Protective Effects of Amino Acids on Plasmid DNA Damage Induced by Therapeutic Carbon Ions.

Radioprotectors with few side effects are useful for carbon-ion therapy, which directly induces clustering damage in DNA. With the aim of finding the most effective radioprotector, we investigated the effects of selected amino acids which might have chemical DNA-repair functions against therapeutic carbon ions. In the current study, we employed five amino acids: tryptophan (Trp), cysteine (Cys), methionine (Met), valine (Val) and alanine (Ala). Samples of supercoiled pBR322 plasmid DNA with a 17 mM amino acid were prepared in TE buffer (10 mM Tris, 1 mM ethylenediaminetetraacetic acid, pH 7.5). Phosphate buffered saline (PBS) was also used in assays of the 0.17 mM amino acid. The samples were irradiated with carbon-ion beams (290 MeV/u) on 6 cm spread-out Bragg peak at the National Institute of Radiological Sciences and Heavy Ion Medical Accelerator in Chiba, Japan. Breaks in the DNA were detected as changes in the plasmids and quantified by subsequent electrophoresis on agarose gels. DNA damage yields and protection factors for each amino acid were calculated as ratios relative to reagent-free controls. Trp and Cys showed radioprotective effects against plasmid DNA damage induced by carbon-ion beam, both in PBS and TE buffer, comparable to those of Met. The double-strand break (DSB) yields and protective effects of Trp were comparable to those of Cys. The yields of both single-strand breaks and DSBs correlated with the scavenging capacity of hydroxyl radicals (rate constant for scavenging hydroxyl radicals multiplied by the amino acid concentration) in bulk solution. These data indicate that the radioprotective effects of amino acids against plasmid DNA damage induced by carbon ions could be explained primarily by the scavenging capacity of hydroxyl radicals. These findings suggest that some amino acids, such as Trp, Cys and Met, have good potential as radioprotectors for preventing DNA damage in normal tissues in carbon-ion therapy.

Potential Mechanisms for Protective Effect of D-Methionine on Plasmid DNA Damage Induced by Therapeutic Carbon Ions.

D-methionine (D-met), a dextrorotatory isoform of the amino acid L-methionine (L-met), can prevent oral mucositis and salivary hypofunction in mice exposed to radiation. However, the mechanism of its radioprotection is unclear, especially with regard to the stereospecific functions of D-met. Radiation is known to cause injury to normal tissue by triggering DNA damage in cells. Thus, in this study we sought to determine whether the chirality of D-/L-met affects radiation-induced events at the DNA level. We selected plasmid DNA assays to examine this effect in vitro, since these assays are highly sensitive and allow easy detection of DNA damage. Samples of supercoiled pBR322 plasmid DNA mixed with D-met, L-met or dimethylsulfoxide (DMSO) were prepared and irradiated with a Bragg peak beam of carbon ions (∼290 MeV/u) with a 6-cm spread. DNA strand breaks were indicated by the change in the form of the plasmid and were subsequently quantified using agarose gel electrophoresis. We found that D-met yielded approximately equivalent protection from carbon-ion-induced DNA damage as DMSO. Thus, we propose that the protective functions of methionine against plasmid DNA damage could be explained by the same mechanism as that for DMSO, namely, hydroxyl radical scavenging. This stereospecific radioprotective mechanism occurred at a level other than the DNA level. There was no significant difference between the radioprotective effect of D-met and L-met on DNA.

Monitoring Mitochondrial Complex-I Activity Using Novel PET Probe 18F-BCPP-EF Allows Early Detection of Radiotherapy Effect in Murine Squamous Cell Carcinoma.

OBJECTIVES: Aerobic glycolysis, the main pathway of energy production in tumors (Warburg effect) allows detection of tumors by positron emission tomography (PET) using 18F-fluoro-2-deoxy-D-glucose (18F-FDG). Since ionizing radiation (IR) is reported to switch aerobic glycolysis to mitochondrial oxidative phosphorylation, radiotherapeutic efficacy was monitored by the activity of mitochondrial complex I (MC-I), using a new PET probe 18F-BCPP-EF, 18F-2-tert-butyl-4-chloro-5-{6-[2-(2-fluoro-ethoxy)-ethoxy] -pyridine-3-ylmethoxy}-2H-pyridazin-3-one, compared with 18F-FDG uptake and the apoptosis index. METHODS: Tumor uptake of 18F-BCPP-EF or 18F-FDG was examined in C3H/HeN mice inoculated with murine squamous cell carcinoma SCCVII at various time points after a single dose of x-ray irradiation at 0, 6, 15, or 30 Gy. Apoptosis incidence was determined by TUNEL staining in excised tumor tissue. RESULTS: Tumor growth suppression was dose-dependent; tumor grew 10-fold (0 Gy), 5-fold (6 Gy), 2-fold (15 Gy), and reduced to half in its volume (30 Gy) 14 days after treatment. 18F-BCPP-EF uptake was significantly increased as early as 3 days after 15 Gy or 30 Gy, when tumor size and apoptosis index showed no difference among radiation doses. In contrast, 18F-FDG uptake was initially increased dose-dependently, remained elevated up to 7 days, and eventually decreased 10 days after 30 Gy and also 14 days after 15 Gy when tumor size was already reduced. Apoptosis index was increased after irradiation but failed to correlate with tumor response. CONCLUSION: Tumor uptake of 18F-BCPP-EF was increased dose-dependently early after effective doses of IR when 18F-FDG uptake as well as apoptosis incidence were not indicative of tumor response. The results suggest that 18F-BCPP-EF is a promising "positive" MC-I imaging PET probe for early detection of efficacy of tumor radiotherapy.

Liposome-Encapsulated Hemoglobin Improves Tumor Oxygenation as Detected by Near-Infrared Spectroscopy in Colon Carcinoma in Mice.

Liposome-encapsulated hemoglobin (LEH) with high (h-LEH, P50 O2  = 10 mm Hg) or low O2 affinity (l-LEH, P50 O2  = 40 mm Hg) may improve O2 delivery to sensitize tumor tissues for radiotherapy. A total of 10 mL/kg of h-LEH, l-LEH, red blood cells (RBCs), or saline was infused in mice transplanted with murine colon carcinoma with near-infrared spectroscopy (NIRS) detectors set at the tumor (right leg) and intact muscle (left leg). NIRS recorded changes in the amount of oxyhemoglobin (oxyHb), deoxyhemoglobin (deoxyHb), and their sum (tHb) with the animals spontaneously breathing room air (10 min), pure O2 (5 min), and then back to room air. The tumor was finally excised for histological examination. In mice treated with h-LEH, tHb significantly increased compared to mice receiving other solutions. The magnitude was significantly attenuated in the tumor compared to the intact muscle under room air. Reciprocal changes in oxyHb and deoxyHb between intact muscle and tumor in response to infused solutions allowed assumption of average tissue PO2 between 30 and 40 mm Hg in muscle and at around 10 mm Hg in tumor. While O2 respiration increased oxyHb and decreased deoxyHb both in muscle and tumor, their sum or tHb consistently decreased in muscle and increased in tumor regardless of preceding infusion. Such responses were totally reversed when mice were placed under hypoxia (10% O2 ), suggesting that a lack of physiological circulatory regulation in tumor may account for heavier immunohistochemical staining for human hemoglobin in tumors of mice treated with h-LEH than with l-LEH. The results suggest that h-LEH may cause significant tumor oxygenation compared to RBC, l-LEH, or saline probably due to its nanometer size (vs. RBC) and high O2 affinity (vs. l-LEH) without increasing O2 content in the intact tissue (vs. O2 respiration) probably due to a lack of physiological circulatory regulation.

Liposome-encapsulated hemoglobin enhances chemotherapy to suppress metastasis in mice.

Liposome-encapsulated hemoglobin with high O2 -affinity (P50 O2 = 10 mm Hg, h-LEH) was reported to enhance tumor radiosensitivity. We hypothesize that targeted O2 delivery to tumor hypoxia by h-LEH may also enhance chemotherapy to suppress tumor growth and metastasis in mice. Doxorubicin (DXR; 0.5 or 2 mg/kg i.p.) or S-1 (4 or 8 mg/kg orally) alone or in combination with h-LEH (5 mL/kg i.v.) was administered for 2 weeks to C57BL/6N mice inoculated with Lewis Lung Carcinoma (LLC) in the leg. After the 2-week therapy in six treatment groups, mice were sacrificed for quantitative assessment of tumor growth and lung metastasis. The tumor was then evaluated for its expression of hypoxia-inducible factor-1α (HIF-1α) and matrix metallopoteinase-2 (MMP-2) activity. Combined use of h-LEH and chemotherapeutic agents (DXR or S-1) showed no additional enhancement on suppression of the tumor growth over the chemotherapeutic agent alone. However, the combination use of h-LEH significantly suppressed the number and total area of metastatic colonies in the lung compared with each chemotherapeutic agent alone. Although HIF-1α expression and MMP-2 activity in the original tumor was significantly suppressed in the groups of mice treated with either DXR or S-1 alone, the addition of h-LEH to either agent showed further enhancement of oxygen-mediated degradation of HIF-1α and suppression of MMP-2 activity. Although the addition of h-LEH to DXR or S-1 had little effect on original LLC tumor growth, it significantly enhanced suppression of lung metastasis in mice.

Effects of liposome-encapsulated hemoglobin on gastric wound healing in the rat.

Liposome-encapsulated hemoglobin (LEH) may improve microcirculation and oxygen (O2 ) metabolism at a surgical wound to accelerate its healing. Ten mL/kg of LEH with high (h-LEH) or low O2 -affinity (l-LEH), homologous red blood cells (RBC), empty liposome or saline as a control was infused before a 10-mm incision and interrupted suture closure of the gastric wall in a total of 110 rats. Two and 4 days later, the stomach was excised for bursting pressure determination and histological sampling. The dose-response relationship was examined in 70 additional rats receiving progressively reduced doses of h-LEH. Hypoxia-inducible factor-1α (HIF-1α) was stained immunohistochemically in 54 other rats to examine its accumulation at the anastomotic sites. Bursting pressure of the surgical wound was significantly higher 2 days after surgery only in the h-LEH-treated rats (P < 0.05), but not at 4 days after surgery, when other rats showed increased bursting pressure to a nonsignificant level. Histological examination revealed less granulocyte infiltration, better granulation, and more macrophage infiltration in h-LEH-treated rats at 2 days, but no longer at 4 days postsurgery. Dose-response study revealed that 0.4 mL/kg of h-LEH (hemoglobin 24 mg/kg) was effective for elevating bursting pressure at 2 days. h-LEH-treated rats had significantly suppressed HIF-1α accumulation in the wound 6, 24, and 48 h after surgery as compared with control animals treated with homologous RBC or saline. In conclusion, the results suggest that h-LEH, but not l-LEH or homologous transfusion, may accelerate wound healing early after gastric incision and anastomosis in the rat. The mechanism(s) appears to be related to improved O2 supply, aerobic metabolism, and suppressed inflammation in the wound.

Liposome-encapsulated hemoglobin accelerates bronchial healing after pneumonectomy in the rat with or without preoperative radiotherapy.

Liposome-encapsulated hemoglobin (LEH) has been reported to accelerate wound healing in the stomach and skin in an experimental setting. LEH was tested in bronchial anastomotic healing after radiation and pneumonectomy in the rat. Sprague-Dawley rats (n = 61) received preoperative radiation (20 Gy) to the chest and underwent left pneumonectomy with bronchial stump closure using the Sweet method 4 days later, when they were randomized to receive intravenous infusion of LEH with high O2 affinity (P50 O2 = 17 mm Hg, 10 mL/kg, n = 32) or saline (n = 29). Additional rats (n = 18) were treated in the same way without preoperative radiation. Bronchial anastomotic healing was evaluated 2 days after surgery by determining the bursting pressure and infiltration of neutrophils, monocytes, and macrophages. Bronchial bursting pressure was elevated in the rats receiving LEH both in the unirradiated group (LEH 212 ± 78 vs. saline 135 ± 63 mm Hg, P < 0.05) and in rats with preoperative radiation (LEH 162 ± 48 vs. saline 116 ± 56 mm Hg, P < 0.01). Moreover, the percentage of rats with bursting pressure <100 mm Hg tended to be smaller in the unirradiated group (LEH 1/9 [11.1%] vs. saline 4/9 [44.4%], NS) and was significantly reduced in irradiated animals (LEH 3/32 [9.4%] vs. saline 11/29 [38%], P < 0.05). There were no morphological differences except for macrophage infiltration to the anastomotic area, which was significantly prominent in the LEH-treated rats (P < 0.05) regardless of the presence or absence of preoperative irradiation (IR). The results suggest that LEH with high O2 affinity may improve mechanical strength and morphological findings in bronchial anastomosis in rats regardless of the presence or absence of preoperative IR. The irradiated rats later treated with LEH had equivalent or better bronchial healing than that of saline-treated naïve animals undergoing pneumonectomy alone.

Biopsy specimens obtained 7 days after starting chemoradiotherapy (CRT) provide reliable predictors of response to CRT for rectal cancer.

PURPOSE: Preoperative chemoradiation therapy (CRT) significantly decreases local recurrence in locally advanced rectal cancer. Various biomarkers in biopsy specimens obtained before CRT have been proposed as predictors of response. However, reliable biomarkers remain to be established. METHODS AND MATERIALS: The study group comprised 101 consecutive patients with locally advanced rectal cancer who received preoperative CRT with oral uracil/tegafur (UFT) or S-1. We evaluated histologic findings on hematoxylin and eosin (H&E) staining and immunohistochemical expressions of Ki67, p53, p21, and apoptosis in biopsy specimens obtained before CRT and 7 days after starting CRT. These findings were contrasted with the histologic response and the degree of tumor shrinkage. RESULTS: In biopsy specimens obtained before CRT, histologic marked regression according to the Japanese Classification of Colorectal Carcinoma (JCCC) criteria and the degree of tumor shrinkage on barium enema examination (BE) were significantly greater in patients with p21-positive tumors than in those with p21-negative tumors (P=.04 and P<.01, respectively). In biopsy specimens obtained 7 days after starting CRT, pathologic complete response, histologic marked regression according to both the tumor regression criteria and JCCC criteria, and T downstaging were significantly greater in patients with apoptosis-positive and p21-positive tumors than in those with apoptosis-negative (P<.01, P=.02, P=.01, and P<.01, respectively) or p21-negative tumors (P=.03, P<.01, P<.01, and P=.02, respectively). The degree of tumor shrinkage on both BE as well as MRI was significantly greater in patients with apoptosis-positive and with p21-positive tumors than in those with apoptosis-negative or p21-negative tumors, respectively. Histologic changes in H&E-stained biopsy specimens 7 days after starting CRT significantly correlated with pathologic complete response and marked regression on both JCCC and tumor regression criteria, as well as with tumor shrinkage on BE and MRI (P<.01, P<.01, P<.01, P<.01, and P=.03, respectively). CONCLUSIONS: Immunohistochemical expressions of p21 and apoptosis together with histologic changes on H&E-stained biopsy specimens obtained 7 days after starting CRT are strong predictors of the response to CRT.

Liposome-encapsulated hemoglobin ameliorates tumor hypoxia and enhances radiation therapy to suppress tumor growth in mice.

We hypothesize that liposome-encapsulated hemoglobin with high O2 affinity (P5002 = 12 mm Hg, h-LEH) may increase O2 delivery to hypoxic tumors and enhance radiation therapy synergistically to suppress tumor growth. First, h-LEH (5, 10, and 20 mL/kg) was intravenously infused 30 min before radiation (20 Gy) of SCCVII tumor grown in C3H/HeN mice. Second, 10 mL/kg of h-LEH was administered 30, 60, 90, and 120 min prior to radiation to determine optimal timing. Tumor size was monitored thereafter to titrate tumor growth suppression. Third, additional mice with SCCVII tumor were infused with h-LEH or empty liposome (EL), and tumors were excised at various time points for immunohistochemical examination of h-LEH and hypoxia-inducible factor-1α (HIF-1α). h-LEH was most effective at 10 mL/kg in comparison to 5 or 20 mL/kg of h-LEH or EL. Tumor growth was most suppressed when the interval between h-LEH infusion and radiation was shortest, 30 min. As a result, 10 mL/kg of h-LEH infusion 30 min prior to radiation prolonged 5-fold tumor-growth time from 20.0 days (radiation and EL) to 26.5 days, P<0.01, synergy ratio 1.42. While human hemoglobin (h-LEH) was detected in tumors 0.5 to 24 h after administration, HIF-1α accumulation was sparse and became significantly reduced compared to controls 48 and 72 h after h-LEH infusion. h-LEH (10 mL/kg) was highly effective in enhancing radiation therapy synergistically under ambient respiration against tumor growth in mice. Decreased accumulation of HIF-1α in h-LEH-treated tumor may suggest targeted tumor oxygenation as a potential mechanism.

Phase I/II study of preoperative concurrent chemoradiotherapy with S-1 for locally advanced, resectable rectal adenocarcinoma.

PURPOSE: To assess the maximum tolerability of a combination of S-1 and preoperative radiotherapy and to evaluate the feasibility and activity in patients with locally advanced rectal cancer. METHODS: Patients (n = 30) with adenocarcinoma of the middle or lower rectum were enrolled in a phase I (n = 9) and/or phase II (n = 21) trial. A total dose of 45 Gy was delivered in 25 fractions over 5 weeks, and S-1 was orally administered twice a day on days 1-14 and 22-35. Surgical resection was scheduled 4-8 weeks after the completion of chemoradiation. RESULTS: In phase I, the recommended dose (RD) of S-1 was 80 mg/m(2)/day, and the maximum-tolerated dose was never reached. A total of 27 cases, including the 6 RD cases in phase I, were enrolled in phase II. In phase II, a pathological complete response (pCR) was observed in 6/27 patients (22%), pathological downstaging was observed in 21/27 patients (78%), and a tumor volume reduction of 69 ± 22% was obtained. These results were similar to the previously reported pCR rates of 16-18%, pathological downstaging rates of 49-59%, and tumor volume reduction of 68% after chemoradiotherapy with capecitabine. Grade 3 adverse events consisted of one case of leukopenia (4%), 2 cases of anemia (7%) and 3 cases of diarrhea (11%). Overall, the adverse events were very mild. Hand-foot syndrome was not observed. CONCLUSION: The efficacy of chemoradiotherapy with S-1 seems to be equivalent to the efficacy reported for chemoradiotherapy with capecitabine, but the adverse events were much milder, although further study is warranted.

Effects of preoperative immunochemoradiotherapy and chemoradiotherapy on immune responses in patients with rectal adenocarcinoma.

BACKGROUND: We previously reported that preoperative chemoradiotherapy (CRT) combined with intraoperative electron irradiation for cT3/T4 adenocarcinoma of the rectum reduced the local recurrence, with significant improvement of survival. Radiotherapy has been reported to reduce immune function. We examined the effects of PSK, a protein-bound polysaccharide, concomitant with preoperative CRT on immune responses. PATIENTS AND METHODS: Thirty patients with cT3/T4 adenocarcinoma of the rectum were randomly assigned to 2 weeks' irradiation and 4 weeks' S-1 administration before surgery (control group), or the same CRT with simultaneous 4 weeks' PSK administration (PSK group). Both systemic and local immune responses were evaluated. RESULTS: Significant increase of natural killer cell count in the peripheral blood and cytotoxic T-cell counts in the peri-tumoral and normal mucosa, and a significant decrease of serum immunosuppressive acidic protein level were observed in the PSK group. CONCLUSION: Combined use of PSK with preoperative CRT may improve immune function.

Molecular determinants of folate levels after leucovorin administration in colorectal cancer.

PURPOSE: Oral leucovorin (LV) is used with uracil/tegafur (UFT) in the treatment of colorectal cancer (CRC). In order to find the factors related to the efficacy of LV in enhancing the antitumour effect of UFT, we investigated the relationships between the reduced folate levels in the CRC tissue after LV administration and the gene-expression levels of folate-metabolizing enzymes and folate transporters. METHODS: The subjects were 60 CRC patients, scheduled to undergo surgery. The control group (n = 30) did not receive LV. Three groups (n = 10 for each) received a single dose of oral LV at 25 mg, 4, 12 or 18 h before surgery (LV 4 h, LV 12 h or LV 18 h groups, respectively). The reduced folate levels in plasma and tissues were measured by high-performance liquid chromatography (HPLC) or a thymidylate synthase-FdUMP binding assay, respectively. The intratumoral expression levels of 34 genes were quantitatively evaluated with a real-time polymerase chain reaction (RT-PCR) assay. RESULTS: The reduced folate levels persisted for a longer period of time in the CRC tissue than in the plasma after LV administration. A multivariate logistic regression analysis revealed that high folylpolyglutamate synthase (FPGS) gene expression, low gamma-glutamyl hydrolase (GGH) gene expression and low ATP-binding cassette sub-family C, number 1 (ABCC1) gene expression in CRC tissues were predictive factors for a high reduced folate level after LV administration. CONCLUSIONS: The expression level of FPGS, GGH and ABCC1 in CRC tissues could predict the reduced folate level after LV administration, and these factors may determine the efficacy of LV treatment.

Prognostic factors in patients with synchronous peritoneal carcinomatosis (PC) caused by a primary cancer of the colon.

BACKGROUND: Peritoneal carcinomatosis (PC) is seen in about 10% of patients with colon cancer during the initial operation and has been considered a preterminal condition. The actual outcome can vary extensively depending on the presence/absence of metastases other than PC. METHODS: A total of 975 consecutive patients with colon cancer who underwent resection were included. The extent of PC was determined at laparotomy. Metastases restricted to the adjacent peritoneum or a few metastases to the distant peritoneum were classified as "limited," whereas numerous metastases to the distant peritoneum were as "extensive" regardless of the sizes of the disseminated nodules. RESULTS: PC group consisted of 75 patients (7.7%). The median survival time (MST) in the PC group was 6.8 months. Survival was significantly better in cases with limited PC (MST, 12.4 months), without lymph node involvement (20.8 months), with preoperative performance status of 0 or 1 (8.5 months), and who received chemotherapy more than 3 months (8.8 months). A multivariate analysis revealed that these four factors were significant predictors of better outcome. CONCLUSIONS: The extent of PC and lymph node involvement, even if the distribution is confined around the primary lesion, are more accurate prognostic factors than distant metastasis in patients with colon cancer and synchronous PC.

Evaluation of D-18F-FMT, 18F-FDG, L-11C-MET, and 18F-FLT for monitoring the response of tumors to radiotherapy in mice.

UNLABELLED: O-18F-fluoromethyl-D-tyrosine (D-18F-FMT) is a promising novel agent for tumor imaging by PET. The aim of this study was to evaluate the potential of D-18F-FMT and the other conventional ligands used for tumor imaging, namely, 18F-FDG, L-11C-methionine (L-11C-MET), and 3'-deoxy-3'-18F-fluorothymidine (18F-FLT), as a PET ligand for monitoring early responses to radiotherapy in tumor-bearing mice. METHODS: C3H/HeN mice inoculated with murine squamous cell carcinomas were treated with a single dose of x-ray irradiation at 2, 6, 20, or 60 Gy. Tumor uptake of each ligand was examined 1, 3, and 7 d after the irradiation. RESULTS: Tumor uptake of D-18F-FMT was decreased on day 1 after irradiation at 6, 20, or 60 Gy, and the decrease persisted until day 7. Tumor uptake of 18F-FDG was elevated on days 1 and 3 after irradiation at 2, 6, or 20 Gy, followed by a decrease in uptake on day 7 in mice irradiated at 20 or 60 Gy. Decreased tumor uptake of L-11C-MET was observed only on day 3 after the irradiation. Decreased tumor uptake of 18F-FLT was detected on day 1 after irradiation at 2, 6, 20, or 60 Gy; thereafter, the dose-dependent decrease in uptake was no longer seen. Only for D-18F-FMT were significant positive correlations found between ligand uptake at all the time points examined and tumor volume on day 14 after various doses of irradiation. CONCLUSION: The findings suggest that D-18F-FMT is a promising PET ligand for early-phase detection and prediction of the effects of radiation therapy.

Long-term postoperative adjuvant chemotherapy of UFT/LV improves survival in a primary tumor resection-pulmonary metastasis model.

BACKGROUND: The optimum regimen and optimum duration of administration of postoperative chemotherapy would vary with the postoperative residual tumor volume. Whether or not prolonged administration of oral uracil/tegafur (UFT) with leucovorin (LV) would prolong the survival period was assessed experimentally. MATERIALS AND METHODS: Murine models of pulmonary metastasis with different volumes of residual tumor after primary tumor resection were prepared, and the efficacy of 12-week and 4-week oral administration of UFT/LV as postoperative adjuvant chemotherapy was compared. RESULTS: In the model with only a small volume of occult residual tumor after early resection of the primary tumor, the survival period in the 12-week UFT/LV group was significantly increased as compared with that in the untreated group, whereas no significant difference was noted between the 4-week UFT/LV group and the untreated group. CONCLUSION: Long-term administration of UFT/LV as postoperative adjuvant chemotherapy may be potentially beneficial.

Phase I/II study of 24-hour infusion of irinotecan combined with oral UFT for metastatic colorectal cancer.

BACKGROUND: To evaluate the efficacy and safety of irinotecan combined with UFT for untreated and pretreated metastatic colorectal cancer. METHODS: Escalating doses of irinotecan (80-110 mg/m(2)) were administered by 24-hour infusion on day 1. UFT was administered orally at 400 mg/m(2)/day on days 3-7 and 10-14. The treatment cycles were repeated every 2 weeks. RESULTS: In the phase I study, the maximum tolerated dose of irinotecan was 110 mg/m(2) and the recommended dose for the phase II study was determined to be 100 mg/m(2). Thirty-five patients including 3 patients at the recommended dose in the phase I study were evaluated in the phase II study. The grade 3/4 toxicities observed were leukopenia, neutropenia, thrombocytopenia and anemia. No grade 3 or more severe nonhematological toxicities were noted. The response rate was 62.9% and the median overall survival 16.7 months. CONCLUSIONS: A 24-hour infusion of irinotecan combined with UFT is feasible and active for metastatic colorectal cancer.

Dihydropyrimidine dehydrogenase activity during long-term adjuvant treatment with oral uracil and tegafur for colorectal cancer.

BACKGROUND: Dihydropyrimidine dehydrogenase (DPD) is a rate-limiting enzyme for the degradation of 5-fluorouracil. The effect of long-term treatment with oral fluoropyrimidines on DPD activity has not been investigated. This study was conducted to examine changes in DPD activity in peripheral mononuclear cells (PMNC) during long-term treatment with oral uracil and tegafur (UFT) for colorectal cancer. METHODS: UFT was administered for 5 consecutive days and not administered the next 2 days for 6 months after surgery. PMNC-DPD activity was measured before and 1, 2, 4 and 6 months after starting UFT administration. RESULTS: In 70 eligible patients, there were no significant variations of PMNC-DPD activity during postoperative administration of UFT for 6 months. Grade 2 or higher adverse events were observed in significantly more patients with low DPD than with high DPD activity (p = 0.018). CONCLUSION: There were no significant variations of PMNC-DPD activity during the postoperative administration of UFT for 6 months. Low PMNC-DPD activity before UFT treatment was considered to be a predicting factor for toxicity.

Predictors of tumor downsizing and regression with preoperative radiotherapy alone and with concomitant tegafur/uracil (UFT) for resectable advanced rectal adenocarcinoma.

BACKGROUND/AIMS: Predictors of sensitivity to preoperative radiotherapy (RT) may differ from those of chemo-radiotherapy (CRT). This study attempts to evaluate retrospectively the significance of apoptosis-related and proliferative indexes in biopsy specimens obtained before treatment as predictors of sensitivity to RT or to CRT for locally advanced rectal adenocarcinoma. METHODOLOGY: The subjects were 96 patients with clinical T3-4/Nx/M0 adenocarcinoma of the middle third or lower third of the rectum. Sixty-one patients were treated with preoperative RT alone (20 Gy in 10 fractions) [RT group] during 1991-1998, and 35 patients received concurrent oral tegafur/uracil (UFT) [CRT group] since 1999. Radical surgery including TME and pelvic nerve preservation with 15 Gy of intraoperative RT was performed two weeks after completion of the preoperative radiation. We evaluated apoptotic index (AI) and p53, p21 and Ki-67 protein expression in the biopsy specimens, and histological differentiation, pathologic regression in the resected specimens and the degree of tumor shrinkage based on the double contrast barium enema images. RESULTS: AI-positivity, p53-negativity, p21-positivity and well differentiated adenocarcinoma were predictors of high sensitivity in RT group, whereas AI-positivity alone was the predictor in CRT group. The addition of UFT to RT increased sensitivity in patients with p53-positivity, p21-negativity and moderately differentiated adenocarcinoma. CONCLUSIONS: Predictors of sensitivity are different between RT and CRT.

Detection of carcinoembryonic antigen messenger RNA-expressing cells in peripheral blood 7 days after curative surgery is a novel prognostic factor in colorectal cancer.

BACKGROUND: The significance of detection of circulating cancer cells in blood during surgery in patients with colorectal cancer (CRC) remains controversial. Experimental study revealed that the cancer cells injected from the vein disappeared completely until 7 days. The aim of this study was to clarify that the detection of circulating cancer cells in blood taken later than 7 days after curative surgery may be a prognostic factor. METHODS: Two hundred consecutive patients with CRC who underwent potentially curative surgery were the subjects. Peripheral blood was collected between 7 and 10 days after resection. Cancer cells were detected using reverse transcriptase-polymerase chain reaction targeting carcinoembryonic antigen (CEA) messenger RNA (mRNA). The median follow-up period was 52 months (range: 34-69 months). RESULTS: The overall positive incidence of CEA mRNA was 22%. Detection of CEA mRNA was not significantly related to conventional clinicopathological findings. Recurrence has been confirmed in 55 patients (28%). The recurrence rate was significantly higher in patients with rectal cancer, deep penetration, lymph node metastasis, preoperative chemoradiotherapy and positive CEA mRNA. The CEA mRNA positive patients showed significantly poorer disease free survival (DFS) and overall survival (OS) than the negative patients (DFS, P = 0.007; OS, P = 0.04). Multivariate analysis revealed that the positive expression of CEA mRNA (P < 0.01) as well as the tumor location and TNM stage classification was identified as the significant risk factors for recurrence. CONCLUSIONS: Detection of CEA mRNA expressing cells in peripheral blood 7 days after curative surgery is a novel independent factor predicting recurrence in patients with CRC.

Contribution of indirect action to radiation-induced mammalian cell inactivation: dependence on photon energy and heavy-ion LET.

The contribution of indirect action mediated by OH radicals to cell inactivation by ionizing radiations was evaluated for photons over the energy range from 12.4 keV to 1.25 MeV and for heavy ions over the linear energy transfer (LET) range from 20 keV/microm to 440 keV/microm by applying competition kinetics analysis using the OH radical scavenger DMSO. The maximum level of protection provided by DMSO (the protectable fraction) decreased with decreasing photon energy down to 63% at 12.4 keV. For heavy ions, a protectable fraction of 65% was found for an LET of around 200 keV/microm; above that LET, the value stayed the same. The reaction rate of OH radicals with intracellular molecules responsible for cell inactivation was nearly constant for photon inactivation, while for the heavy ions, the rate increased with increasing LET, suggesting a reaction with the densely produced OH radicals by high-LET ions. Using the protectable fraction, the cell killing was separated into two components, one due to indirect action and the other due to direct action. The inactivation efficiency for indirect action was greater than that for direct action over the photon energy range and the ion LET range tested. A significant contribution of direct action was also found for the increased RBE in the low photon energy region.