Protective effects of cerebrolysin against chemotherapy (carmustine) induced cognitive impairment in Albino mice.

Chemotherapy-induced cognitive impairment (CICI) comprises different neurological problems, including difficulty in learning new things, concentrating and making decisions that affect daily life activities. Clinical reports indicate that around 70% of cancer patients receiving chemotherapy suffer from cognitive impairment. The purpose of the present study is to examine the effects of widely used anticancer medication (Carmustine) on cognitive function using mice model and investigation of the neuroprotective effects of Cerebrolysin (CBN). Cerebrolysin (CBN) is a mixture of several neurotrophic factors and active peptides with anti-inflammatory, antioxidant, and neuroprotective actions. Our study aimed to establish a mice model of Carmustine (BCNU)-induced cognitive deficits and determine the protective effects of CBN. BCNU (10 mg/kg, i.v.) was administered to mice for 28 days, and behavioral parameters were measured on a weekly basis. CBN (44 and 88 mg/kg, i.p.) was administered daily from day 1 to 28 to BCNU treatment mice. All animals were sacrificed on day 29 and brain hippocampus tissues were used for biochemical, neuroinflammatory, neurotransmitters analysis. BCNU administration animals showed impaired cognition and memory, confirmed from behavioral analysis. Further, BCNU increased oxidative stress, inflammatory cytokines release and altered neurotransmitters concentration as compared to the control group (p < 0.01). However, mice treated with CBN (44 and 88 mg/kg, i.p.) significantly and dose-dependently improved cognitive functions, reduced oxidative stress markers, inflammatory cytokines and restored neurotransmitters concentration as compared to BCNU administered mice (p < 0.05). The finding of current study suggested that CBN could be the promising compound to reverse cognitive impairment associated with use of chemotherapy.

Role of deltaNp63(pos)CD44v(pos) cells in the development of N-nitroso-tris-chloroethylurea-induced peripheral-type mouse lung squamous cell carcinomas.

The role of cells expressing stem cell markers deltaNp63 and CD44v has not yet been elucidated in peripheral-type lung squamous cell carcinoma (pLSCC) carcinogenesis. Female A/J mice were painted topically with N-nitroso-tris-chloroethylurea (NTCU) for induction of pLSCC, and the histopathological and molecular characteristics of NTCU-induced lung lesions were examined. Histopathologically, we found atypical bronchiolar hyperplasia, squamous metaplasia, squamous dysplasia, and pLSCCs in the treated mice. Furthermore, we identified deltaNp63(pos)CD44v(pos)CK5/6(pos)CC10(pos) clara cells as key constituents of early precancerous atypical bronchiolar hyperplasia. In addition, deltaNp63(pos)CD44v(pos) cells existed throughout the atypical bronchiolar hyperplasias, squamous metaplasias, squamous dysplasias, and pLSCCs. Overall, our findings suggest that NTCU induces pLSCC through an atypical bronchiolar hyperplasia-metaplasia-dysplasia-SCC sequence in mouse lung bronchioles. Notably, Ki67-positive deltaNp63(pos)CD44v(pos) cancer cells, cancer cells overexpressing phosphorylated epidermal growth factor receptor and signal transducer and activator of transcription 3, and tumor-associated macrophages were all present in far greater numbers in the peripheral area of the pLSCCs compared with the central area. These findings suggest that deltaNp63(pos)CD44v(pos) clara cells in mouse lung bronchioles might be the origin of the NTCU-induced pLSCCs. Our findings also suggest that tumor-associated macrophages may contribute to creating a tumor microenvironment in the peripheral area of pLSCCs that allows deltaNp63(pos)CD44v(pos) cancer cell expansion through activation of epidermal growth factor receptor signaling, and that exerts an immunosuppressive effect through activation of signal transducer and activator of transcription 3 signaling.

Pivotal role of glutathione depletion in eNOS uncoupling of LPS-Treated HUVECs.

The present study investigated the relationship between uncoupling of endothelial nitric oxide synthase (eNOS) and vascular endothelial cell (VEC) oxidative stress (OS) during sepsis and the role of eNOS glutathionylation in eNOS uncoupling of septic VECs. Human umbilical vein endothelial cells (HUVECs) cultured in vitro (EA.hy269 cell line) were incubated with Dulbecco's modified Eagle's medium (DMEM) (normal control group), lipopolysaccharide (LPS) (sepsis group), 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) (glutathionylation group), and LPS+ dithiothreitol (DTT) (deglutathionylation sepsis group). As result, compared with the DMEM group, malondialdehyde (MDA) level and uncoupling eNOS activity significantly increased in the LPS and BCNU groups. However, in the LPS + DTT group, only the NO level increased. Compared with the LPS group, MDA level, NO concentration, and normal functional eNOS activity significantly decreased, and uncoupling eNOS activity significantly increased in the BCNU group. In the LPS + DTT group, MDA level and uncoupling eNOS activity significantly decreased, and NO concentration and normal functional eNOS activity significantly increased. During sepsis, the main mechanism for VEC OS was eNOS uncoupling mediated by eNOS glutathionylation.

Overexpression of Nrf2 attenuates Carmustine-induced cytotoxicity in U87MG human glioma cells.

BACKGROUND: Malignant glioma is one of the most devastating tumors in adults with poor patient prognosis. Notably, glioma often exhibits resistance to conventional chemotherapeutic approaches, complicating patient treatments. However, the molecular mediators involved in tumor chemoresistance remain poorly defined, creating a barrier to the successful management of glioma. In the present study, we hypothesized that the antioxidant transcription factor, Nrf2 (nuclear factor erythroid-derived 2 like 2), attenuates glioma cytotoxicity to Carmustine (BCNU), a widely used chemotherapeutic agent known to modulate cellular oxidative balance. METHODS: To test the hypothesis, we employed human malignant glioma cell line, U87MG and overexpression of Nrf2 in glioma cells was achieved using both pharmacological and genetic approaches. RESULTS: Notably, induction of Nrf2 was associated with increased expression of heme oxygenase-1 (HO-1), a stress inducible enzyme involved in anti-oxidant defense. In addition, over expression of Nrf2 in U87MG cells significantly attenuated the cytotoxicity of Carmustine as evidenced by both cellular viability assay and flow cytometry analysis. Consistent with this, antioxidants such as glutathione and N-acetyl cysteine significantly reduced Carmustine mediated glioma cytotoxicity. CONCLUSIONS: Taken together, these data strongly implicate an unexplored role of Nrf2 in glioma resistance to Carmustine and raise the possible use of Nrf2 inhibitors as adjunct to Carmustine for the treatment of malignant glioma.

Novel medium-term carcinogenesis model for lung squamous cell carcinoma induced by N-nitroso-tris-chloroethylurea in mice.

Targeted treatments for lung cancer based on pathological diagnoses are required to enhance therapeutic efficacy. There are few well-established animal models for lung squamous cell carcinoma although several highly reproducible mouse models for lung adenoma and adenocarcinoma are available. This study was carried out to establish a new lung squamous cell carcinoma mouse model. In the first experiment, female A/J mice were painted topically on back skin twice weekly with 75 µL 0.013 M N-nitroso-tris-chloroethylurea for 2, 4, and 8 weeks (n = 15-20 per group) as initiation of lung lesions, and surviving mice were killed at 18 weeks. In the second experiment, mice were treated as above for 4 weeks and killed at 6, 12, or 18 weeks (n = 3 per group). Lung lobes were subjected to histopathological, immunohistochemical, immunoblotting, and ultrastructural analyses. In the case of treatment for 2, 4, and 8 weeks, incidences of lung squamous cell carcinoma were 25, 54, and 71%, respectively. Cytokeratin 5/6 and epidermal growth factor receptor were clearly expressed in dysplasia and squamous cell carcinoma. Desmosomes and tonofilaments developed in the squamous cell carcinoma. Considering the carcinogenesis model, we conclude that 2 or 4 weeks of N-nitroso-tris-chloroethylurea treatment may be suitable for investigating new chemicals for promotional or suppressive effects on lung squamous cell carcinoma.

Long-Term Results with Thiotepa-Containing Conditioning Regimens for Autologous Stem Cell Transplantation.

Autologous stem cell transplantation (ASCT) remains a cornerstone in the treatment of both Hodgkin lymphoma (HL) and various non-Hodgkin lymphoma (NHL) subtypes. BEAM (carmustine, etoposide, cytarabine, and melphalan) is the most frequently used conditioning regimen; however, owing due to limited availability and toxicity of carmustine, thiotepa-containing regimens have been suggested. We previously reported encouraging results in ASCT with a TECAM (thiotepa, etoposide, cyclophosphamide, cytarabine, and melphalan) conditioning regimen from 2000 to 2013. We aimed to update our experience with the TECAM regimen by adding our experience from 2013 to 2020 to the previously reported cohort. Moreover, we aimed to use the detailed data for the 2 transplant cohorts to identify improvements in ASCT outcomes in the recent era. We retrospectively analyzed all lymphoma patients who underwent ASCT at our center between January 2000 and December 2020. A total of 353 lymphoma patients were included (142 in the newer cohort added to 211 previously reported patients), all of whom were treated with our standard TECAM conditioning regimen. The cohort included 127 patients with HL, 107 with DLBCL, and 119 with other NHL subtypes. The newer cohort was characterized by significantly poorer Eastern Cooperative Oncology Group Performance Status (ECOG-PS) prior to ASCT (45.7% versus 19.3% with ECOG-PS ≥1; P < .01), whereas a higher proportion of patients entered transplantation in complete response (CR) (71.9% versus 47.8%; P < .01). The median follow-up after ASCT was 136.4 months (95% confidence interval [CI], 91.4 to 181.4 months). The 3-year progression-free survival (PFS) and overall survival (OS) rates post-ASCT for the entire cohort were 59.8% and 79.3%, respectively. Evaluating the 303 of 353 patients (86.4%) who entered ASCT with a responsive disease-a population that represents today's approach to the selection of patients for ASCT-the 3-year PFS and OS rates were 61.5% and 81.9%, respectively. In this population, the 3-year PFS rate was 62.2% for HL, 62.6% for DLBCL, 64.3% for primary central nervous system lymphoma (PCNSL), and the 3-year OS rate were 90.1%, 75.2%, and 78.6%, respectively. OS was significantly better in the newer cohort (P < .01), but not when evaluating only patients who entered ASCT with responsive disease. Dose reductions, poor disease status, and poor ECOG-PS at ASCT entry were associated with worse outcomes across all lymphoma subtypes. In accordance with our previous report, patients entering transplantation for DLBCL with a partial response achieved similar outcomes as those with a CR. Eighteen patients died within the first 100 days, 8 due to disease progression and 10 due to transplantation-related complications (2.8%). There were no cases of interstitial pneumonitis syndrome. Twenty-two cases (6.2%) of secondary malignancies were documented. Our results confirm that TECAM is an effective and safe conditioning regimen for ASCT in patients with HL and various NHLs, including favorable results in PCNSL. Despite a higher proportion of frail patients, the newer cohort's outcomes were favorable, driven by better lymphoma control pretransplantation. In the DLBCL cohort, ECOG-PS had more prognostic value than achieving a CR pre-ASCT, a finding relevant to the optimal allocation of patients to different treatment options in the era of chimeric antigen receptor T cell availability.

Protection by Salvia extracts against oxidative and alkylation damage to DNA in human HCT15 and CO115 cells.

DNA damage induced by oxidative and alkylating agents contributes to carcinogenesis, leading to possible mutations if replication proceeds without proper repair. However, some alkylating agents are used in cancer therapy due to their ability to induce DNA damage and subsequently apoptosis of tumor cells. In this study, the genotoxic effects of oxidative hydrogen peroxide (H2O2) and alkylating agents N-methyl-N-nitrosourea (MNU) and 1,3-bis-(2-chloroethyl)-1-nitosourea (BCNU) agents were examined in two colon cell lines (HCT15 and CO115). DNA damage was assessed by the comet assay with and without lesion-specific repair enzymes. Genotoxic agents were used for induction of DNA damage in both cell lines. Protective effects of extracts of three Salvia species, Salvia officinalis (SO), Salvia fruticosa (SF), and Salvia lavandulifolia (SL), against DNA damage induced by oxidative and alkylating agents were also determined. SO and SF protected against oxidative DNA damage in HCT15 cells. SO and SL decreased DNA damage induced by MNU in CO115 cells. In addition to chemopreventive effects of sage plant extracts, it was also important to know whether these plant extracts may interfere with alkylating agents such as BCNU used in cancer therapy, decreasing their efficacy. Our results showed that sage extracts tested and rosmarinic acid (RA), the main constituent, protected CO115 cells from DNA damage induced by BCNU. In HCT15 cells, only SF induced a reduction in BCNU-induced DNA damage. Sage water extracts and RA did not markedly change DNA repair protein expression in either cell line. Data showed that sage tea protected colon cells against oxidative and alkylating DNA damage and may also interfere with efficacy of alkylating agents used in cancer therapy.

Protective role of the dried white mulberry extract on the reproductive damage and fertility in rats treated with carmustine.

The present study investigated the protective effect of dried white Mulberry extract (DWME) against carmustine (Crm) induced biochemical alterations and spermatological, histopathological, and fertility damage in Wistar albino rats. Male rats were divided into four groups (control, Crm, Crm + DWME, and DWME group). It was found that Crm decreased the motility. Crm decreased the concentration (not different from control group) compared to DWME groups. Total blood MDA levels were reduced during the recovery period. Also, the recovery period reduced the MDA levels in the Crm group/testicular tissue. The GSH levels in the Crm + DWME group were the highest among all groups in the testicular tissue/experiment period. In the immunohistochemical evaluation of the testicular tissue, a high level of caspase-3 was observed in the cells that underwent meiosis in the Crm group. The most pronounced DNA damage was also detected in the Crm group. The Crm + DWME group showed the highest number of offspring born during recovery period. In conclusion, dried white mulberry extract protects against the spermatological damages caused by carmustine. Moreover, recovery period played a positive effect on spermatological parameters and fertility.

An Improved Murine Premalignant Squamous Cell Model: Tobacco Smoke Exposure Augments NTCU-Induced Murine Airway Dysplasia.

Tobacco smoke-induced squamous cell lung cancer (SCC) develops from endobronchial dysplastic lesions that progress to invasive disease. A reproducible murine model recapitulating histologic progression observed in current and former smokers will advance testing of new preventive and therapeutic strategies. Previous studies show that prolonged topical application of N-nitroso-tris-chloroethylurea (NTCU) generates a range of airway lesions in sensitive mice similar to those induced by chronic tobacco smoke exposure in humans. To improve the current NTCU model and better align it with human disease, NTCU was applied to mice twice weekly for 4-5 weeks followed by a recovery period before cigarette smoke (CS) or ambient air (control) exposure for an additional 3-6 weeks. Despite the short time course, the addition of CS led to significantly more premalignant lesions (PML; 2.6 vs. 0.5; P < 0.02) and resulted in fewer alveolar macrophages (52,000 macrophages/mL BALF vs. 68,000; P < 0.05) compared with control mice. This improved NTCU + CS model is the first murine SCC model to incorporate tobacco smoke and is more amenable to preclinical studies because of the increased number of PML, decreased number of mice required, and reduced time needed for PML development.

Investigation of maternal melatonin effect on the hippocampal formation of newborn rat model of intrauterine cortical dysplasia.

OBJECTIVES: Cortical dysplasia is a cortical malformation resulting from any developmental defects during different periods of development. This study aims to investigate the hippocampal histopathological alterations in the neonates with cortical dysplasia due to the prenatal exposure to carmustine (1,3-bis (2-chloroethyl)-1-nitrosourea; BCNU) and the possible effects of prophylaxis with melatonin, a neuroprotective agent. METHODS: Wistar albino female rats were randomly divided into four experimental groups; control, melatonin-treated, BCNU-exposed and BCNU-exposed+melatonin-treated. Light microscopy and immunohistochemistry were carried out on the newborn hippocampus. RESULTS: Histopathology of hippocampus from the control and melatonin-treated groups showed continuity of migration and maturation as pathognomonic signs of the normal newborn hippocampus. Hippocampal cortex from the newborns exposed in utero to BCNU showed the histology of early embryonic hippocampal formation with immunohistochemical increase in the number of nestin positive cells and decreases in the immunoreactivity of glial fibrillary acidic protein (GFAP) and synaptophysin. These findings indicate a significant delay in hippocampal maturation, migration, and synaptogenesis. Intrauterine treatment of BCNU-exposed rats with melatonin resulted in histopathological features almost similar to control group. CONCLUSION: It has been concluded that cortical dysplasia induced by intrauterine BCNU administration results in delayed hippocampal maturation, which is successfully restored by intrauterine melatonin treatment.

TRPC5 channel instability induced by depalmitoylation protects striatal neurons against oxidative stress in Huntington's disease.

Protein S-palmitoylation, the covalent lipid modification of the side chain of Cys residues with the 16­carbon fatty acid palmitate, is the most common acylation, and it enhances the membrane stability of ion channels. This post-translational modification (PTM) determines a functional mechanism of ion channel life cycle from maturation and membrane trafficking to localization. Especially, neurodevelopment is regulated by balancing the level of synaptic protein palmitoylation/depalmitoylation. Recently, we revealed the pathological role of the transient receptor potential canonical type 5 (TRPC5) channel in striatal neuronal loss during Huntington's disease (HD), which is abnormally activated by oxidative stress. Here, we report a mechanism of TRPC5 palmitoylation at a conserved cysteine residue, that is critical for intrinsic channel activity. Furthermore, we identified the therapeutic effect of TRPC5 depalmitoylation by enhancing the TRPC5 membrane instability on HD striatal cells in order to lower TRPC5 toxicity. Collectively, these findings suggest that controlling S-palmitoylation of the TRPC5 channel as a potential risk factor can modulate TRPC5 channel expression and activity, providing new insights into a therapeutic strategy for neurodegenerative diseases.

Bone marrow-derived cells exhibiting lung epithelial cell characteristics are enriched in vivo using methylguanine DNA methyltransferase-mediated drug resistance.

Previous studies have suggested that donor bone marrow-derived cells can differentiate into lung epithelial cells at low frequency. We investigated whether we could enrich the number of donor-derived hematopoietic cells that have type II pneumocyte characteristics by overexpression of the drug resistance gene methylguanine DNA methyltransferase (MGMT). MGMT encodes O(6)-alkylguanine DNA alkyltransferase (AGT), a drug resistance protein for DNA damage induced by N,N'-bis(2-chloroethyl)-N-nitrosourea (BCNU), and the mutant P140K MGMT confers resistance to BCNU and the AGT inactivator O(6)-benzylguanine (BG). For this study, we used two MGMT selection models: one in which donor cells had a strong selection advantage because the recipient lung lacked MGMT expression, and another in which drug resistance was conferred by gene transfer of P140K MGMT. In both models, we saw an increase in the total number of donor-derived cells in the lung after BCNU treatment. Analysis of single-cell suspensions from 28 mice showed donor-derived cells with characteristics of type II pneumocytes, determined by surfactant protein C (SP-C) expression. Furthermore, an increase in the percentage of donor-derived SP-C cells was noted after BCNU or BG and BCNU treatment. This study demonstrates that bone marrow cells expressing MGMT can engraft in the lung and convert into cells expressing the type II pneumocyte protein SP-C. Furthermore, these cells can be enriched in response to alkylating agent-mediated lung injury. These results suggest that expression of MGMT could enhance the capacity of bone marrow-derived cells to repopulate lung epithelium, and when used in combination with a gene of interest, MGMT could have therapeutic applications.

Vitamin E reversal of the effect of extracellular calcium on chemically induced toxicity in hepatocytes.

Isolated rat hepatocytes were incubated in the presence or absence of extracellular calcium and alpha-tocopherol succinate with three different toxic chemicals; namely, adriamycin in combination with 1,3-bis(2-chloroethyl)-1-nitrosourea, ethyl methanesulfonate, and the calcium ionophore A23187. In the absence of extracellular calcium these three compounds were far more toxic to the cells than in its presence. The addition of vitamin E to calcium-free medium, however, protected hepatocytes against toxic injury, whereas cells incubated in medium containing calcium were not protected. Hepatocyte viability during each toxic insult correlated well with the cellular alpha-tocopherol content but not with the presence or absence of extracellular calcium. These results suggest that cellular alpha-tocopherol maintains the viability of the cell during a toxic insult and that the presence or absence of vitamin E in the incubation medium probably explains the conflicting reports on the role of extracellular calcium in toxic cell death.

Metallothionein attenuates carmustine-induced oxidative stress and protects against pulmonary fibrosis in rats.

The present study was carried out to evaluate the effect of exogenously administered metallothionein (MT) against carmustine (BCNU)-induced lung toxicity in rats. A total of 60 rats were randomly divided into four groups (15/group): control group in which the animals received 0.5 ml physiologic saline containing 10% ethanol (IP) weekly, MT-administered group in which rats received MT (30 micromol/kg, IP) weekly, BCNU-administered group in which rats received BCNU (5 mg/kg, IP) weekly and MT + BCNU group in which rats received weekly doses of BCNU (5 mg/kg, IP) followed 24 h later by MT (30 micromol/kg, IP). At the end of the experiment (after 6 weeks), lung histological changes, collagen staining, the activity of glutathione reductase (GR) and contents of reduced glutathione (GSH) and hydroxyproline (Hpr) in the lung as well as serum level of tumor necrosis factor-alpha (TNF-alpha) were evaluated. The obtained data revealed that BCNU induced pathological changes and markedly increased lung collagen and level of Hpr but decreased GSH content and GR activity and increased serum TNF-alpha compared to both control and MT-administered rats. Administration of MT + BCNU markedly improved histological features and decreased staining of collagen along with increased GR activity, GSH content but decreased level of Hpr in lung tissue as well as decreased serum level of TNF-alpha compared with BCNU-treated rats. Based on our results, it is possible to postulate that exogenous MT can act against BCNU-induced lung toxicity by a mechanism related, at least in part, to its ability to decrease oxidative stress and fibrosis.

The Effect of Mouse Strain, Sex, and Carcinogen Dose on Toxicity and the Development of Lung Dysplasia and Squamous Cell Carcinomas in Mice.

In order to translate new treatments to the clinic, it is necessary to use animal models that closely recapitulate human disease. Lung cancer develops after extended exposure to carcinogens. It has one of the highest mutation rates of all cancer and is highly heterogenic. Topical treatment with N-nitrosotris-(2-chloroethyl)urea (NTCU) induces lung squamous cell carcinoma (SCC) with nonsynonymous mutation rates similar to those reported for human non-small cell lung cancer. However, NTCU induces lung cancer with variable efficacy and toxicity depending on the mouse strain. A detailed characterization of the NTCU model is needed. We have compared the effect of three different NTCU doses (20, 30, and 40 mmol/L) in female and male of NIH Swiss, Black Swiss, and FVB mice on tumor incidence, survival, and toxicity. The main findings in this study are (1) NIH Swiss mice present with a higher incidence of SCC and lower mortality compared with Black Swiss and FVB mice; (2) 30 mmol/L NTCU dose induces SCC at the same rate and incidence as the 40 mmol/L dose with lower mortality; (3) female mice present higher grade and incidence of preinvasive lesions and SCC compared with males; (4) NTCU-induced transformation is principally within the respiratory system; and (5) NTCU treatment does not affect the ability to elicit a specific adaptive immune response. This study provides a reference point for experimental designs to evaluate either preventive or therapeutic treatments for lung SCC, including immunotherapies, before initiating human clinical trials.

BCNU toxicity presenting with a large pericardial and pleural effusion.

BACKGROUND: Carmustine (BCNU), 1, 3-bis (2-chloroethyl)-1-nitosourea, is an alkylating agent which is commonly used as a part of conditioning regimen for autologous peripheral blood stem cell transplantation. Despite the widespread use of BCNU therapy in adults, cardiopulmonary toxicity presenting with cardiac tamponade has not been reported. CASE REPORT: We present a patient who received BCNU as part of her conditioning regimen followed by autologous peripheral blood stem cell transplantation. This patient subsequently developed bilateral upper lobe pulmonary infiltrates with bilateral pleural effusions and a large pericardial effusion which was a result of BCNU toxicity. CONCLUSIONS: Early institution of glucocorticoids for patients receiving high dose BCNU containing chemotherapeutic regimens has shown to significantly reduce BCNU induced cardio-pulmonary toxicity.

Tamoxifen accelerates proteasomal degradation of O6-methylguanine DNA methyltransferase in human cancer cells.

Tamoxifen, a synthetic triphenyl-ethylene compound, is a member of a class of anticancer drugs known as selective estrogen receptor modulators. It may block tumor growth by mimicking estrogen and binding to the estrogen receptors, preventing cancerous growth. Clinical studies have demonstrated that a combination chemo/hormonal therapy regimen with tamoxifen and O(6)-alkylating drugs increased the tumor response rate in cancer patients. The mechanism of action of this combined regimen remains undefined. In this study, we demonstrated that treatment of human colorectal HT-29 carcinoma cells with tamoxifen decreased the repair activity and expression level of O(6)-methylguanine DNA methyltransferase (MGMT) protein in a concentration- and time-dependent manner. This inhibition was also shown in other malignant human cells, regardless of their estrogen receptor status. Furthermore, MGMT inactivation by tamoxifen was associated with a significantly increased susceptibility of cells to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). No alteration in MGMT mRNA levels was observed in tamoxifen-treated cells. The half-life of MGMT protein was markedly decreased in the presence of tamoxifen. Tamoxifen-induced MGMT degradation could be blocked by MG-132, a proteasome inhibitor. An increased level of ubiquitinated MGMT protein was found after tamoxifen treatment. We conclude that tamoxifen decreased the MGMT protein level by accelerating protein degradation through the ubiquitin-dependent proteasomal pathway. These findings provide a strong rationale for combined chemo/hormonal therapy with tamoxifen and BCNU in the treatment of human cancers.

O6-benzylguanine and BCNU in multiple myeloma: a phase II trial.

PURPOSE: Carmustine (BCNU) is known to have modest activity in multiple myeloma; however, resistance to BCNU manifests by the activity of O6-methylguanine methyltransferase (MGMT). The objective of this study was to determine the safety and efficacy of depletion of MGMT activity in plasma cells using O6-benzylguanine (O6-BG) with BCNU in patients with multiple myeloma. METHODS: Patients with previously treated or untreated multiple myeloma were eligible. Cycles of O6-BG at a dose of 120 mg/m2 and BCNU at a dose of 40 mg/m2 were repeated every 6 weeks. RESULTS: Seventeen patients were enrolled on the study, with a median follow-up of 24.5 (range 5-69) months. One complete response (7%) and 3 partial responses (20%) were observed. Nine patients (60%) had stable disease. Bone marrow studies demonstrated 94% depletion of MGMT activity in CD38+ marrow cells. The most frequent grade 3 and 4 adverse events were neutropenia (71%), lymphocytopenia (53%), and thrombocytopenia (53%). CONCLUSIONS: Chemotherapy utilizing the MGMT inhibitor O6-benzylguanine and BCNU results in inhibition of MGMT activity in malignant plasma cells and produces meaningful responses in a modest proportion of patients with multiple myeloma. Hematologic toxicity with this regimen is significant and dose-limiting.

Fatal outcome related to carmustine implants in glioblastoma multiforme.

Following the resection of newly diagnosed or recurrent glioblastomas, local implantation of carmustine-impregnated biodegradable wafers (Gliadel) in the resection cavity constitutes an adjuvant therapy that can improve the possibilities of survival. However, some precautions should be taken regarding Gliadel implantation. We report three cases in whom patients with glioblastoma multiforme were implanted with fibrin glue-secured Gliadel after the lateral ventricles had been opened, and who later developed severe hydrocephalus leading to death. Although Gliadel may be an important adjunct to treatment, opening of the ventricles during surgery as part of its application should be considered a contra-indication.

Higher glioblastoma tumour burden reduces efficacy of chemotherapeutic agents: in vitro evidence.

INTRODUCTION: Glioblastoma is the most common primary brain tumour with poor overall survival. Surgical resection followed by radiation and chemotherapy is the mainstay of treatment. The role of aggressive resection in improving overall survival remains contentious, although there is evolving data to suggest this trend. Definitive evidence will necessitate a well-designed randomized prospective trial, although it is not likely that this will be possible or feasible. One possible advantage of aggressive tumour resection is a rapid reduction in oncological burden which may augment the effect of adjuvant chemotherapy. MATERIALS AND METHODS: Three glioblastoma cell lines were seeded in concentrations from 5000-20,000 cells per well onto 96-well plates. The cells were incubated for 24 hours before treatment with varying concentrations of 1,3-Bis(2- chloroethyl)-1-nitrosourea (BCNU) ranging from 25 to 175 microg/mL. After 24 hours of treatment with BCNU, the cells were then examined microscopically and subjected to a cell proliferation assay to determine cytotoxicity effects of BCNU. RESULTS: The drug concentration required to achieve greater than 90% growth inhibition (IC90) was taken as the reference for efficacy of chemotherapy dose. With tumour loading of 5000 cells per well, BCNU concentrations of 75-100 microg/mL resulted in greater than IC90, whereas BCNU concentration of 150-175 microg/mL was required with tumour loading of 20,000 cells per well. A higher concentration of chemotherapeutic agent is therefore required to bring about cell death in the presence of greater tumour burden. CONCLUSION: Higher glioblastoma loading confers chemoresistance to BCNU. This is possibly secondary to complex interactions between tumour cells and neighbouring cells acting via autocrine or paracrine signaling pathways.