspondent eight post-chemotherapy samples and other 16 paired pre/post-chemotherapy samples. No differences in clinical and tumor parameters could explain this clustering. Another group of I I patients with paired samples had expression of selected genes determined by real-time RT-PCR and CTGF and DUSP1 were confirmed more expressed in post- as compared to pre-chemotherapy samples. After neoadjuvant chemotherapy some residual samples may retain their molecular signature while others present significant changes in their gene expression, probably induced by the treatment. CTGF and DUSP1 overexpression in residual samples may be a reflection of resistance to further administration of AC regimen.

Influence of Glomerular Filtration Rate on the Pharmacokinetics of Cyclophosphamide Enantiomers in Patients With Lupus Nephritis

The pharmacokinetics of cyclophosphamide (CYC) enantiomers were evaluated in patients with lupus nephritis distributed in 2 groups according to creatinine clearance: group 1 (90.6-144.6 mL/min/1.73 m(2)) and group 2 (42.8-76.4 mL/min/ 1.73 m(2)). All patients were treated with 0.75 to 1.3 g of racemic CYC as a 2-hour infusion and with 1 mg intravenous midazolam as a drug-metabolizing marker. CYC enantiomers and midazolam concentrations in plasma were measured by liquid chromatography/tandem mass spectrometry (LC/MS/MS). The following differences (Wilcoxon test, P <= .05) were observed between the (S)-(-) and (R)-(+) enantiomers: AUC(0-infinity) 152.41 vs 129.25 mu g.h/mL, CL 3.28 vs 3.89 L/h, Vd 31.38 vs 29.74 L, and t(1/2) 6.79 vs 5.56 h for group 1 and AUC(0-infinity) 167.20 vs 139.08 mu g.h/mL, CL 2.99 vs 3.59 L/h, and t(1/2) 6.15 vs 4.99 h for group 2. No differences (Mann test, P <= .05) were observed between groups 1 and 2 in the pharmacokinetic parameters of both enantiomers. No significant relationship was observed between midazolam clearance (2.92-16.40 mL/min.kg) and clearance of each CYC enantiomer. In conclusion, CYC kinetic disposition is enantioselective, resulting in higher exposures of the (S)-(-) enantiomer in lupus nephritis patients, and the pharmacokinetic parameters of both enantiomers are not altered by the worsening of renal condition.

Determination of Cyclophosphamide Enantiomers in Plasma by LC-MS/MS: Application to Pharmacokinetics in Breast Cancer and Lupus Nephritis Patients

This article describes the enantioseleclive analysis of cyclophosphamide (CPA) in human plasma using LC-MS/MS. CPA enantiomers were extracted from plasma using a mixture of ethyl acetate and chloroform (75:25, v/v). The enantiomers were separated on a Chiralcel(R) OD-R column, with the mobile phase consisting of a mixture of acetonitrile and water (75:25, v/v) plus 0.2% formic acid. The protonaled ions and their respective product ions were monitored using two functions, 261 > 141 for CPA enantiomers and 189 > 104 for the internal standard (antipyrine). Recovery rates were higher than 95% and the quantification limit was 2.5-ng/ml plasma for both enantiomers. The coefficients of variation and the relative errors obtained for the validation of intra- and interassay precision and accuracy were less than 10%. The method was applied for the investigation of the enantioselective pharmacokinetics of CPA in a lupus nephritis patient treated with 1 g CPA infused over 2 h and in a breast cancer patient treated with 0.9 g infused over 1 h. No stereoselectivity in the pharmacokinetic parameters was observed for either patient. Clearance values of 2.63 and 2.93 l/h and of 3.36 and 3.61 l/h for (-)-(S) and (+)-(R)-CPA were obtained for the breast cancer and lupus nephritis patient., respectively. Chirality 21:383-389, 2009. (C) 2008 Wiley-Liss, Inc.

Cyclophosphamide effect on coccidioidomycosis in the rat

Coccidioidomycosis is a systemic mycosis, endemic in arid areas of the American continent. The rat was employed as an experimental host, since it had been shown to reproduce human lesions and present a chronic course of disease with granulomas mainly restricted to lungs. Given the influence of immunosuppressive therapy on the clinical course of human coccidioidomycosis, we studied the effect of cyclophosphamide (CY) in the experimental rat model. Accordingly, animals were inoculated with 400 Coccidioides immitis arthroconidia of the Acosta strain, by intracardiacal route. As single CY doses failed to alter the course of disease, three schedules were used: A) 4 daily doses of 20 mg/kg each, prior to C. immitis inoculation; B) 4 similar daily doses after infection; and C); 6 doses of 20 mg/kg each, given from day +1 to +4, then on days +8 and +9, post infection (pi), taking day 0 as the time of fungal inoculation. The first two schedules inhibited antibody formation up to day 28 pi, without modifying cellular response to coccidioidin as measured by foodpad swelling. Initially, there was greater fungal spread than in controls receiving C. immitis alone, which proved self-limiting in the latter. In contrast, schedule C led to 559r mortality, with both humoral and cellular response abrogation, accompanied by extensive C. immitis dissemination. Histology disclosed significant alterations, such as the persistence of primary infection sporangia, corresponding to the acute stage of coccidioidomycosis in the absence of granuloma development. Therefore, the observed depression in cellular immunity seems responsible for the lack of inflammatory reaction capable of restricting sporangia proliferation in tissues which, in turn, enhances pathogen spread and mortality rate.

Evolution of subpatent parasitaemia in Trypanosoma cruzi chronically infected mice with the help of a cyclophosphamide amplification transfer assay

We have evaluated the sensitivity of the classical blood subinoculation method, modified through cyclophosphamide treatment of transferred mice, for the detection of occult parasitaemias in Trypanosoma cruzi chronically infected mice. Besides its simplicity, the method was shown to be highly sensitive for both the "chronic" phase parasites (99% of chronic cases were shown to harbour occult parasitaemias) and for the acute phase parasites (T. cruzi could be detected in 53.8% of animals transferred with one Y strain parasite and in 20% of animals transferred with one CL strain parasite). Using acute phase bloodforms, the assay proved to be more sensitive than conventional subinoculation when dealing with the CL, but not the Y strain of the parasite. With the help of this parasite detection tool, we have studied during a one year period, the evolution of subpatent parasitaemias in a group of mice which survived through chemotherapy from lethal acute phase of T. cruzi infection. Cyclophosphamide transfer assay revealed occult parasitaemias in 100% of the chronic animals, nevertheless, continuous and discontinuous patterns of positivity were observed.

Modulation of parasitemia and antibody responce to Trypanosoma cruzy by cyclophosphamide in Calomys callosus (Rodentia, Cricetidae)

Calomys callosus a wild rodent, previously described as harboring Trypanosoma cruzi, has a low susceptibility to infection by this protozoan. Experiments were designed to evaluate the contribution of the immune response to the resistance to T. cruzi infection exhibited by C. calossus. Animals were submitted to injections of high (200 mg/kg body weight) and low (20 mg/kg body weight) doses of cyclophosphamide on days -1 or -1 and +5, and inoculated with 4 x 10³ T. cruzi on day O. Parasitemia, mortality and antibody response as measured by direct agglutination of trypomastigotes were observed. Two hundred mg doses of cyclophosphamide resulted in higher parasitemia and mortality as well as in suppression of the antibody response. A single dose of 20 mg enhanced antibody levels on the 20th day after infection, while an additional dose did not further increase antibody production. Parasitemia levels were not depressed, but rather increased in both these groups as compared to untreated controls. Passive transfer of hyperimmune C. callosus anti-T. cruzi serum to cyclophosphamide immunosuppressed animals resulted in lower parasitemia and mortality rates. These results indicate that the immune response plays an important role in the resistance of C. callossus to T. cruzi.

Cyclophosphamide effect on paracoccidioidomycosis in the rat

Paracoccidioidomycosis is an endemic fungal disease widely distributed throughout Latin America. The potent immunosuppressor cyclophosphamide (CY) has been used to modulate host immune response to Paracoccidioides brasiliensis in an experimental model. Inbred male Buffalo/Sim rats weighing 250-300 g were inoculated with 5 x 10(6) P. brasiliensis cells of the yeast phase form by intracardiac route. One group of animals was treated with 20 mg/kg body weight at days +4, +5, +6, +7, +11 and +12 post-infection (pi.), while a control group was infected alone. No mortality was recorded in either group. Treated rats presented: a) a decrease in granuloma size, which contained less fungal cells; b) a lack of specific antibodies up to 35 days pi., and c) a significant increase in the footpad swelling test (DTH) against paracoccidioidin. Splenic cell transfer from CY-treated P. brasiliensis-infected donors to recipients infected alone led to a significant increase in DTH response in the latter versus untreated infected controls. Likewise, in treated infected recipients transferred with untreated infected donor spleen cells, footpad swelling proved greater than in controls. Thus, it would seem that each successive suppressor T lymphocyte subset belonging to the respective cascade may be sensitive to repeated CY doses administered up to 12 days pi.. Alternatively, such CY schedule may induce the appearance of a T cell population capable of amplifying DTH response.

Inhibitory effect of cyclophosphamide on the biosynthesis of a perchloro-soluble protein fraction of ehrlich ascites carcinoma cells

The perchloro-soluble mucroptotein fraction was determined in the cells of Ehrlich ascites carcinoma on the 10th and 12th days post-inoculation of the tumor. After 3 days of a single subcutaneous dose of cyclophosphamide (200 mg/kg) the mucoprotein levels were found considerable lower. This difference was highly significant statistically.

Recrudescence induced by cyclophosphamide of chronic Trypanosoma cruzi infection in mice is influenced by the parasite strain

Reactivation of chronic chagasic patients may occur upon use of immunosuppressive drugs related to kidney or heart transplantation or when they are affected by concomitant HIV infection. This recrudescence, however, does not occur in all chagasic patients exposed to immunosuppressive agents. We therefore investigated the influence of Trypanosoma cruzi strains in the recrudescence of the parasitism in mice at the chronic phase treated with cyclophosphamide, an immunosuppressor that blocks lymphocytes DNA synthesis and therefore controls B cells response. A large variation was detected in the percentages of newly established acute phases in the groups of mice inoculated with the different strains. We suggest that reactivation of chronic T. cruzi infections is influenced by the parasite intrinsic characteristics, a phenomenon that might occur in the human disease.

Shrinking lung syndrome successfully treated with rituximab and cyclophosphamide.

Shrinking lung syndrome (SLS) is an uncommon feature of systemic lupus erythematosus (SLE) characterized by dyspnea, pleuritic chest pain, diaphragmatic elevation, restrictive ventilatory defect and reduced respiratory muscle strength as measured by volitional tests. We report the case of a 28-year-old woman with overlapping features of SLE and Sjögren syndrome who developed severe SLS while receiving corticosteroids and azathioprine for severe polyarthritis. She was treated with a combination of rituximab and cyclophosphamide, which led to a dramatic improvement in her clinical condition and respiratory function tests. The increase in vital capacity was one of the highest among 35 published cases of SLS. Thus, restoring a near-normal lung function is an achievable goal in SLS, and the use of rituximab, with or without concomitant cyclophosphamide, certainly deserves further study in this setting.

Randomized phase 3 trial of fluorouracil, epirubicin, and cyclophosphamide alone or followed by Paclitaxel for early breast cancer.

BACKGROUND Taxanes are among the most active drugs for the treatment of metastatic breast cancer, and, as a consequence, they have also been studied in the adjuvant setting. METHODS After breast cancer surgery, women with lymph node-positive disease were randomly assigned to treatment with fluorouracil, epirubicin, and cyclophosphamide (FEC) or with FEC followed by weekly paclitaxel (FEC-P). The primary endpoint of study-5-year disease-free survival (DFS)-was assessed by Kaplan-Meier analysis. Secondary endpoints included overall survival and analysis of the prognostic and predictive value of clinical and molecular (hormone receptors by immunohistochemistry and HER2 by fluorescence in situ hybridization) markers. Associations and interactions were assessed with a multivariable Cox proportional hazards model for DFS for the following covariates: age, menopausal status, tumor size, lymph node status, type of chemotherapy, tumor size, positive lymph nodes, HER2 status, and hormone receptor status. All statistical tests were two-sided. RESULTS Among the 1246 eligible patients, estimated rates of DFS at 5 years were 78.5% in the FEC-P arm and 72.1% in the FEC arm (difference = 6.4%, 95% confidence interval [CI] = 1.6% to 11.2%; P = .006). FEC-P treatment was associated with a 23% reduction in the risk of relapse compared with FEC treatment (146 relapses in the 614 patients in the FEC-P arm vs 193 relapses in the 632 patients in the FEC arm, hazard ratio [HR] = 0.77, 95% CI = 0.62 to 0.95; P = .022) and a 22% reduction in the risk of death (73 and 95 deaths, respectively, HR = 0.78, 95% CI = 0.57 to 1.06; P = .110). Among the 928 patients for whom tumor samples were centrally analyzed, type of chemotherapy (FEC vs FEC-P) (P = .017), number of involved axillary lymph nodes (P < .001), tumor size (P = .020), hormone receptor status (P = .004), and HER2 status (P = .006) were all associated with DFS. We found no statistically significant interaction between HER2 status and paclitaxel treatment or between hormone receptor status and paclitaxel treatment. CONCLUSIONS Among patients with operable breast cancer, FEC-P treatment statistically significantly reduced the risk of relapse compared with FEC as adjuvant therapy.