Inhibition of chronic ulcerative colitis-associated colorectal adenocarcinoma development in a murine model by N-acetylcysteine

Long-term ulcerative colitis (UC) patients are at increased risk for developing colorectal cancer. In order to develop strategies for preventing UC-associated carcinogenesis, we studied the effect of the antioxidant N-acetylcysteine (NAC) on UC-associated cancer development in a mouse model. Female C57BL/6J mice were subjected to long-term administration of dextran sulfate sodium (DSS) in the drinking fluid and 2-fold iron-enriched AIN76A diet, with or without NAC. In the DSS-plus-2-fold iron positive control group, gross tumor incidence was 88.5% (23/26 mice) after 12 DSS cycles (1 DSS cycle = 7 day DSS treatment period followed by 10 day recovery period). The tumor multiplicity was 2.1 +/- 0.2 tumors/tumor-bearing mouse, and the tumor volume was 0.054 +/- 0.019 cm3. With 0.2% NAC administration, tumor incidence was significantly reduced (68%, 17/25 mice; P < 0.05), as was the tumor multiplicity (1.5 +/- 0.1 tumors/tumor-bearing mouse; P < 0.05). The tumor volume was lower (0.014 +/- 0.004 cm3), but not significantly decreased. The proliferation index was significantly decreased in non-cancerous epithelia (48.5 +/- 6.0% vs 32.0 +/- 3.7%; P < 0.05), but not in tumor cells. NAC significantly induced apoptosis in both non-cancerous epithelia and colorectal adenocarcinoma. The number of cells immunostained-positive for nitrotyrosine was markedly decreased in the non-cancerous mucosa of NAC-treated mice (102.4 +/-16.6 positive cells/mm2 mucosa vs 53.6 +/- 14.9 cells/mm2; P < 0.05). In addition, the number of inducible nitric oxide synthase (iNOS)-positive inflammatory cells in the non-cancerous mucosa of the distal colon was markedly decreased by NAC. This study indicates that the antioxidant NAC has the potential to serve as a preventive agent for UC-associated colorectal cancer, possibly via inhibition of cellular proliferation and nitrosative stress-caused cellular damage.     

Identification of mucin-depleted foci in the unsectioned colon of azoxymethane-treated rats: correlation with carcinogenesis

We tested the association between aberrant crypt foci (ACF) and tumor induction by feeding azoxymethane-induced rats (15 mg/kg x 2, s.c.) with synbiotics (Raftilose Synergy 1, a derivative of inulin, 10% of the diet, along with lactobacilli and bifidobacteria). After 16 weeks of feeding, synbiotics significantly increased ACF multiplicity. On the contrary, after 32 weeks, synbiotics significantly decreased intestinal tumors. When the same unsectioned colon used for ACF determination was stained with high-iron diamine Alcian blue, foci of crypts with scarce or absent mucins were identified. We defined these lesions as mucin-depleted foci (MDF), and they were visible in all azoxymethane-treated rats and correlated with tumor induction (MDF/colon: 8.2 +/- 0.9 and 3.8 +/- 0.9 in controls and synbiotic-fed rats, respectively, P < 0.01; crypts/MDF: 12.2 +/- 2 and 6.4 +/- 1 in controls and synbiotic-fed rats, respectively, P < 0.05, means +/- SE, n = 7). There were fewer MDF/colon than ACF, and they were histologically more dysplastic than mucinous lesions identified as ACF in high-iron diamine Alcian blue-stained colon. In conclusion, MDF may be premalignant lesions that predict colon carcinogenesis.     

SMS 201.995 inhibits in vitro and in vivo growth of human colon cancer.

The effect of a long-acting somatostatin analogue SMS 201.995 (SMS; Sandoz) on basal and gastrin-stimulated growth of 4 human colon cancer lines was studied in vitro and in vivo. Proliferation assay was done with overnight [75Se]selenomethionine uptake after 5 days of incubation. Gastrin concentrations used were 5e-10 M and 1e-7 M. SMS concentrations were from 2e-12 M to 2e-7 M. Cell lines LIM 1215, LIM 2405, and LIM 2412 were inhibited dose-dependently in both basal and gastrin-stimulated groups. LIM 1863 was slightly stimulated. Based on in vivo growth characteristics, LIM 2412 and LIM 2405 were selected for xenograft study. The dose of 50 micrograms/kg/day was arrived at after a preliminary experiment showed it to be safe and effective. The LIM 2412 xenografts in the SMS-treated animals were 473.3 +/- 99.9 (SD) versus 838.1 +/- 111.3 mm3 in control (P less than 0.05) after 20 days. The LIM 2405 tumors were also significantly inhibited (81.2 +/- 30.0 versus 245.7 +/- 48.3 mm3, P less than 0.01). The effect of SMS appeared to be reversible. Oral SMS at 200 micrograms/kg/day was not absorbed. This study suggests that SMS may have direct antitumor effects in human colon cancer.     

Osteopontin silencing by small interfering RNA suppresses in vitro and in vivo CT26 murine colon adenocarcinoma metastasis

Hepatic metastasis is a primary cause for failure of locoregional therapy in colorectal cancer. Increased expression of osteopontin (OPN), a ligand for alpha(v)beta3 integrin and CD44 receptors, is associated with metastasis in several types of cancer. However, the mechanism by which OPN mediates metastasis in colorectal cancer remains unknown. We hypothesized that OPN mediates invasion of colon cancer cells through basement membrane and migration through extracellular matrix (ECM). In this study, we used CT26 murine colon adenocarcinoma cells syngeneic to BALB/c mice to generate cell lines (pS-OPN) in which OPN expression was suppressed through small interfering RNA (siRNA) plasmids. CT26 wild-type cells (WT) and CT26 cells stably expressing murine-mismatch siRNA (pS-MM) served as controls. Western blotting quantified OPN protein levels and our most downregulated clone, pS-OPN-A4, demonstrated a mean 3.0-fold decrease in OPN protein expression versus WT. In vitro cell motility and invasiveness were decreased in pS-OPN-A4 by 3.6-fold (P = 0.004 versus WT) and 4.1-fold (P = 0.01 versus WT), but proliferation was similar amongst cell lines. We demonstrated that OPN suppression significantly correlates with MMP-2 downregulation. In vivo hepatic metastasis was assessed by quantifying liver weights and surface tumor nodules in 33 BALB/c mice (11/group) subjected to intrasplenic injection of tumor cells. pS-OPN-A4 resulted in a 50.4% decrease in mean liver weight compared with WT (3.79 +/- 1.49 g versus 1.88 +/- 1.34 g, P = 0.009). Only 18% of pS-OPN-A4 livers had >20 metastatic surface nodules compared with 89% for WT and 75% for pS-MM-V6. This study demonstrates that RNA interference stably reduces CT26 tumor expression of OPN and significantly attenuates CT26 colon cancer metastasis by diminishing tumor cell motility and invasiveness.     

Increased susceptibility of chronic ulcerative colitis-induced carcinoma development in DNA repair enzyme Ogg1 deficient mice

Ogg1 DNA repair enzyme recognizes and excises oxidative stress-caused 8-hydroxyl-deoxyguanosine (8-OHdG) from GC base-pairs. Ogg1 knockout mice are phenotypically normal, but exhibit elevated levels of 8-OHdG in nuclear and mitochondrial DNA, as well as moderately elevated mutagenesis and spontaneous lung tumors and UV-induced skin tumors. To elucidate the mechanistic role of inflammation-caused oxidative stress in carcinogenesis, the development of chronic ulcerative colitis (UC)-induced carcinoma in Ogg1 knockout mice was studied using a dextran sulfate sodium (DSS)-induced UC model without the use of a carcinogen. Ogg1 (-/-), Ogg1 (+/-), and wild type C57BL/6 mice were subjected to long-term, cyclic DSS treatment to induce chronic UC and carcinogenesis. In wild type C57BL/6 control mice after 15 cycles of DSS treatment, colorectal adenocarcinoma incidence was 24.1% (7/29 mice), with a tumor volume of 27.9 +/- 5.2 mm(3). Ogg1 (-/-) mice showed significantly increased adenocarcinoma development in the colon with a tumor incidence of 57.1% (12 of 21 mice, P < 0.05) and a tumor volume of 35.1 +/- 6.1 mm(3). Ogg1 mice (+/-) also exhibited significantly increased tumor development in the colon with a tumor incidence of 50.0% (13/26 mice) and a tumor volume of 29.1 +/- 7.2 mm(3). Histopathologic analyses revealed that colorectal tumors were well-differentiated tubular adenocarcinomas or mucinous carcinoma and adjacent colonic mucosa showed mild to moderate chronic UC. Using immunohistochemical approaches, Ogg1 (-/-) and (+/-) mice exhibited similar numbers and staining intensities of macrophages in UC areas as seen in Ogg1 (+/+) mice, but markedly increased numbers and staining intensities of 8-OHdG positive inflammatory and epithelial cells. These results provide important evidence on the relationship between inflammation-caused oxidative stress, DNA repair enzyme Ogg1, and carcinogenesis.     

NO-donating aspirin inhibits angiogenesis by suppressing VEGF expression in HT-29 human colon cancer mouse xenografts

The inhibitory effect of NO-donating aspirin (NO-ASA) on colon cancer has been demonstrated in vivo and in vitro but its mechanism is still obscure. We investigated the effect of NO-ASA on angiogenesis. Four groups of athymic mice (N = 12) bearing subcutaneous xenotransplants of HT-29 human colon cancer cells were injected intratumorally twice a week for 3 weeks with vehicle or m-NO-ASA or p-NO-ASA; the fourth group received no injections. The necrotic area of tumors, expressed as percentage of total area, was similar in the non-injected and vehicle-injected groups (51.8 +/- 2.8 versus 52.2 +/- 4.1, P > 0.05; mean +/- SEM for these and subsequent values). Compared with the vehicle group, the necrotic area of tumors was higher in the m-NO-ASA-treated (61.0 +/- 2.7, P < 0.02) and p-NO-ASA (65.8 +/- 2.4, P < 0.001)-treated groups. NO-ASA decreased microvessel density: vehicle = 11.7 +/- 0.8; m-NO-ASA = 7.8 +/- 0.6 (P = 0.0003 versus vehicle) and p-NO-ASA 6.2 +/- 0.7 (P = 0.0001 versus vehicle). The expression of vascular endothelial growth factor (VEGF) was significantly reduced in response to NO-ASA, with the p- isomer being more potent than the m-. NO-ASA altered the spatial distribution of VGEF expression, with 16.7% of the vehicle-treated xenografts displaying diminished VEGF in the inner region of the area between necrosis and the outer perimeter of the tumor, compared with those treated with m- (58.3%) or p-NO-ASA (75%, P < 0.01 for both versus control). Our findings indicate that NO-ASA suppresses the expression of VEGF, which leads to suppressed angiogenesis. The antiangiogenic activity of NO-ASA may be part of its antineoplastic effect.     

胃、结肠癌的癌组织周边区淋巴管的形态学特征

背景与目的:关于淋巴转移机制,大多数研究局限于淋巴管的分布。本研究的目的是观察在癌组织周边区的淋巴管形态、分布和淋巴管内皮细胞的变化。方法:取人胃癌、大肠癌手术标本各10例,制作大鼠结肠癌模型20例。在光镜下观察淋巴管形态及其内皮细胞的超微结构。使用图像分析仪,测量癌组织周边区和正常区淋巴管的数密度和体密度,计算淋巴管开放率和破坏率。结果:癌周边区的淋巴管管腔扩张、管壁被破坏;淋巴管内皮细胞被溶解破坏,遗留断裂的碎片,细胞器出现病理变化。癌周边区淋巴管的密度高于正常区。人结肠癌癌周边区和正常区的淋巴管数密度分别为(10.2±1.7)/mm2和(5.1±0.8)/mm2(P<0.05),体密度分别为(1.5±0.2)%和(0.7±0.0)%(P<0.05)。人胃癌癌周边区和正常区的淋巴管数密度分别为(8.0±0.9)/m m 2和(3.4±0.6)/m m 2 (P <0.01),体密度分别为(1.6±0.3)%和(0.8±0.2)%(P <0.05)。鼠结肠癌模型、人胃癌和结肠癌的癌周边区淋巴管开放率(22.2%、35.0%、25.8%)明显高于正常区(7.8%、8.0%、5.0%)(P <0.05)。鼠结肠癌模型和人胃癌的癌周边区淋巴管破坏率(20.1%和35.3%)也明显高于正常区(0和2.0%)(P <0.05)。结论:与正常组织比较,癌组织周边区淋巴管管腔扩张,管壁破坏,内皮细胞被溶解破坏,淋巴管密度增大。     

Increased susceptibility of Sf1(+/-) mice to azoxymethane-induced colon tumorigenesis

Aberrant transactivation of a certain set of target genes by the beta-catenin and T-cell factor-4 nuclear complex has been considered crucial for the initiation of colorectal carcinogenesis. We previously identified splicing factor-1 (SF1) as a novel component of the beta-catenin and T-cell factor-4 complex, and showed that the overexpression of SF1 inhibited the gene transactivational activity of the complex and markedly suppressed beta-catenin-evoked colony formation by human embryonic kidney 293 cells. However, the involvement of SF1 in the process of carcinogenesis in vivo remains unclear. In the present study, we established SF1-knockout mice using the gene trapping method. Homozygous mice (Sf1(-/-)) died during embryonic development before embryonic day (E)8.5, whereas heterozygous (Sf1(+/-)) mice were born alive and developed normally. Azoxymethane (AOM) was given at a dose of 10 mg/kg body weight once a week for 6 weeks to 7-week-old Sf1(+/-) and Sf1(+/+) mice. At 23 weeks after the start of AOM the average number (5.5 +/- 0.6 versus 2.2 +/- 0.2 in females [P = 0.003, Mann-Whitney U-test], 3.7 +/- 0.2 versus 1.7 +/- 0.7 in males [P = 0.014]) and volume of colon tumors per mouse (8.7 +/- 1.6 versus 2.2 +/- 0.5 mm(3) per female [P = 0.0008], 11.3 +/- 3.4 versus 0.6 +/- 0.2 mm(3) per male [P = 0.001]) were significantly higher in Sf1(+/-) than in Sf1(+/+) mice. The increased susceptibility of Sf1(+/-) mice to AOM-induced colon tumorigenesis indicates the crucial involvement of SF1 in the beta-catenin-mediated regulation of proliferation and differentiation of intestinal epithelial cells.     

C-C chemokine receptor 5 on stromal cells promotes pulmonary metastasis

We have shown that mice that express the C-C chemokine receptor 5 (CCR5) have enhanced local tumor growth and an impaired response to vaccine therapy compared with CCR5 knockout (CCR5(-/-)) mice. Here, we extend these observations to evaluate the function of CCR5 in pulmonary metastasis and the mechanism underlying the diminished tumor growth in CCR5(-/-) mice. Lung metastases were counted in wild-type (WT) and CCR5(-/-) mice following the injection of 1 x 10(6) B16-F10 melanoma cells. These results were compared with those from syngeneic bone marrow chimeric mice formed by the transfer of WT bone marrow into irradiated CCR5(-/-) and CCR5(-/-) marrow into irradiated WT mice. Intact CCR5(-/-) mice developed fewer metastases than WT mice (40.2 versus 70.6; P < 0.05). Bone marrow chimeras formed by the transfer of WT bone marrow into CCR5(-/-) hosts had fewer metastases than WT hosts injected with knockout marrow (46.6 versus 98.6; P < 0.01). Adoptive transfer of CCR5-expressing leukocytes also failed to promote metastasis in CCR5(-/-) mice. However, the i.v. transfer of WT pulmonary stromal cells into CCR5(-/-) mice increased the number of metastases compared with transfer of CCR5(-/-) stromal cells (102.8 versus 26.0; P < 0.05). These results show for the first time that CCR5 expression on stromal and not hematopoietic cells contributes to tumor metastasis. Therefore, recently developed CCR5 inhibitors may have a novel benefit in cancer therapy.     

PIK3CA mutation/PTEN expression status predicts response of colon cancer cells to the epidermal growth factor receptor inhibitor cetuximab

Cetuximab is a monoclonal antibody that targets the human epidermal growth factor receptor (EGFR). Although approved for use in EGFR-overexpressing advanced colorectal cancer, recent studies have shown a lack of association between EGFR overexpression and cetuximab response, requiring the identification of novel biomarkers predictive of response to this agent. To do so, 22 colon cancer cell lines were screened for cetuximab response in vitro and sensitive and resistant lines were identified. In sensitive cell lines, cetuximab induced a G(0)-G(1) arrest without inducing apoptosis. Notably, cetuximab-sensitive but not cetuximab-resistant cell lines were preferentially responsive to EGF-stimulated growth. Whereas neither EGFR protein/mRNA expression nor gene copy number correlated with cetuximab response, examination of the mutation status of signaling components downstream of EGFR showed that cell lines with activating PIK3CA mutations or loss of PTEN expression (PTEN null) were more resistant to cetuximab than PIK3CA wild type (WT)/PTEN-expressing cell lines (14 +/- 5.0% versus 38.5 +/- 6.4% growth inhibition, mean +/- SE; P = 0.008). Consistently, PIK3CA mutant isogenic HCT116 cells showed increased resistance to cetuximab compared with PIK3CA WT controls. Furthermore, cell lines that were PIK3CA mutant/PTEN null and Ras/BRAF mutant were highly resistant to cetuximab compared with those without dual mutations/PTEN loss (10.8 +/- 4.3% versus 38.8 +/- 5.9% growth inhibition, respectively; P = 0.002), indicating that constitutive and simultaneous activation of the Ras and PIK3CA pathways confers maximal resistance to this agent. A priori screening of colon tumors for PTEN expression status and PIK3CA and Ras/BRAF mutation status could help stratify patients likely to benefit from this therapy.     

Mutation accumulation in the intestine and colon of mice deficient in two intracellular glutathione peroxidases

Mice deficient in two glutathione peroxidases (GPX), Gpx1 and Gpx2, [Gpx1/2-double knockout (DKO) mice] are prone to ileocolitis on a mixed C57BL/6 and 129S1/SvJ (B6.129) genetic background. We reported previously that approximately 25% of B6.129 Gpx1/2-DKO mice develop ileocolonic tumors by 6 to 9 months of age, when their non-DKO littermates [having at least one wild-type (WT) Gpx1 or Gpx2 allele] rarely have inflammation and none have tumors. Because genetic background affects tumor susceptibility, we have generated a B6 Gpx1/2-DKO colony and discovered that these mice have fewer inflammatory cells, milder ileocolitis, and low mortality, and only 2.5% of B6 mice developed tumors. The mutant frequency of a cII reporter gene was about 2- to 3-fold higher in 28-day-old Gpx1/2-DKO and 4-fold higher in 8-month-old Gpx1/2-DKO ileal mucosa than in controls in both genetic backgrounds. In contrast, mutant frequencies in the unaffected B6 liver were not significantly different between WT and Gpx1/2-DKO mice. The mutant frequency of 8-month-old B6.129 Gpx1/2-DKO ileum was 38.94 +/- 15.5(-5), which was not significantly higher than the age-matched B6 ileum, 25.54 +/- 10.33(-5). The mutation spectra analysis has shown that B6 Gpx1/2-DKO ileum had a 3-fold increase in small nucleotide deletions at mononucleotide repeats over control B6, which are a signature mutation associated with oxidative stress. Unexpectedly, B6 Gpx1/2-DKO mice had fewer C to T transitions at CpG dinucleotides than the WT B6 (18.0% versus 40.1%; P < 0.001). Our results suggest that inflammation drives gene mutations, which leads to neoplastic transformation of intestinal epithelium in the B6.129 Gpx1/2-DKO mice but rarely in the B6 Gpx1/2-DKO mice.     

Inhibitory effect of cryptoporic acid E, a product from fungus Cryptoporus volvatus, on colon carcinogenesis induced with N-methyl-N-nitrosourea in rats and with 1,2-dimethylhydrazine in mice.

The antitumorigenic effect of cryptoporic acid E (CPA-E), a dimeric drimane sesquiterpenoid isolated from the fungus Cryptoporus volvatus, on colon carcinogenesis was investigated. Female F344 rats given an intrarectal instillation of 2 mg of N-methyl-N-nitrosourea 3 times weekly in weeks 1 and 2 were fed diet containing 0.2% CPA-E from week 3. Female ICR mice given 15 weekly intraperitoneal injections of 10 mg of 1,2-dimethylhydrazine/kg body weight during weeks 1 to 15 were fed diet containing 0.06% CPA-E from week 1. The experiment was terminated at week 35 for rats and at week 25 for mice. The incidence and the number of tumors per animal were reduced in CPA-E-fed animals compared to the controls: 31% vs. 75% (P less than 0.05) and 0.4 +/- 0.2 (SEM) vs. 0.9 +/- 0.2 (0.1 greater than P greater than 0.05) in rats, and 31% vs. 63% (0.1 greater than P greater than 0.05) and 0.4 +/- 0.2 vs. 2.4 +/- 0.8 (P less than 0.05) in mice (16 animals in each group). Intrarectal deoxycholic acid-induced colonic mucosal ornithine decarboxylase activity was significantly lowered in CPA-E-fed animals compared to controls. This shows an antipromoting activity of CPA-E against colon carcinogenesis. Thus, it was concluded that CPA-E inhibits colon cancer development in both rats and mice treated with 2 different colon carcinogens.     

Endostatin对结肠癌生长和转移抑制作用的实验研究

背景与目的:结肠癌的生长、转移是血管生成依赖性的,血管生成抑制剂有望通过抑制肿瘤血管生成,诱导结肠癌细胞凋亡,有效地抑制结直肠癌的生长和转移。肿瘤的抗血管生成治疗对选择手术时机和方式、制定综合治疗方案,提高结肠癌患者5年生存率都具有重要意义。本实验研究血管生成抑制因子Endostatin对结肠癌生长和转移的抑制作用,并探讨其对结肠癌细胞凋亡的影响。方法:采用人结肠腺癌细胞株SW1116完整组织块裸鼠原位种植,建立类似于临床的结肠癌转移裸鼠模型。种植后第1周开始皮下注射Endostatin,每天一次,剂量为0mg/kg(对照组)、5mg/kg、10mg/kg、20mg/kg(治疗组),共用6周。种植后第7周处死动物,测量原位肿瘤瘤重、抑瘤率、肿瘤微血管密度(intratumoralmicrovesseldensity,MVD)、肿瘤细胞凋亡指数(apoptoticindex,AI),观察肿瘤细胞腹膜、肝、其他脏器转移及腹水情况。结果:Endostatin剂量为0mg/kg、5mg/kg、10mg/kg、20mg/kg时,原位肿瘤瘤重分别为(1.31±0.36)g、(0.42±0.17)g、(0.21±0.09)g、(0.13±0.05)g;抑瘤率分别为0%、67.9%、84.0%、90.1%;MVD分别为(12.8±4.1)、(5.9±2.5)、(2.2±1.4)、(0.74±O.3);AI分别为(3.87±2.61)%、(6.89±5.18)%、(13.24±4.76)%、(20.97±9.04)%;腹膜转移率分别为9     

Metastasis stage, adjuvant treatment, and residual tumor are prognostic factors for medulloblastoma in children: conclusions from the Children's Cancer Group 921 randomized phase III study.

PURPOSE: From 1986 to 1992, "eight-drugs-in-one-day" (8-in-1) chemotherapy both before and after radiation therapy (XRT) (54 Gy tumor/36 Gy neuraxis) was compared with vincristine, lomustine (CCNU), and prednisone (VCP) after XRT in children with untreated, high-stage medulloblastoma (MB). PATIENTS AND METHODS: Two hundred three eligible patients with an institutional diagnosis of MB were stratified by local invasion and metastatic stage (Chang T/M) and randomized to therapy. Median time at risk from study entry was 7.0 years. RESULTS: Survival and progression-free survival (PFS) +/- SE at 7 years were 55%+/-5% and 54%+/-5%, respectively. VCP was superior to 8-in-1 chemotherapy, with 5-year PFS rates of 63%+/-5% versus 45%+/-5%, respectively (P = .006). Upon central neuropathology review, 188 patients were confirmed as having MB and were the subjects for analyses of prognostic factors. Children aged 1.5 to younger than 3 years had inferior 5-year estimates of PFS, compared with children 3 years old or older (P = .0014; 32%+/-10% v 58%+/-4%, respectively). For MB patients 3 years of age or older, the prognostic effect of tumor spread (MO v M1 v M2+) on PFS was powerful (P = .0006); 5-year PFS rates were 70%+/-5%, 57%+/-10%, and 40%+/-8%, respectively. PFS distributions at 5 years for patients with M0 tumors with less than 1.5 cm2 of residual tumor, versus > or = 1.5 cm2 of residual tumor by scan, were significantly different (P = .023; 78%+/-6% v 54%+/-11%, respectively). CONCLUSION: VCP plus XRT is a superior adjuvant combination compared with 8-in-1 chemotherapy plus XRT. For patients with M0 tumors, residual tumor bulk (not extent of resection) is a predictor for PFS. Patients with M0 tumors, > or = 3 years with < or = 1.5 cm2 residual tumor, had a 78%+/-6% 5-year PFS rate. Children younger than 3 years old who received a reduced XRT dosage had the lowest survival rate.     

Direct and bystander killing of sarcomas by novel cytosine deaminase fusion gene

Soft tissue and bone sarcomas of the extremities can be difficult to eradicate, and standard treatment may require limb amputation. New therapies to decrease tumor size could improve the effectiveness of treatment and decrease the frequency of limb amputation. Cytosine deaminase (CD)-based gene therapy has been shown to be effective in decreasing growth of solid tumors when animals with CD-expressing tumor cells are treated with 5 fluorocytosine (5FC), an inert prodrug that is converted to 5-fluorouracil (5FU) by CD. In this investigation, we used a novel CD-containing fusion gene to determine whether CD-based gene therapy affected soft tissue or bone sarcomas. The novel fusion gene (NGFR-CD) encodes for a protein with extracellular and transmembrane domains of human nerve growth factor receptor (NGFR) and cytoplasmic CD. Murine 2472 (2) sarcoma cells were transduced with fusion genes containing either the bacterial (NGFR-(b)CD) or yeast (NGFR-(y)CD) CD gene. 5FC treatment killed NGFR-(b)CD- and NGFR-(y)CD-transduced sarcoma cells in vitro through direct and bystander effects (P < 0.01). In contrast, 5FC treatment of mice with s.c. 2NGFR-(b)CD or 2NGFR-(y)CD tumors affected only 2NGFR-(y)CD tumors. 5FC had no effect on growth of NGFR-(b)CD tumors but caused significant decrease in the size of 2NGFR-(y)CD tumors (51 +/- 60 versus 938 +/- 767 mm(3), treated versus control, P < 0.01). Evaluation of bystander killing in vivo revealed significant tumor killing, with a 5-fold reduction in s.c. tumor volume evident in saline versus 5FC-treated mice when tumors were comprised of 90% 2472 cells and 10% 2NGFR-(y)CD selected for fluorescence-activated cell sorting (P < 0.01). Bone sarcomas were eliminated in 9 of 10 5FC-treated mice, compared with 11.8 +/- 6.0 mm(2) in saline-treated mice (P < 0.002). In addition, 5FC treatment of bone sarcomas caused a significant reduction in cancer-induced bone destruction (P < 0.002) and resulted in a reduction in the number of osteoclasts. Finally, 5FC treatment had no effect on animal weight or survival, whereas doses of 5FU providing equivalent tumor reduction as 5FC resulted in treatment-associated deaths and significant weight loss (P < 0.001).     

A fish oil derived concentrate enriched in eicosapentaenoic and docosahexaenoic acid as ethyl ester suppresses the formation and growth of intestinal polyps in the Min mouse

We examined whether the n-3 polyunsaturated fatty acid ethyl ester enriched fish oil K85 (54.4% of eicosapentaenoic acid and 30.3% of docosahexaenoic acid as ethyl esters) could inhibit the intestinal tumorigenesis in Min mice, a murine model of familial adenomatous polyposis (FAP). Min mice that are heterozygous for a nonsense mutation in the Apc gene, develop spontaneously multiple intestinal neoplasms, primarily in the small intestine. K85 was dissolved in corn oil (vehicle) and mixed into the AIN-76A diet. The total oil content (K85 + corn oil) was 12% in all diets. The various experimental diets contained 0 (vehicle control), 0.4, 1.25 or 2.5% of K85. In the small intestine, the mean number of tumors/mouse was 105 +/- 18 (SEM) in control males and 70 +/- 11 in control females. Dietary K85 treatment reduced the number of small intestinal tumors: in males, the maximum reduction was 66% (P = 0.002) with 0.4% of K85; and in females, the maximum reduction was 48% (P = 0.043) with 2.5% of K85, but the inhibition was only slightly increased from 0.4% to 2.5% of K85. The mean tumor diameter was 1.33 +/- 0.08 mm in control males and 1.06 +/- 0.08 in control females, and the diameter ranged from <0.1 mm (monocryptal adenomas) to 4 mm. The small intestinal tumor diameter was reduced by K85 in a dose-dependent manner: in males, with a maximum reduction of 26% (r = -0.64, P = 0.004); and in females, with a maximum reduction of 38% (r = -0.61, P < 0.004). In the large intestine, the mean number of tumors/mouse was 1.0 +/- 0.5 in males and 0.8 +/- 0.2 in females. Although K85 treatment tended to reduce the number and diameter of the large intestinal tumors, these effects did not reach statistical significance. Aberrant crypt foci not elevated from the flat mucosa (ACF(Min)) occurring in the colon of Min mice were also scored. The mean number of ACF(Min)/colon (3.8 +/- 0.9) and the crypt multiplicity (1.49 +/- 0.28) in females were reduced by 73% (P = 0.03) and 60% (P = 0.048) with 2.5% of K85, respectively, whereas no significant effect could be observed in the males.      

Hint1 is a haplo-insufficient tumor suppressor in mice

The HINT1 protein, a member of the histidine triad (HIT) family, is highly conserved in diverse species and ubiquitously expressed in mammalian tissues. However, its precise function in mammalian cells is not known. As a result of its structural similarity to the tumor-suppressor protein FHIT, we used homozygous-deleted Hint1 mice to study its role in tumorigenesis. We discovered that after 2 to 3 years of age the spontaneous tumor incidence in Hint1 -/- mice was significantly greater than that in wild-type Hint1 +/+ mice (P < 0.05). Using a well-established mouse model of 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary carcinogenesis we found a marked and significant (P < 0.05) increase in the incidence of mammary and ovarian tumors in both, Hint1 -/- and +/- mice versus +/+ mice. The Hint1 -/- and +/- mice had similar tumor incidence and similar tumor histologies. Therefore, deletion of Hint1 in mice enhances both spontaneous tumor development and susceptibility to tumor induction by DMBA. In addition, since the Hint1 +/- tumors retained expression of the unmutated wild-type allele, Hint1 is haplo-insufficient with respect to tumor suppression in this model system.