Anticancer Research

Published by International Institute of Anticancer Research
Online ISSN: 1791-7530
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Article
The effect of treatment schedule on the antitumor activity of a new platinum derivative, glycolate-0,0'-diammineplatinum (II) (254S) compared with cis- diamminedichloroplatinum (II) (CDDP) was investigated using ascites L1210 leukemia and solid Lewis lung carcinoma. The drugs were given i.p. by three treatments: as a single injection, as three injections at 4 day intervals and as 9 daily continuous injections. 254S produced a marked increase of lifespan in mice by all three treatment schedules (about 100% ILS), although the consecutive treatment of 254S needed more total doses against L1210 leukemia. The antitumor activity of 254S was, however, inferior to that of CDDP. Moreover, 254S did not show certain dependence on treatment schedule, while CDDP was rather dependent on treatment schedule. The single injection (day 1) of CDDP exhibited the most potent antitumor activity. On the other hand, although the single injection of CDDP showed more host toxicity than the other treatment schedules and the consecutive treatment needed more total doses, neither drug showed any definite schedule dependency against Lewis lung carcinoma. Moreover, the tumor growth inhibitory activity of 254S was almost the same as or slightly superior to that of CDDP. Both drugs produced about 70% (days 14-17) and 50% (day 20) tumor weight inhibitions against early and advanced Lewis lung carcinoma, respectively.
 
Article
Cancer therapeutic drugs that inhibit DNA topoisomerase (topo) II by stabilizing the cleavable complex are collectively known as topo II poisons. Phenoxodiol is a synthetic derivative of the plant isoflavone daidzein and is currently undergoing clinical testing as a cancer therapeutic drug. The development of this agent as an antitumor drug was based to a large extent on its low toxicity in normal tissues but potent topo II inhibitory effects in rapidly dividing tumor cells. To evaluate phenoxodiol as a potential inhibitor of topoisomerases, we used the relaxation and nicking assays that can identify topo I inhibitors, and the unknotting and DNA cleavage assays that can identify topo II inhibitors. Phenoxodiol inhibited the catalytic activity of topo II in a dose-dependent manner and it stabilized the topo II-mediated cleavable complex, demonstrating that this agent is a topo II poison. Phenoxodiol's topo II inhibitory effects were comparable to those of other antitumor agents such as VP-16 and were stronger than those of genistein. Phenoxodiol did not inhibit topo I catalytic activity nor did it stabilize the topo I-mediated cleavable complex. These results demonstrate that phenoxodiol is a topo II-specific poison and suggest that this novel agent may find applications in cancer chemotherapy.
 
Article
A mouse leukemia L1210 cell line (Y8), selected for resistance to deoxyadenosine, has a markedly altered phenotypic expression that includes loss of sensitivity to dATP as an allosteric inhibitor of ribonucleotide reductase, increased expression of c-myc, c-fos and WAF1/p21, but decreased expression of p53. In addition, the Y8 cells have a Very strong apoptotic response to a variety of agents under conditions in which the parental wild-type cells do not apoptose. In these studies, we show that flavopiridol (a cdk inhibitor) causes the Y8 cells to undergo apoptosis via a caspase-3 activation process. The apoptotic response to flavopiridol is markedly enhanced by LY294002. Data also show that the apoptotic response of the Y8 cells to roscovitine (a cdk inhibitor) is enhanced by UCN-01 (a PKC inhibitor). These data show that simultaneous blockage of specific pathways leads to increased apoptosis in the Y8 cells with essentially no effects on the parental wild-type L1210 cells.
 
NaB and UCN-01 induce apoptosis in human cervical carcinoma cells. HeLa and CaSki cells were treated for 48 h with increasing concentrations of NaB, or for 24 h with increasing concentrations of UCN-01, as described in the Materials and Methods. A: Following treatment with NaB or UCN-01, mitochondrial membrane depolarization was analysed by flow cytometry using MitoTracker Red dye. B: Sub-G1 DNA levels were assessed following cell incubation with propidium iodide. All results represent the mean±s.d of 3 determinations. They are representative of 3 independent experiments. 
NaB and UCN-01 combinatory treatment induces G1 phase accumulation and enhances apoptosis in HeLa and CaSki cells through induction of cell cycle regulators. A: Cells were treated for 48 h with 6 mM NaB, or for 24 h with 250 nM UCN-01, or with a combination of both drugs. Flow cytometric analyses show cell cycle distribution in the G1, S and G2/M phases (upper panel) and sub-G1 DNA content after staining with PI (lower panel). Data are representative of multiple independent experiments. B: Histograms show the percentage of apoptotic cells after co-treatment with NaB and UCN-01, either with a fixed concentration of 250 nM UCN-01 plus increasing concentrations of NaB (upper panels), or with a fixed concentration of 6 mM NaB plus increasing concentrations of UCN-01 (lower panels). Each bar represents the mean±s.d of 3 determinations. Similar results were obtained from 3 independent experiments. *P<0.05, ***P<0.001. C and D: Cells were cultured for 48 h with 6 mM NaB or for 24 h with 250 nM UCN-01 or 20 nM STS, alone or in combination followed by Western blotting to detect p53, p21 and p73. Blottings using an anti-β-actin antibody served as protein loading controls. NaB and UCN-01 (C) or STS (D) combination increased the expression of the three cell cycle regulators studied. 
NaB/UCN-01 or STS-mediated cell death is associated with caspase activation and PARP cleavage. HeLa and CaSki cells were cultured for 48 h with 6 mM NaB, or for 24 h with 250 nM UCN-01 or 20 nM STS, alone or in combination. A: Flow cytometric analysis of HeLa and CaSki cells showing cleaved caspase-3 immunostaining after the different treatments. The values indicate the percentage of cells with active caspase-3 which significantly increased in co-treated cells compared to cells treated with a single agent. Data are the means±s.d of 3 determinations. B and C: Western blots of cell lysates were performed and were probed with the corresponding antibodies. They confirm procaspase-3 processing, as well as PARP cleavage, which were more important in cells co-treated with NaB and UCN-01 (B), or STS (C) compared with single treatment. Blottings using an anti-β-actin antibody served as protein loading controls. D: Representative images of HeLa cells after exposure to NaB, UCN-01, or both drugs. More apoptotic cells are visualized after co-treatment compared to single treatment. 
Analysis of xenografted tumours. A: Tumours were dissected, fixed in 4% buffered formalin and paraffin-embedded. Five μm sections were treated for histology (a, b), Ki 67 immunohistochemistry (c, d) and HPV DNA in situ hybridization (e, f). CaSki cells were used as positive control for the latter (e). Microphotographs were taken with ×5 (a, c), ×10 (d) and ×20 objectives (b, e, f). B: Schematic representation of the experimental design: CaSki-bearing mice were treated for 5 days with NaB (200 or 800 mg/kg) or with UCN-01 (1.875 or 7.5 mg/kg) intra-peritoneal (i.p.) injection 4 weeks after grafting. Mock mice were treated with vehicle. C: Tumour growth was evaluated by plotting the mean of the relative tumour volume per group (each group consisted of 5 mice) against time after first treatment. D: Relative tumour volume at the end of the experiment, 4 weeks after treatment beginning. 
Article
The effect of combining sodium butyrate (NaB), a histone deacetylase inhibitor, and 7-hydroxy-staurosporine (UCN-01) on cytotoxicity in human cervical carcinoma cells was evaluated. HeLa and CaSki cells were treated using NaB alone or in combination with staurosporine (STS) or its analog UCN-01. Cytotoxicity was determined by flow cytometry and morphological assays. Apoptotic pathways were characterized by Western blotting and immunostaining. CaSki cells were also xenografted into nude mice to assess the in vivo effects of NaB/UCN-01 combination. Treatment with NaB and STS or UCN-01 resulted in enhanced apoptosis of cancer cells. Apoptosis involved mitochondrial pathways and overexpression of p53 and p73. In concordance, co-treatment modulated some p53/p73 downstream targets such as p21, BAX, BCL-2 and BCL-X(L), leading to increased caspase-3 and poly(ADP-ribose) polymerase cleavage. In vivo, NaB/UCN-01 combination exerted a substantial tumour growth suppression effect compared with single treatment. UCN-01 was shown to be a potentiator of NaB therapy for cervical cancer cells.
 
Article
The prognosis for advanced gastric cancer is poor, with surgery as the only treatment for resectable advanced gastric cancer. Therefore, treatment options that might improve the prognosis are needed. To that end, neoadjuvant chemoradiotherapy with S-1 and cisplatin (CDDP) was investigated. The chemotherapy schedule included one cycle repeated after 6 weeks. S-1 was administered orally every day on days 1-21 and CDDP was infused on days 1, 8 and 15. Radiation therapy was started concurrently with chemotherapy and repeated daily on days 1-5, 8-12, 15-19, and 22-26. A total of 10 patients were recruited. The first four patients were entered into level 1 (CDDP, 20 mg/m²). The next six patients were entered into level 0 (15 mg/m²), because of dose-limiting toxicity (delaying the second course of chemotherapy in two patients) that had been observed at level 1. The maximum tolerated dose (MTD) of CDDP was 20 mg/m². Seven patients underwent surgery and all had an R0 (no residual tumor) resection without surgical complications. Neoadjuvant chemoradiotherapy with S-1 and CDDP may cause surgery to be delayed, but shows promise for resectable advanced gastric cancer.
 
Article
7-Hydroxystaurosporine (UCN-01) is a novel antitumor agent as well as a potent inhibitor of a variety of protein kinases with a preference to protein kinase C. Because an intimate interaction exists between PKC signaling and the ER signaling pathways, sex steroid agonists/antagonists might modulate the cytotoxicity of UCN-01. Effects of sex steroid agonists and antagonists on the UCN-01 cytotoxicity were analyzed by MTT assay in MCF-7/WT and MCF-7/ADR, a multiple drug resistance phenotype lacking ER and PR. MCF-7/ADR is 23-fold more resistant to UCN-01 than MCF-7. In MCF-7/WT, danazol and mifepristone enhanced the cytotoxicity of UCN-01, while these agents did not exert any significant effects on it in MCF-7/ADR. These results suggest that danazol or mifepristone might enhance the antitumor activity of UCN-01 in hormone-dependent cancer cells by interacting with PR.
 
Article
The primary focus of this study was to investigate the role of human alpha1-acid glycoprotein (hAGP) on the pharmacokinetics and tissue distribution of the antitumor drug UCN-01 (7-hydroxystaurosporine) in rats, following bolus administration and at steady-state blood concentration. To evaluate plasma pharmacokinetics, the rats received UCN-01 alone, UCN-01 + hAGP (87:1 ratio), or UCN-01 + hAGP (26:1 ratio) i.v. Additional rats were studied after i.m. administration of UCN-01 and i.v. administration of human AGP (87:1 ratio). For tissue distribution, rats received UCN-01 alone, UCN-01 + hAGP (87:1 ratio). One hour after drug administration, blood samples as well as various tissues and organs were collected. Plasma concentrations of UCN-01 as well as tissue accumulation were measured by HPLC using a fluorescence detector. Following i.v. bolus administration, the UCN-01 concentration-time profile declined bi-exponentially. The distribution half-life was 0.2 hours, while the elimination half-life was 6.65 hours. The volume of distribution of the central compartment (Vc) was 1000 ml/kg and the volume of distribution during the elimination phase (Vdb) was 2551 ml/kg. The total body clearance (TBC) was 4.4 ml/min/kg. Co-administration of hAGP with UCN-01 at 1:87 ratio did not affect the elimination half-life of UCN-01 during our sampling period, however the distribution half-life was delayed by approximately 2.7-fold. Furthermore, the Vc, Vd(beta) and TBC were significantly reduced to 395 ml/kg, 735 ml/kg and 1.34 ml/min/kg, respectively. UCN-01 Vd's and TBC were reduced further by increasing human hAGP:UCN-01 ratio to 26:1. Also, hAGP administration did not significantly affect the pharmacokinetic profile of UCN-01 after i.m. administration, which was similar to that measured after i.v. administration. One hour after i.v. bolus administration, UCN-01 was distributed extensively to all tissues with a tissue/plasma ratio ranging from 10-times in the brain to more than 1000-times in the lungs. The presence of hAGP drastically reduced tissue accumulation of UCN-01, although the tissue to plasma ratio remained > 1.0 except for the brain. At steady-state blood concentration following the infusion of UCN-01 over 180 minutes, the ratio of the drug concentration to the concomitant plasma concentration remained > 1.0, even in the presence of hAGP. The data showed that the binding of UCN-01 to hAGP drastically altered its pharmacokinetics and tissue distribution, even at the plasma steady-state concentration.
 
Article
To evaluate the clinical usefulness of 7-hydroxystaurosporine (UCN-01), we compared the antitumor spectrum of UCN-01 with those of conventional antitumor agents against 40 fresh gastric and 40 fresh colorectal cancer specimens using the MTT assay. At a cut-off concentration of 30 micrograms/ml, UCN-01 showed a higher efficacy rate than mitomycin C (MMC), cisplatin, and 5-fluorouracil (5-FU) against both gastric and colorectal cancers. With respect to the gastric cancer specimens, the antitumor spectrum of UCN-01 was independent from those of the other agents, while the patterns of antitumor effects of the conventional agents all correlated significantly with each other. For the colorectal cancer specimens, the pattern of UCN-01-sensitivity did not correlate with the patterns for 5-FU or MMC. In conclusion, UCN-01 may be useful for clinical application against gastric and colorectal cancer due to its different antitumor spectrum from conventionally available agents.
 
Article
The present exploratory phase II study was performed to evaluate the activity and tolerability of adding a second agent (gemcitabine) to the well-tolerated mitoxantrone/prednisone regimen in patients with locally advanced or metastatic prostate cancer no longer responsive to hormonal treatment. Forty-three patients with hormone-refractory prostate cancer (HRPC) were included in the study from May 2000 to April 2004. Their median age was 71 years (range, 56-81) and their median Karnofsky performance status (KPS) was 90 (range, 70-100). The treatment schedule consisted of intravenous (i.v.) mitoxantrone (8 mg/m2 on day 1), i.v. gemcitabine 800 mg/m2 on days 1 and 8, recycled every 21 days and oral prednisone administered at a dose of 10 mg per day. Hormonal treatment with LHRH was continued in all patients. Up to six cycles of treatment were planned in the absence of progressive disease. Sixteen patients had measurable disease (six patients only measurable disease, ten patients bone disease plus measurable disease) and 27 patients had only bone disease. Concerning the PSA levels, a partial response (PR) was observed in 15 patients (38%), stable disease (SD) in 16 patients (41%) and progressive disease (PD) in eight patients (21%). The objective response was evaluable in 16 patients; one patient was not evaluable because he had received only one cycle. Ten patients (63%) had SD and five patients (31%) PD. In the ten evaluable patients with objective SD, depending upon the PSA response, three PR, six SD and one PD were observed. Among the five patients who progressed, three PD and two SD were observed as a PSA response. Pain remission was recorded in 15/41 patients (36%) and the KPS remained stable in most patients. The median overall survival was 15 months (range, 1-41) (95% CI: 10-20 months). The 1-year survival rate was 61%. Hematological toxicity was mild: G 3-4 neutropenia was observed in five (12%) patients. There were no neutropenic, fevers. No significant non-hematological toxicity was observed. The mitoxantrone, gemcitabine and prednisone combination, in accordance with the present regimen, was feasible, had a palliative effect, good tolerance and antitumor activity. Nonetheless, our results do not seem to be superior to those previously described for mitoxantrone plus prednisone.
 
Article
Background/Aim: Patients with human papillomavirus (HPV)-positive tonsillar and base of tongue cancer have a better outcome than those with corresponding HPV-negative tumors (80% vs. 40% 5-year disease free survival with conventional radiotherapy). They should not all need chemoradiotherapy, but before tapering treatment, more markers are needed to predict treatment response. In the present study, human leukocyte antigen (HLA) - HLA-A*02 was analyzed with HPV as a prognostic factor for tonsillar and base of tongue cancer. Pre-treatment biopsies, previously tested for HPV DNA, from 425 patients diagnosed with tonsillar and base of tongue cancer between 2000-2009 at the Karolinska University Hospital were examined for HLA-A*02. HLA-A*02 was present in 144/305 (47.2%) of the HPV-positive and 63/120 (52.8%) of the HPV-negative tumours. Among 383 patients treated with curative intent, absence of HLA-A*02 was correlated with increased disease-free survival in the HPV-positive (p=0.016), but not in the HPV-negative group. Absence of HLA-A*02 correlated with better disease-free survival for patients with HPV-positive tonsillar and base of tongue cancer.
 
Article
Triplet combination chemotherapy has the potential to improve the prognosis of patients with unresectable gastric cancer. We conducted a phase I trial of triplet combination chemotherapy consisting of paclitaxel, cisplatin, and S-1 (PCS) for unresectable gastric cancer. Patients with metastatic or incurable disease were enrolled. S-1 was administered on days 1-14. Paclitaxel and cisplatin were infused on days 1 and 15. The starting doses of paclitaxel and cisplatin were 100 and 20 mg/m(2), respectively. Dose levels of paclitaxel and cisplatin were escalated as follows: 120 and 20 mg/m(2), respectively (level 2); 120 and 30 mg/m(2), respectively (level 3). End-points: Dose-limiting toxicities included grade 3 nausea, vomiting, and general fatigue, and grade 4 febrile neutropenia. The maximum tolerated dose and recommended dose were established at level 3 and level 2, respectively. Although further clinical trials are recommended to more thoroughly evaluate safety and efficacy, PCS appears to be an excellent candidate for a standard treatment strategy for unresectable gastric cancer.
 
Article
Neoadjuvant chemotherapy (NAC) is one of the main strategies for patients with locally advanced breast cancer. In our previous study, biological markers such as estrogen receptor (ER), progesterone receptor (PgR), and HER2 were essential predictors of the effectiveness of NAC to help individualize treatment. This study examined the effect of NAC on the disease-free survival (DFS) of breast cancer patients. Furthermore, the study was expanded by adding Ki-67 as a biological marker, and examined the correlation between Ki-67 and the prognosis. Between September 2005 and September 2007, 43 patients with breast cancer received NAC and surgery. Four cycles of DC (doxorubicin: 60 mg/m(2) and cyclophosphamide: 500 mg/m(2)) were administered intravenously (i.v.) on day 1 every 21 days, followed by 12 cycles of paclitaxel i.v. (80 mg/m(2)) every 7 days, prior to surgery. The primary endpoint was the pathological complete response (pCR) rate and the secondary endpoint was DFS; the pCR rate was estimated for each groups stratified by the presence or absence of different factors (PcR, ER/PgR, and Ki-67). The clinical response (cCR+cPR) rate was 81.0%, and the pCR rate was 25.6%. The pCR rate was 75, 50, 9 and 0% in HER2(+)/ER(-), HER2(+)/ER(+), HER2(-)/ER(-), and HER2(-)/ER(+) patients, respectively. The 4-year DFS rate was estimated at 78% for all patients. The HER2 status was an independent predictor of pathological complete response (pCR). The DFS rate of patients with lower Ki-67 values (<15%) was higher than that of patients with higher Ki-67 values (≥15%). The treatment-related adverse events were manageable: the majority were mild, but five patients experienced grade 3 (neutropenia and sensory neuropathy) adverse events. DC followed by weekly paclitaxel is an active and manageable preoperative regimen for breast cancer patients. HER2 overexpression may be a good predictive marker of pCR, and the Ki-67 value after NAC may be a prognostic factor for DFS.
 
Article
We reported that doxorubicin and cyclophosphamide (DC) followed by weekly paclitaxel is an active and manageable preoperative regimen for breast cancer patients. However, as one of the side effects of paclitaxel, neuropathy was noted in up to 30% of patients. Cyclooxygenase-2 (COX-2) and its derived prostaglandins play a role in stimulating angiogenesis, inhibiting apoptosis, and suppressing the immune response. Some recent studies showed that COX-2 inhibitors, such as meloxicam, have the potential to enhance tumor suppression and reduce the severity of paclitaxel-induced neuropathy. Four cycles of DC (doxorubicin: 60 mg/m(2) and cyclophosphamide: 600 mg/m(2)) administered intravenously (i.v.) on day 1 every 21 days were followed by 12 cycles of paclitaxel i.v. (80 mg/m(2)) every 7 days, prior to surgery. During paclitaxel therapy, breast cancer patients were administered meloxicam (10 mg per day) daily, when experiencing symptoms of grade 2 neuropathy (motor or sensory). The primary endpoint was the pCR rate achieved with the treatment. Forty-three patients received preoperative chemotherapy between April 2004 and March 2007 at six centers. The patient population was identified from a database of the Japan Breast Cancer Research Network. Clinical responses were rated as clinically complete response (cCR) in 9 patients (22%), clinically partial response (cPR) in 25 patients (59%), and clinically stable disease (cSD) in 9 patients (19%). pCR was seen in 25.6%. In addition, we identified 15 patients, who developed grade 2 neuropathy during paclitaxel therapy and subsequently received meloxicam. Meloxicam application had a marked effect within 28 days of initiation. The sensory neuropathy of the patients was reduced gradually, but their motor neuropathy did not improve. Five out of the 15 patients with neuropathy experienced symptom improvement after meloxicam treatment (p<0.05; before versus after 2 months of meloxicam administration). Furthermore, among the 15 patients, who received meloxicam, clinical responses were rated as cCR in 2 patients, cPR in 4 patients, and cSD in 9 patients. The pCR was seen in 4 patients (26.7%). Although meloxicam in combination with DC and weekly paclitaxel chemotherapy did not show promising therapeutic activity, it may provide some relief for neuropathy.
 
Article
Vascular disrupting agents (VDAs) are designed to cause a rapid and selective shutdown of the established tumor vasculature, which leads to secondary ischemic tumor cell death. We examined the efficacy of a novel VDA, MN-029, in the rodent KHT sarcoma model. A significant reduction in the functional vessel number was observed after intraperitoneal injection of MN-029 at a dose of 100 mg/kg. Histological evaluation showed extensive necrosis (approximately 90%) by 24 h. MN-029 treatment to the tumor-bearing mice also resulted in a dose-dependent tumor cell killing. When used in combination with radiation or cisplatin chemotherapy, a 100 mg/kg dose of MN-029 significantly enhanced tumor killing compared to that seen with radiation or cisplatin alone. The results demonstrated that MN-029 could cause rapid vascular shutdown in solid tumors, dose-dependent secondary tumor cell killing, and effective enhancement of the antitumor effects of radiation and cisplatin chemotherapy.
 
Article
The aim of this phase I study was to adjust the dose of cisplatin as adjuvant combination chemotherapy with S-1 in an outpatient setting for gastric cancer. The first course was initiated with S-1 monotherapy on days 1-28. From the second to the sixth course, S-1 was administered on days 1-28 and cisplatin was added on days 1, 15, and 29. The dose level of cisplatin was escalated as follows: 20 mg/m(2) (level 1); 25 mg/m(2) (level 2); 30 mg/m(2) (level 3). Dose-limiting toxicity was a delay factor of the start of the next course due to incomplete recovery. The maximum tolerated and recommended doses were confirmed as level 3 and level 2, respectively. Although further clinical trials are recommended to evaluate efficacy, this combination of S-1 plus cisplatin regimen is expected to become a standard adjuvant treatment for gastric cancer in the outpatient setting. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.
 
Cell-cycle analysis. Cells were either kept as untreated controls or treated with 100 nM PD-0332991 for 48 hours. Cells were analyzed by flow cytometry after propidium iodide staining. The figure depicts the percentage of cells that transitioned from G 0 -G 1 to S and G 2 -M phases of the cell cycle. SW 156, a sensitive cell line, was the only cell line to demonstrate a significant difference between treatment and control for G 0 /G 1 arrest.  
Apoptosis analysis. Cells were either kept as untreated controls (Un) or treated with 100 nM PD-0332991 for 48 hours. Identification of apoptotic subpopulations were completed by labeling phosphatidylserine residues of the cell surface with annexin V-FITC and staining cells with propidium iodide. This staining allows differentiation of apoptotic subpopulations. SW 156, a sensitive cell line, was the only cell line to demonstrate a significant difference between treatment and control in apoptosis, specifically the sub-population of late apoptosis.
Article
PD-0332991 is an inhibitor of cyclin-dependent kinases (CDK) 4 and 6, and was evaluated to determine its anti-proliferative effects in 25 renal cell carcinoma (RCC) cell lines. Half-maximal inhibitory concentrations (IC50) of PD-0332991 were determined with cell line proliferation assays, as were its effects on the cell cycle, apoptosis, and retinoblastoma (RB) phosphorylation. Molecular markers for response prediction, including p16, p15, cyclin D1 (CCND1), cyclin E1 (CCNE1), E2F transcription factor 1 (E2F1), RB, CDK4 and CDK6, were studied using array comparative genomic hybridization (CGH) and gene expression. IC50 values for PD-0332991 ranged from 25.0 nM to 700 nM, and the agent demonstrated G0/G1 cell-cycle arrest, induction of late apoptosis, and blockade of RB phosphorylation. Through genotype and expression data p16, p15 and E2F1 were identified as having significant association between loss and sensitivity to PD-0332991: p16 (p=0.021), p15 (p=0.047), and E2F1 (p=0.041). PD-0332991 has antiproliferative activity in RCC cell lines, and molecular markers predict for sensitivity to this agent.
 
Article
S-1 is an orally administered fluorinated pyrimidine with high activity in metastatic breast carcinoma (MBC) and in chemotherapy-pretreated metastatic breast carcinoma. Forty patients with MBC who did not respond to capecitabine-based chemo-therapy and then received S-1 were identified from our data base of records between 2006 and 2008. The clinico-pathological data and outcomes of these patients were then reviewed. The overall response rate was 27.8%. The median survival was 19.2 months, and the median time to disease progression was 6.2 months. The most common treatment-related adverse events (all grades) were hand-foot syndrome (15%), nausea (15%), vomiting (7.5%), disorder of taste (7.5%), and diarrhea (5%). However, the majority were mild to moderate in intensity, and only one patient experienced grade 3 (according to the National Cancer Institute of Canada Common Toxicity criteria) adverse events. Myelosuppression and alopecia were rare, and there were no reported treatment-related deaths. The results of the current study demonstrate that S-1 is an effective and well-tolerated treatment in patients with capecitabine-resistant MBC. In addition, it is a convenient, orally administered drug, which makes it an attractive agent for use in outpatient treatment.
 
Article
Wilms' tumor gene 1 (WT1) product is a pan-tumor-associated antigen. We previously identified WT1 protein-derived promiscuous helper peptide, WT1332. Therefore, isolation and characterization of the WT1332-specific T-cell receptors (TCRs) are useful to develop broadly applicable TCR gene-based adoptive immunotherapy. A novel HLA-DRB1*04:05-restricted WT1332-specific TCR gene was cloned and transduced into human CD4(+) T-cells by using a lentiviral vector. The WT1332-specific TCR-transduced CD4(+) T-cells showed strong proliferation and Th1-cytokine production in an HLA-DRB1*04:05-restricted, WT1332-specific manner. Furthermore, the WT1332-specific TCR-transduced CD4(+) T-cells could lyse HLA-DRB1*04:05-positive, WT1-expressing leukemia cells in vitro. The novel TCR gene cloned here should be a promising tool to develop adoptive immunotherapy of WT1332-specific TCR-transduced CD4(+) T-cells for the treatment of WT1-expressing cancer, such as leukemia. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.
 
Article
The aims of this multiple-institution phase II study were to evaluate the efficacy and tolerability of docetaxel, cisplatin and 5-fluorouracil (DCF) for the therapy of patients with metastatic squamous cell carcinoma of the esophagus (SCCE). Eligible patients included those with previously untreated SCCE, score of ECOG 0-2 and adequate organ function. Patients received 60 mg/m(2) docetaxel and 70 mg/m(2) cisplatin on day 1, and 600 mg/m(2) 5-fluorouracil on days 1-5 every four weeks. Twenty-nine (22 male, 7 female) patients with metastatic SCCE (M1a: 20, M1b: 9) were enrolled. Three cases achieved complete response and seven a partial response. In addition to these patients, three patients achieved good response and underwent surgical resection, giving an overall response rate of 34.5% (95% Confidene Interval=17.9-54.3) in confirmed cases and 44.8% (95% CI=26.4-64.3) in unconfirmed cases. Grade 3 or 4 hematological toxicities were as follows: leukopenia in 15 patients (52%), neutropenia in 22 patients (76%) and febrile neutropenia in 6 patients (21%), while grade 3 or 4 non-hematological toxicities were relatively rare. This DCF regimen was well tolerated; the results of this study provide information on the potential of DCF for treatment of patients with metastatic SCCE.
 
Article
Cisplatin resistance is an important issue in lung cancer. We aimed at investigating if the Hedgehog pathway inhibitor GDC-0449 is effective in cisplatin-resistant cells and if it alters intracellular Ca(2+)-homeostasis. The cytoplasmatic ([Ca(2+)](cyto)) and endoplasmatic ([Ca(2+)])(ER) Ca(2+) concentration of HCC (adeno carcinoma of the lung) and H1339 (small cell lung carcinoma) cells were measured with the calcium indicator dye Fura-2 AM. The expression of the inositol-1,4,5-trisphosphate receptor (IP(3)R) and sarco/endoplasmic reticulum Ca(2+)-ATPase (SERCA) were analyzed using western blot analysis. GDC-0449 inhibited cell growth in cisplatin-naïve and -resistant cells. In both cell types, GDC-0449 increased [Ca(2+)](cyto) and reduced endoplasmatic [Ca(2+)](ER). Cisplatin failed to considerably alter Ca(2+) homeostasis in resistant cells. The effects of GDC-0449 on intracellular Ca(2+) homeostasis were not mediated by an altered expression of IP(3)R or SERCA. GDC-0449 alters intracellular Ca(2+) homeostasis and inhibits cell growth in cisplatin-resistant lung cancer cells.
 
Article
Cisplatin is a widely used agent for treatment of solid tumors, but its clinical utility is limited by toxicity. Preclinical studies have shown less acute toxicity when STEALTH liposomal cisplatin (SPI-077) is used, with antitumor effects equivalent to those of intravenously administered free cisplatin. We previously reported that systemic treatment with low-dose tumor necrosis factor-alpha (TNF) augments the activity of STEALTH liposomal doxorubicin (Doxil). In this study, we examined the effect of repeated systemic applications of low-dose TNF on the antitumor activity of SPI-077 in rats with soft-tissue sarcoma or osteosarcoma. Addition of TNF to SPI-077 treatment showed an improved tumor growth delay of the soft-tissue sarcoma. The combined SPI-077/TNF treatment resulted in a more prolonged antitumor activity, whereas free cisplatin showed a better tumor response, however with a rapid outgrowth a few days after the end of therapy. In the osteosarcoma, free cisplatin did not have an antitumor effect, but addition of TNF caused a clear tumor growth delay. SPI-077 alone resulted in a tumor growth delay, but combination with TNF had no additive effect. SPI-077 yielded less systemic toxicity than cisplatin. Depending on the type of tumor, the addition of TNF to SPI-077 results in a better tumor growth delay with a prolonged antitumor effect and, in combination with the reduced toxicity of SPI-077, this combination may be preferable to cisplatin.
 
Article
In in vitro experiments, the localized lipophilic cation, MKT-077, demonstrated time- and concentration-dependent antitumor activity. In the present experiment, the in vivo antitumor activity of MKT-077 was evaluated using human tumor xenografts serially transplanted into nude mice. The antitumor efficacy of MKT-077 against five xenografts inoculated subcutaneously into nude mice was studied. Treatment with MKT-077 was initiated when the tumors started exponential growth. The antitumor activity of MKT-077 was assessed by a) the lowest value of the relative mean tumor weight of the treated:control tumors (T/CRW), where relative weight (RW) represented the change from baseline weight; and by b) the unadjusted weight of treated tumors at the end of the experiment. When MKT-077 was administered continuously using the Osmotic Micropump, antitumor activity was positively correlated with exposure time and drug concentration from 7.5 to 40 mg/kg/day. The maximum tolerated dose was 20 mg/kg/day for 7 days. Co-4 (human colon cancer), St-4 (human gastric cancer), and CRL1420 (human pancreatic cancer) were evaluated as sensitive to MKT-077 in tested 5 strains. The antitumor activity of MKT-077 was confirmed using human tumor xenografts in nude mice.
 
Article
To assess the efficacy of multiple treatment of phosphatidylinositol-3-kinase (PI3K) inhibitor on autochthonous tumours in phosphatase and tensin homologue (Pten)-deficient genetically engineered mouse cancer models using a longitudinal magnetic resonance imaging (MRI) protocol. Using 3D MRI, B-cell follicular lymphoma growth was quantified in a Pten(+/-)Lkb1(+/hypo) mouse line, before, during and after repeated treatments with a PI3K inhibitor GDC-0941 (75 mg/kg). Mean pre-treatment linear tumour growth rate was 16.5±12.8 mm(3)/week. Repeated 28-day GDC-0941 administration, with 21 days 'off-treatment', induced average tumour regression of 41±7%. Upon cessation of the second treatment (which was not permanently cytocidal), tumours re-grew with an average linear growth rate of 40.1±15.5 mm(3)/week. There was no evidence of chemoresistance. This protocol can accommodate complex dosing schedules, as well as combine different cancer therapies. It reduces biological variability problems and resulted in a 10-fold reduction in mouse numbers compared with terminal assessment methods. It is ideal for preclinical efficacy studies and for phenotyping molecularly characterized mouse models when investigating gene function.
 
Article
Since the advent of microvascular free tissue transfer, adjunctive surgical techniques have become sought as a means to improving overall surgical time and outcomes in oncological surgery. Anastomotic devices have provided one such avenue, with staples and coupling devices suggested as an alternative to traditional suturing. We describe our experience with two such devices, automatic staples and the anastomotic ring coupler, in 1,000 cases of free tissue transfer. In 1,400 anastomoses, there was a significant reduction in anastomotic time from suturing (22 minutes) with the use of staples (15 minutes) or the ring coupler (4 minutes), p<0.01. This was without any increase in complication rates. These findings support the use of these devices and suggest an increasing role for modern devices in microvascular surgery.
 
Article
Minimizing normal tissue toxicity can enhance the therapeutic gain of thermochemotherapy. For this purpose, we investigated the optimal duration of whole body hyperthermia (WBH) (41.5 degrees C) when administered simultaneously with carboplatin (CBDCA). Using a transplantable fibrosarcoma in Fischer 344 rats, we measured tumor growth delay (TGD) as well as normal tissue toxicities (body weight loss, thrombocytopenia) induced by various durations of WBH (0.5, 1.0, 1.5, 2.0 or 2.5 hours) when combined with CBDCA (30 mg/kg, i.v.). When combined with CBDCA, 1.0 hour WBH increased the TGD compared to 0.5 hour of WBH, but with WBH durations greater than 1.0 hour, the TGD did not further significantly increase. Measuring CBDCA-induced myelosuppression, the platelet count on day 6 post-treatment decreased from a control mean of 6.8 x 10(8)/ml to 1.8 x 10(8)/ml after 2.5 hour WBH exposure in a duration-dependent manner (p < 0.001). To estimate the specific therapeutic efficacy (STE), we calculated a ratio of TGD to myelosuppression (thrombocytopenia). Compared to other WBH exposure times, 1.0 hour duration of WBH combined with CBDCA produced the highest STE (2.8) and over 1.5 hour duration of WBH did not result in any additional increase in STE. We conclude that 1.0 hour WBH exposure is optimal when combined with CBDCA in order to maximize the therapeutic gain.
 
Article
The antitumor mechanism of (-)-(R)-2- aminomethylpyrrolidine (1.1-cyclobutanedicarboxylato)platinum(II) (DWA2114R) was examined using cultured murine L1210 leukemia cells by estimating its effects on parameters such as proliferation, macromolecular synthesis, morphology and cell cycle progression. Each parameter was estimated in cells concomitantly exposed to the drug for 24-48 hr. More than 0.1 microM of DWA2114R markedly inhibited cell proliferation as well as DNA synthesis, and it decreased in mitotic index in a concentration-dependent manner. One microM of DWA2114R decreased DNA synthesis by 80% in the cells treated for 24 hr, while the inhibition of RNA synthesis was less than 40%. A significant inhibition of protein synthesis was caused only by treatment with a high concentration (100 microM) of the drug. Under complete cytostatic conditions (10 microM of DWA2114R), cell volume markedly increased and about 40% of the total cells were polynucleate. In addition, flow cytometrical analysis revealed that most of these cells were accumulated in the G2/M phase of the cell cycle, and a new peak located in the G2/M phase of tetraploid cells emerged. On the other hand, the cells treated with 100 microM of the drug did not increase in volume and their progress in the cell cycle was almost completely blocked.
 
Article
1,1-Diallylhydrazine was administered as a 0.03125% solution in drinking water for life to Swiss mice, from 6 weeks of age. Compared to untreated controls, in treated animals the lung tumor incidence rose from 25 to 76% in females and from 26 to 76% in males, whereas the incidence of forestomach tumors increased from 4 to 14% in females and from 0 to 34% in males. Histopathologically, the tumors were classified as adenomas and adenocarcinomas of the lungs and squamous cell papillomas and carcinomas of the forestomach. This work is part of our structure activity inquiry and demonstrates the relative carcinogenic potency of another disubstituted allylhydrazine.
 
Article
Z-1,1-dichloro-2,3-diphenylcyclopropane (a.k.a. Analog II, AII) is known as a potential anti-breast cancer agent and has previously been studied as an antiestrogen (AE). We hypothesized that its activity is independent of estrogen receptor (ER) status. AII and its known and potential metabolites were synthesized and characterized by NMR and MS. ER positive/estrogen and AE sensitive MCF-7, ER positive/estrogen and AE resistant MCF-7/LY2, and ER negative/estrogen and AE resistant MDA-MB231 cells were used in metabolism, cytostasis, cytotoxicity, and serum binding/wash out assays. Bacterial mutation assays were performed with Salmonella typhimurim strain TM677. AII underwent slow solvolysis in culture medium to Z-2-chloro-1,3-diphenyl-2-propen-1-ol and its oxidized form Z-alpha-chlorochalcone (ZCC). ZCC was the major metabolite of AII in all three cell lines. Cytostasis and clonogenic assays showed AII to be cytostatic to each of the lines, and was more potent against MCF-7 and MCF-7/LY2 than MDA-MB231 cells. ZCC was cytotoxic, with IC50 values of 89, 0.5, and 170 nM in MCF-7, MCF-7/LY2, and MDA-MB231 cells, respectively. Cytotoxicity from ZCC was delayed compared to loss in cell viability, suggesting a non-necrotic mechanism. Serum protected against loss of cell viability caused by AII, but had no effect on the action of ZCC. The effects of ZCC could be partially reversed by washing the drug out of cells. The effects of AII persisted after wash out. AII was also shown to be nonmutagenic in forward Salmonella mutation assays both with and without metabolic activation. In conclusion, AII, a chemical with weak antiestrogenicity, anti-breast cancer activity, and low toxicity in whole animals, shows growth inhibitory properties against both ER positive and negative human breast cancer cells in culture. Its direct action appears to be cytostatic and longlived. AII is converted by the cells to a less-retained and -protein bound metabolite, ZCC, that is more cytotoxic. Neither AII nor ZCC appear to have mutagenic activity. Both AII and ZCC thus appear to have potential for use against estrogen-dependent and -independent human breast cancers.
 
Article
Z-1,1-dichloro-2,3-diphenylcyclopropane (a.k.a. Analog II, AII) is a known anti-breast cancer agent with apparent antiestrogenic effects and remarkably low toxicity in rodents. We have recently shown that AII and its major metabolite Z-alpha-chlorochalcone (ZCC) inhibit proliferation of both estrogen-responsive and -nonresponsive human breast cancer cells, suggesting its mechanism is not mediated by the type I estrogen receptor (ER). The present studies were performed to begin to define the molecular targets of AII and ZCC. Based on the compounds' structures and actions, we hypothesized that their effects could be due to interaction at type II estrogen binding sites (EBSII) and/or cellular microtubules. The affinities of AII ZCC and the positive control diethylstilbestrol (DES) for the ER (in MCF-7 and MCF-7/LY2 cells) and EBSII (in MCF-7, MCF 7/LY2, and MDA-MB231 cells) were determined with a whole cell assay for displacement of [3H]estradiol. The kinetics of their effects on cellular microtubules and cell cycle distribution of human breast cancer cells were measured by indirect immunofluorescence and flow cytometry. Their abilities to inhibit assembly of isolated tubulin in vitro were determined. AII, ZCC, and DES had similar affinities for the EBSII in the three cell lines. Neither AII nor ZCC displaced [3H]estradiol from the ER in MCF-7 cells, whereas DES did. The microtubule network of MDA-MB231 cells exposed to 100 microM AII or 10 microM ZCC began to disassemble within 1 hour of treatment and was completely diffuse after 6 hour of exposure to either drug. AII inhibited in vitro assembly of tubulin, with an IC50 of 6.7 +/- 0.9 microM, while ZCC was inactive below 40 microM. Both drugs caused accumulation of the cells in the G2/M phase of the cell cycle. The evidence suggests that the antitumor action of AII is mediated, at least in part, through the EBSII and/or perturbation of tubulin-microtubule dynamics. AII thus represents a new lead compound for design and discovery of novel antitumor agents directed against the EBSII and/or microtubules.
 
Article
The in vitro antimetabolic effect of a newly synthesized platinum antineoplastic agent, DWA2114R, on 72 fresh human tumors was compared with that of cis-platinum (CDDP). DWA2114R is reported to have a lower nephrotoxicity than CDDP, but is as strong an inhibitor of DNA synthesis as CDDP. In vitro IC50 was assessed in 10 tumors; the IC50 of DWA2114R ranged between 9.2 and 107 microM (mean +/- S.D., 45.9 +/- 36.6) and that of CDDP ranged between 0.9 and 40 microM (18.3 +/- 15.1). DWA2114R had an antitumor spectrum similar to that of CDDP. Primary esophageal and pancreatic cancers were sensitive to DWA2114R, and cells separated from malignant effusion or metastatic lymph nodes were more sensitive than those from primary lesions. Nude mice were transplanted with 3 kinds of human tumor xenografts (esophageal, pancreatic and bile duct cancer lines), and were then treated with CDDP or DWA2114R at 4 times the clinical doses. CDDP significantly inhibited the growth of all the 3 lines, and DWA2114R inhibited the growth of 2 of the lines. The effect of DWA2114R on body weight was smaller than that of CDDP. These results suggest that DWA2114R may be less potent than CDDP but may be useful as a new platinum antineoplastic agent with lower grade of side effects than CDDP.
 
Article
(-)-(R)-2-Aminomethylpyrrolidine(1,1-cyclobutanedicarboxylato++ +)platinum(II) monohydrate (DWA2114R), cis-diammine(1,1-cyclobutanedicarboxylato)platinum(II) (CBDCA) and cis-diamminedichloroplatinum(II) (CDDP) were compared for their antitumor effects and nephrotoxicity-inducing activities at the same dosage (1/8, 1/4, 1/3, 1/2, 2/3 or 3/4 of the LD10 or LD10) on the basis of their intravenous lethal doses in mice. DWA2114R was effective against murine tumor lines, Colon 26 and Colon 38 carcinomas, M5076 ovarian sarcoma and P388 L1210 leukemias, implanted subcutaneously (s.c.). Triple injection every other day of DWA2114R was more effective than a single injection at each sublethal dose. The antitumor effects of DWA2114R against these tumors were more effective than or were similar to those of CBDCA and CDDP. The antitumor effect against CDDP-resistant L1210 leukemia implanted s.c. was only observed in the treatment of DWA2114R, but not in CBDCA and CDDP. No excellent antitumor effects of three platinum complexes were observed against Lewis lung carcinoma and B16 melanoma implanted s.c. even at triple injection every other day, and no effect was obtained against Meth-A fibrosarcoma under similar conditions. While the treatment of CDDP showed marked increases in levels of blood urea nitrogen and of urinary protein and sugar at effective doses in the antitumor evaluations, the treatment of DWA2114R as well as CBDCA showed no increase in these parameters. These results indicate that DWA2114R represents a desirable second generation antitumor platinum complex.
 
Article
FK 973, a novel substituted dihydrobenzoxazine structurally similar to mitomycin C, is a derivative of the product isolated from Streptomyces sandaensis. In vitro and in rodents, it is a potent antitumor agent. During Phase I clinical trials, we evaluated the pharmacologic properties of FK 973 in eight adenocarcinoma patients after a 30-min i.v. infusion of doses ranging from 7 to 45mg/m2. An established enzyme immunoassay that measures the stable deacetylated active metabolite FR66980 showed that the plasma drug levels declined biphasically with a terminal half life (t1/2 beta) of 4.7 +/- 1.6hr (mean +/- S.D.) The total clearance rate was 438 +/- 113ml/(min/m2). Both the maximum plasma drug concentrations (Cmax) and the area under the concentration-versus-time curve (AUC) were dose related. In addition to nausea and vomiting, alopecia, and myelosuppression, three patients experienced a delayed vascular-leak syndrome. The 3 patients had received doses among the highest studied, and the toxicity appeared to be dose related and cumulative. The evidence suggests that higher doses generated higher Cmax and AUC values, which may be correlated with toxic effects.
 
i: R 4 CH 2 COOH (ref. 14), ii: LiAlH 4 /THF (ref. 15), iii: R 3-substituted oxirane/EtOH (65-92%). 
i: R 1 NCS/toluene (85-91%), ii: R 3-substituted oxirane/EtOH (78%) 
Article
Cancer treatment often fails due to multidrug resistance (MDR) of the tumor cells. One of the major causes is overexpression of P-glycoprotein (P-gp). By N-substitution reactions of diamine, amino acid and amino alcohol derivatives with 1-substituted tetrahydroisoquinoline skeleton, structurally diverse 1,2-disubstituted 1,2,3,4-tetrahydroisoquinolines were synthesized. The compounds were assayed as P-gp inhibitors using a standard functional assay with rhodamine (6G) on MCF-7/Adr cells. Cytotoxicity was investigated on HeLa cells using an antiproliferative assay. Five of the 24 compounds showed greater P-gp inhibition than the control compound verapamil with AC50 values (concentration of the compound eliciting 50% of the maximal rhodamine 6G accumulation) significantly lower than that of verapamil. Novel compounds were synthesized that showed MDR-reversal effect. One of them, (1'R*,2R*)-2-[2'-[2''-hydroxy-3''-(alpha-naphthyloxy)propyl]-6',7'-dimethoxy-1',2',3',4'-tetrahydro-1'-isoquinolyl]propan-1-ol hydrochloride, showed two times higher efficacy than verapamil at 10 times lower concentrations. The outcome makes this molecule an attractive subject for further investigation and development.
 
Article
Chlorophyllin (CHL) was tested for its effect on 1,2-dimethylhydrazine (DMH)-induced nuclear aberrations in rat colonic epithelium. Rats were given water containing CHL (1.5 mM) and food ad libitum. After 5 wks on this dietary regimen, the rats were given a single intraperitoneal injection of DMH (20 mg DMH base/kg body weight). The rats were sacrificed 18 hours later and their colons were removed. Ten consecutive crypts from the proximal and distal portion of each specimen were scored for nuclear aberrations and the karyorrhectic index (KI) was determined. Rats receiving CHL + DMH had significantly fewer nuclear aberrations (lower KI) in the colonic epithelium than rats given DMH alone. This implies that CHL, a known antimutagen, may have anticarcinogenic properties as well.
 
Synthesis of compounds 17-23. 
Synthesis of compound 30. 
Article
We report the synthesis of new guanylhydrazones from imidazo[2,1-b]thiazoles and of a bis-guanylhydrazone from diimidazo[1,2-a:1,2-c]pyrimidine. The compounds were tested as potential antitumor agents at the National Cancer Institute. Two derivatives are now under review by BEC: one of these was also subjected to a test for positive inotropic activity in view of a possible coanthracyclinic activity. Tyrosine kinase receptors may be involved as molecular targets in the mechanism of action of the guanylhydrazones described.
 
Morphological features of classic elevated ACF and flat lesions in the colon of DMH-treated F344 rats as observed by surface examination and histological examination under the light microscope. A) Classic elevated ACF at week 31; the same lesions were examined by scanning electron microscopy (Figure 2 A). B) Flat lesion at week 31; the same lesion is examined by scanning electron microscopy (Figure 2 B). C) Classic elevated ACF at week 16. D) Flat ACF at week 16. E) Cross-section of the lesion in C. F) Cross-section of the lesion in D. Magnification: A and B, 50x; C-F 100x. 
Scanning electron microscopy of the colonic mucosa of F344 rats, after DMH treatment. A) Classic elevated ACF, recognised as distinct structures with epithelial cells of similar ultrastructure as the surrounding mucosa (see also Figure 1A). This is illustrated in C) by enlarging the ACF indicated by the arrow C. Normal epithelium with goblet cells are illustrated in E), by enlarging the area indicated by the arrow E. B) A flat lesion recognised as an area with small crypt openings and loss of goblet cells (see also Figure 1B). D) Illustrates the transition between normal epithelium with goblet cells (left side of image) and the flat lesion demonstrating dysplastic epithelium with loss of goblet cells (middle and right side of the image). The areas indicated by the arrows F and G are enlarged to show the normal epithelium F) in contrast to the dysplastic epithelium of the flat lesion G). Magnifications: 50x, 200x and 1125x. 
Scannning electron microscopy of a large tumour in the colon of a F344 rat after DMH treatment. A) Gross morphology of the tumour. The arrows B and C indicate areas enlarged to show small crypt openings B) and dysplastic epithelial cells C), in contrast to the normal surrounding mucosa D). Magnifications: 20x, 150x and 900x. 
Article
The surface morphology of late colonic lesions in F344 rats treated with 1,2-dimethylhydrazine was studied by scanning electron microscopy. At week 31 after carcinogen treatment, the surface epithelial characteristics of different types of lesions observed in the colonic mucosa were compared, namely classic elevated aberrant crypt foci (ACF), flat lesion and gross tumour. Classic elevated ACF were easily observed as structures with enlarged crypts elevated from the background mucosa. When the various ACF were compared, or when the ACF were compared with the background mucosa, no ultrastructural differences, or differences in the density of goblet cells were found. The flat lesion showed an epithelium without goblet cells and crypts with small openings harbouring a large number of loose, undefined, dysplastic epithelial cells. These changes appeared to be linked to the malignant development since they were also characteristic of the examined tumour.
 
Article
We showed the possibility of significant decreasing of the frequency of chemically induced colon cancer in rats by vaccination with polyclonal rabbit IgG generated against purified tumor-associated antigens (TAA). TAA were isolated from benign rat colon tumors by the method developed in our laboratory (Zusman et al 1994) using affinity chromatography columns with gel fiberglass membranes (R. Zusman, 1992) containing anti-tumor IgG. The IgG was isolated from rabbits following their vaccination with TAA. Sprague Dawley rats were vaccinated with anti-TAA IgG (100 micrograms/rat) suspended in Freunds adjuvant by weekly subcutaneous injections for 5 weeks. The induction of colon cancer was caused by weekly injections with 1,2-dimethylhydrazine (DMH) (20 mg/kg) for 7 weeks and was started one week after the end of the vaccination. The results of experiments were evaluated 6 months after the start of cancer induction. IgG protected against the carcinogenic effects of DMH. The number of tumor-bearing rats decreased to 64% as compared with 90% in the control group. In vaccinated rats, the incidence of tumors was almost 3 times less than of control, i.e. 3.6 and 9.3, respectively. The number of malignant tumors was also significantly smaller in vaccinated rats than in controls, being 24% and 58%, respectively. Metastases were found only in controls, 4 of 30 rats. The results of our experiments have shown that anti-TAA IgG not only has anti-tumor effects but also prevents the malignization of benign tumors. As one of the main components of TAA which was isolated from colon cancer rats was soluble p53 antigen (Zusman et al 1994), we suggest that the vaccine which has been generated in our experiments may be regarded as acting mainly against p53 antigen, and its antitumor effects should also be considered as effects of p53 antibodies. The further studies will be performed to clarify this.
 
Article
PHIC [NSC-350602; (1,2-diaminocyclohexane) (isocitrato) platinum (II)] is a new highly water-soluble platinum derivative. Preclinical studies showed antitumor activity on a wide range of tumors, no cross-resistance with cisplatin and little or no nephrotoxicity in mice and baboons. A phase I clinical trial was then initiated at doses of 300 mg/m2 infused intravenously over one hour without induced diuresis or hydration. Dosages were escalated up to 1500 mg/m2. A total of 29 patients received 52 courses of treatment. The most important side-effect is thrombocytopenia which is rapidly reversible. Nausea and/or vomiting were mild or moderate with onset 1 hr after the end of the infusion and seldom persisted beyond 24 hrs. Measurements of biological parameters did not reveal significant evidence of nephrotoxicity except in one patient who developed urinary tract infection and for whom hemodialysis became necessary. No change in the audiogram could be demonstrated. Peripheral neuropathy was documented in one patient, and in two other patients to whom morphine was given confusional episodes were observed. Although no antitumor effect was observed, there was apparent stabilization of disease. Pharmacokinetic parameters were calculated in twelve out of these 29 patients. Based upon total platinum plasma concentrations, the elimination half-life is 58.3 hrs and the plasma clearance is 21.2 ml/min with an apparent volume of distribution of 7.6 liters. However, considering both plasma concentrations and urinary excretion, we could estimate the half-life of free filterable species (60 min), the plasma clearance (125 ml/min) and the renal clearance (86 ml/min). Mean urinary excretion is 64.4% of the dose after 6 days and 53.1% at 24 hrs. Other administration protocols are suggested, based upon these pharmacokinetic parameters.
 
Article
Iron salts supplemented in a basal ratio, were fed to young Sprague-Dawley male rats for prolonged periods and the frequency of colonic adenocarcinomas induced by repeated s.c. injection of 1,2-dimethylhydrazine (DMH) at unit base doses of 9.0 mg/kg, ascertained and compared with the respective controls. In a series employing ferric ammonium citrate (0.46% Fe), in addition to ferrous sulfate (0.11% Fe) and ferric ammonium sulfate (0.12% Fe), DMH injection was started on day 15 and the animals necropsied 22 weeks after the last of 23 doses. The general condition was more involved with the 0.46% Fe diet and the total colonic lesion numbers were in the control range. However, the ferric ammonium sulfate-fed group showed a significant increase in tumors in the distal colon portion. In the second experiment, 15% guar gum as such and in admixture with ferric ammonium sulfate (0.12%) were compared with the respective controls, the first of 20 weekly dosages of DMH being administered on day 28 of the feeding. At 32 weeks following injection 1, the overall lesion differences were not remarkable, but the guar gum ratios engendered decreases in the distal colon tumor frequencies. In general, lesion incidence was extensive, involving 80-100% of the animals per group of the 2 series. Adenocarcinomas occurred in the small intestine and were more prominent in the control and 15% guar gum dietary groups but fewer with the ferric ammonium sulfate supplement.
 
Article
A series of derivatives belonging to a new class of compounds (R4-todit) were highly cytotoxic to a panel of leukaemia- and solid tumour-derived cell lines (IC50 = 0.06-20 microM). The most potent compound was the butyl4 derivative (IC50 = 0.06-5.1 microM); T leukaemia and melanoma cells were the most susceptible cells to this inhibitor (IC50 0.06 microM and 0.1 microM, respectively). The effect of butyl4-todit was irreversible, and led to progressive cell death. The compound showed a comparable potency against exponentially growing and stationary phase cells, and against cell lines expressing the MDR phenotype. The cytotoxicity of butyl4-todit in human normal PBL was up to 20 fold lower than that shown against T leukaemia cells. When tested for antiangiogenic activity in vivo, 1.5 mg/Kg butyl4-todit resulted in over 70% inhibition of the angiogenesis process induced in mice by Kaposi's sarcoma cell secreted products.
 
Article
The activity and subcellular distribution of Protein Kinase C (PKC) was determined in the colons of Sprague-Dawley rats that were fed either a low fat rat chow or rat chow supplemented with 17% corn oil (40% ingested calories as fat). Rats given the high fat diets were either given no carcinogen or treated prior to or subsequent to the initiation of the test diets with 1,2 dimethylhydrazine (DMH). Rats were sacrificed and PKC activity determined in the soluble and particulate fractions of the colonic tissue 13 weeks after the initiation of the diets or DMH treatment, which was before tumor induction. In addition several rats were maintained on their diets until colon tumor formation occurred and PKC activity determined in the colonic tumor and compared to age matched control colonic tissue. In the absence of DMH, fish oil and corn oil equally augmented PKC activity and decreased the ratio of soluble/particulate PKC. With DMH treatment, corn oil augmented PKC as above, but fish oil supplementation resulted in a pattern of PKC activity and distribution more typical of a low fat diet, particularly when fish oil supplementation preceded DMH treatment. PKC activity in DMH induced colonic carcinomas was markedly depressed regardless of the fat source in the diet, when compared to colonic tissue from a non-DMH treated age matched low fat control.
 
Article
Thymidylate synthase and thymidine kinase are key enzymes involved in the de novo and salvage pathways for pyrimidine nucleotide synthesis, respectively. Thymidylate synthase is inhibited by 5-fluorodeoxyuridine monophosphate, forming an inactive ternary complex with intracellular folate. We investigated the effects of 1-(2-tetrahydrofuryl)-5-FU plus uracil (UFT) with or without leucovorin on 1,2-dimethylhydrazine-induced rat colorectal carcinomas. Thirty-week administration of UFT with or without leucovorin markedly suppressed both colorectal carcinogenesis and tumor growth, resulted in the increase of thymidylate synthase inhibition and the decrease of thymidine kinase activity in the tumor cells. These results indicate that the combination of UFT with leucovorin could be useful in the development of pre- and post-operative adjuvant chemotherapy programs.
 
Effect of PSC on the size and location of colorectal tumors. Each square dot represents a tumor with respect to its size (largest diameter in mm) and location (distance from the cecum in centimeters). The colon length was arbitrarily divided at 9 cm ( ~ mid point) to categorize tumor location as proximal (at the cecum end) or distal. The horizontal dotted line at 6.5 mm represents the median tumor diameter in the BD group. All grossly visible colorectal tumors were counted, measured and are represented in the figure. 
Effect of PSC exposure on colorectal tumor (A) multiplicity and (B) burden in rats. Each dot represents a rat. Plot (A) shows the total number of tumors per rat, while (B) depicts the total tallied diameter of lesions per rat. The horizontal bar displays the median for each group. p<0.01 denotes statistical significance by unpaired two-tailed t-test analyses. The results indicate that PSC significantly inhibited colorectal tumor multiplicity by 53% and tumor burden by 74%. 
Article
The expression of P-glycoprotein (Pgp) is intimately associated with cancer development. In order to explore the therapeutic value of Pgp as a target for chemotherapy, we studied the effect of PSC 833 (PSC), a potent inhibitor of Pgp, on 1,2-dimethylhydrazine (1,2-DMH)-initiated colorectal carcinogenesis in rats. Male Fischer 344 rats, initiated with 1,2-DMH coupled with partial hepatectomy, were exposed to dietary 1% orotic acid for 22 weeks. They were then fed either the AIN93G basal diet (BD) or BD containing PSC (a daily dose of 15 mg/kg body weight) for 35 weeks. PSC significantly inhibited colorectal tumor multiplicity by 53% and tumor burden by 74%. PSC-mediated inhibition was evident in tumors as small as 2 mm in diameter and remained effective throughout the course of tumor growth. Histological assessment showed that PSC significantly inhibited tumor progression to colorectal adenocarcinoma by 63%. Collectively, this study indicates that PSC inhibited experimental colorectal carcinogenesis initiated with 1,2-DMH in rats.
 
Article
The 1-acyl- and 1,2-diacyl-4,4-diethyl-3,5-pyrazolinediones proved to be cytotoxic against the growth of a number of cell lines, including murine and human leukemias. HeLa suspended carcinoma, colon adencarcinoma SW480, KB nasopharynx and glioma tumors. Selected compounds were also active in the human lung bronchogenic MB-9812, and osteosarcoma TE418 screens. These derivatives were active in vivo in the Ehrlich ascites carcinoma screen in CF-1 mice at 8 mg/kg/day I.P. The mode of action in Tmol3 leukemia cells showed that the compounds reduced de novo synthesis of purines and pyrimidines and inhibited dihydrofolate reductase and ribonucleoside reductase activities. The DNA molecule was not a target although limited DNA strand scission may be possible.
 
Article
Thymidylate synthase, which is a key enzyme involved in the de novo pathway for pyrimidine nucleotide synthesis, is inhibited by 5-fluorodeoxyuridine monophosphate, forming an inactive ternary complex with intracellular folate. We investigated the effect of a 5-fluorouracil derivative (UFT) with or without low dose leucovorin on the number of 5-fluorodeoxyuridine monophosphate binding sites, thymidine kinase activity and intracellular folate concentration in 1,2-dimethylhydrazine-induced rat colorectal carcinomas. A 10-day administration of UFT with or without leucovorin enhanced the thymidine kinase activity and the number of 5-fluorodeoxyuridine monophosphate binding sites, with an increase of thymidylate synthase mRNA expression. Thymidylate synthase inhibition was slightly increased as the intracellular folate concentration increased. These results indicate that thymidylate synthase inhibition increases when the intracellular folate is exogenously supplemented and maintained at an adequate concentration.
 
Article
The effect of the oral administration of 10 compounds on 1,2-dimethylhydrazine (DMH) carcinogenesis was investigated in 180 male Wistar rats and 510 male BD6 rats. DMH, administered s.c. once per week for 20 consecutive weeks (20 mg/kg body wt/dose), produced intestinal (mainly colon) tumors of various histological type in 100% of both rat strains and, in addition, caused Zymbal gland carcinomas in 79.7% of Wistar rats. Pretreatment with disulfiram (DSF, 500 mg/kg), a known inhibitor of DMH metabolism, totally prevented intestinal and Zymbal gland tumors in Wistar rats. When DSF treatment started after the first DMH injection, the protective effect was not total, the incidence and multiplicity of both types of tumors being comparable to those observed following a single injection of the carcinogen alone. This confirms the involvement of DSF in the initiation stage only of DMH carcinogenesis. A complete prevention of intestinal tumors in BD6 rats was also produced not only by the DSF metabolite carbon disulfide (250 mg/kg) but also by the hepatotoxic agent carbon tetrachloride (1.5 ml/kg), which suggests that the block of DMH metabolism in rat liver is not an exclusive property of thiono-sulfur compounds. Butylated hydroxytoluene (BHT) decreased the multiplicity of intestinal tumors, but not to a significant extent. BHT and the aforementioned metabolic inhibitors were administered by gavage in corn oil, which per se did not significantly decrease intestinal or Zymbal gland tumors. All remaining modulators were administered with drinking water. Two additional antioxidants triggered opposite effects on the multiplicity of intestinal tumors. In fact, sodium selenite (10 mg/l) significantly decreased the number of tumors, whereas ascorbic acid (10 g/l), irrespective of its combination with CaCl2, produced a marked enhancement. The alkali metal salts CaCl2 and KCl (both at 5 g/l) as well as the methylxanthines caffeine and theophylline (both at 600 mg/l) were devoid of significant effects. Neither treatment with DMH alone nor its association with test modulators was accompanied by significant changes in body weight gain or survival of animals. On the whole, depending on the mechanisms involved, the comparative study of test compounds led to a broad array of effects on DMH carcinogenesis, ranging from complete inhibition to significant enhancement. The resulting picture can be visualized at a glance in Figure 1 of this article.
 
Article
The synthesis of enantiomerically pure unsaturated long chain 1,2-diamines and amino alcohols was carried out starting from the corresponding non-natural alpha-amino acids. The in vitro cytotoxicity of the compounds prepared was evaluated against six solid tumor cell lines (A2780, H322, LL, WiDr, C26-10 and UMSCC-22B). Free 1, 2-diamines proved to be the most active compounds exhibiting IC50 values between 2.0 mM and 3.3 mM.
 
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A dose response study in carcinogenesis by 1,2-dimethylhydrazine dihydrochloride (1,2-DMH) in Swiss mice was performed. The compound was administered continuously in drinking water for life from 6 weeks of age at dose levels of 0.002, 0.001, 0.0005, 0.00025, 0.000125, 0.0000625, 0.00003125 and 0.000015625%. A positive correlation was established between the dose levels of 1,2-DMH and the yield of blood vessel tumors. In addition, the latency periods for these tumors diminished when the dose levels of the carcinogen decreased. Although some of the treatments induced lung tumors, no association was observed between the dose levels of the carcinogen and the lung tumor incidence. The study thus provides an example in which the continuous lifespan administration of a carcinogen resulted in a partial dose response effect. This method of administration closely resembles some of the human exposure situation.
 
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1-Acyl- and 1,2-diacyl-1,2,4-triazolidine-3,5-diones were found to be potent cytotoxic agents in murine and human cancer cell lines, e.g. L1210, P388, Tmolt3, colon adenocarcinoma, Hela cells and glioma. In vivo activity was demonstrated at 8 mg/kg/day against Ehrlich ascites carcinoma growth. In L1210 cells, 1-acetyl-4-phenyl-1,2,4-triazolidine-3,5-dione, 41, reduced DNA synthesis significantly with moderate reduction in RNA synthesis. Enzyme sites in L1210 cells which were markedly affected were m- and r-RNA polymerase, PRPP amidotransferase, IMP dehydrogenase, dihydrofolate reductase, thymidine, TMP and TDP kinases. Kinetic studies suggest the inhibition of rate limiting enzymes in the purine pathway by 41 was responsible for its cytotoxicity. Acute toxicity studies in mice indicated 41 was safe for therapeutic use at 20, 50, and 100 mg/ky/day.
 
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1,2-Dimethylhydrazine dihydrochloride (1,2-DMH) was given to Swiss mice in 1 or 10 weekly intracolonic instillations of 60 and 40 micrograms/g body weight, respectively. In mice that received a single treatment, the tumor incidences in the lungs, blood vessels and subcutis were 34% (p less than 0.006), 8 and 25 in females and 2, 20 and 0% in males, respectively. In mice treated repeatedly with the carcinogen, the corresponding tumor incidences were 34% (p less than 0.006), 38% (p less than 0.000001) and 12% (p less than 0.03) in females and 34, 10 and 0% in males, respectively. In control mice the tumor incidences in lungs, blood vessels and subcutis were 15, 8 and 2% in females and 22, 5 and 2% in males, respectively. The induction of large intestinal cancer, which was the main objective of this investigation, however, failed to materialize.
 
Top-cited authors
Da-Tian Bau
  • China Medical University (ROC)
Kosei Hirakawa
  • Osaka City University
Ondrej Topolcan
  • Charles University in Prague
Hitoshi Fujiwara
  • Kyoto Prefectural University of Medicine
Wen G Jiang
  • Cardiff University