In vivo and in vitro assays of immunocompetence in bronchogenic carcinoma.

To determine the degree of correlation among the various in vivo and in vitro assays that could be used to assess immunocompetence in bronchogenic carcinoma, the response to common recall skin antigens, primary sensitization to DNCB and lymphocyte function tests based on blastogenic response to mitogens and in mixed lymphocyte culture (MLC) were tested in 48 patients with bronchogenic carcinoma. The values were compared to responses in 94 age-matched healthy control subjects. There were impaired skin tests reactions among squamous cell carcinoma patients, with little impairment of their lymphocyte blastogenesis reactions. Anaplastic carcinoma patients had notable defects in lymphocyte function tests but less impairment of the skin test reactions. These data suggest that the mitogen concanavalin A, phytohemagglutinin, and the MLC are more useful screening assays of in vitro immunocompetence than are the other commonly used mitogens.     

In vitro assessment of immunocompetence in patients with malignant melanoma

Ninety-four patients with malignant melanoma and 96 healthy controls were tested for lymphocyte proliferative capacity in a microblastogenesis assay. Each lymphocyte sample was assayed for incorporation of (³H)thymidine after stimulation with PHA, PWM, Con A (two doses), PPD, and allogeneic lymphocytes (MLC). MLC was the only assay that revealed a substantial and significant difference between the melanoma patients and controls. Almost all assays showed lower values in patients with more advanced disease. However, it was not possible to accurately predict clinical outcome from data obtained from in vitro immunocompetence tests. These results indicate the relatively limited clinical usefulness of assays of lymphocyte proliferative capacity in melanoma patients.     

Unaltered immunocompetence in patients with non-disseminated breast cancer at the time of diagnosis

Immunologic parameters were examined preoperatively in 104 patients with breast cancer, staged according to the TNM classification and in 95 age-matched healthy women. The immunologic evaluation in the peripheral blood included lymphocyte and monocyte counts, determination of E-rosette-forming T-lymphocytes (SER+) and B-lymphocytes (MER+), T-lymphocyte subsets defined with monoclonal antibodies (Leu-1, Leu-2a, Leu-3a) and with lectin fractionation (soybean agglutinin), lymphocyte transformation test with phytohemagglutinin (PHA) and concanavalin A (ConA), and colony formation of T-lymphocytes in agar (T-lymphocyte colony-forming cells, [TL-CFC]). Two age groups (Group A: 30-50 years; Group B: 51-70 years) and the different tumor stages (Stage I-IV) were analyzed. Patients and controls did not differ in the absolute numbers of lymphocytes, T- and B-cells. In patients of Group B, the absolute number of monocytes was increased slightly in Stage II and III and significantly in Stage IV (P less than 0.05). Similarly, the lymphocyte response to PHA was significantly reduced in Stage IV Group B only (P less than 0.05). ConA-induced lymphocyte proliferation and TL-CFC capacity were not different in patients and controls. In the small number of patients and age-matched controls in whom T-lymphocyte subsets were determined, the absolute numbers of T-cells with helper or suppressor phenotype as defined with Leu-3a, Leu-2a, or lectin fractionation with soybean agglutinin were similar. This study demonstrates that in patients with early breast cancer (Stage I-III), immunocompetence as defined by either functional in vitro studies or surface marker analysis is not significantly altered at the time of diagnosis. In contrast, patients with advanced disease (Stage IV) show a significant increase in the absolute number of monocytes and a depressed PHA responsiveness of mononuclear cells.     

Correlation of in vitro cytotoxicity with preclinical in vivo antitumor activity

Several human and murine tumor cell lines were evaluated in an in vitro cytotoxicity assay as prescreens for fermentation extracts and pure materials subsequently tested in vivo against P388 leukemia or B16 melanoma. Each material, regardless of its in vitro cytotoxicity, was evaluated in vivo. At the criteria levels of in vitro positivity and in vivo activity invoked, a highly significant relationship between these two endpoints was demonstrated for each cell line. When cell lines were compared, most of them performed in a similar manner, with HCT-116 human colon carcinoma cells providing a modest advantage predicting for P388 activity in some comparisons. Using the data from any two cell lines in concert did not improve the acuity of the prescreen beyond that associated with the better cell lines used singularly and only a minority of active materials was predicted for uniquely. Overall, the in vitro cytotoxicity assay provided a useful prescreen for selecting P388 and B16 in vivo active materials.     

PC-SPES inhibits colon cancer growth in vitro and in vivo

PC-SPES is a mixture of eight herbs with antiproliferative activity in prostate cancer cell lines and antitumor effects in animal models of prostate cancer. In addition, evidence of clinical efficacy in advanced prostate cancer has been reported. PC-SPES has also been shown to have antitumor activity against several other cancer cell lines including breast and neuroepithelial cancer, melanoma, and leukemia cell lines. Because of these findings, we investigated the effects of PC-SPES in vitro in colon cancer cell lines SW480, SW620, and DLD-1 and in vivo in the Apc(min) mouse, a murine model for intestinal carcinogenesis. For the in vitro studies, colon cancer cell lines were exposed to an ethanolic extract of PC-SPES compared with a diluent control [ethanol < or = 0.3% (v/v)]. PC-SPES resulted in a marked suppression of cell proliferation in all colon cancer cells studied. PC-SPES (3 micro l/ml) caused a 95% inhibition of cell proliferation of the DLD-1 colon cancer cell line, and similar results were observed in the SW480 and SW620 colon cancer cell lines. Cell cycle analysis demonstrated a drastic (> or =60%) accumulation of cells in the G(2)-M phase with a concomitant decrease of cells in the G(0)-G(1) phase in all colon cancer cell lines studied after treatment with PC-SPES (1.5 micro l/ml for 48 h). Western blot analysis demonstrated a decrease in protein levels of beta-tubulin in the SW620 cell line exposed to PC-SPES. Terminal deoxynucleotidyl transferase-mediated nick end labeling analysis revealed an increase in apoptotic colon cancer cells incubated with PC-SPES. For the in vivo studies, female 4-5-week-old Apc(min) mice were randomized to two groups: a PC-SPES-treated group (n = 11) received 250 mg/kg/day (0.2 ml) PC-SPES via gastrointestinal gavage; and a control group (n = 10) received 0.2 ml of the vehicle solution (1.5% carboxymethylcellulose with 0.2% Tween 20) via gastrointestinal gavage. Both groups were treated five times a week for 10 weeks. After treatment, the gastrointestinal tract was dissected for polyp scoring by two observers blinded to treatment. The Apc(min) mice given PC-SPES had a 58% reduction in tumor number and a 56% decrease in tumor load. No effect on either food intake or body weight was observed in the treated versus sham groups. The present study is the first to report the potent activity of PC-SPES against colon cancer. Both cell cycle arrest and apoptosis occurred after treatment with PC-SPES. This suggests that the components of this herbal mixture, either independently or in combination, acted in colon cancer, resulting in a drastic effect on tumor initiation and tumor progression.     

Effect of adriamycin and high-dose methotrexate chemotherapy on in vivo and in vitro cell-mediated immunity in cancer patients

Adjuvant chemotherapy with adriamycin (ADR) and high-dose methotrexate (HDM) with citrovorum rescue appears to hold promise for preventing recurrence of sarcomas following surgery. However, the effect of prolonged treatment with these agents on both in vivo and in vitro cell-mediated immunity (CMI) is unknown. To assess in vitro CMI, cryopreserved lymphocytes from 18 patients, collected before and at monthly intervals following initiation of biweekly ADR and HDM therapy, were tested simultaneously for stimulation to phytohemagglutinin, poke-weed mitogen, Concanavalin A, and pooled allogeneic lymphocytes by a micro-test technique. In vivo CMI was determined by serial skin-testing with 2,4,-dinitrochlorobenzene (DNCB). No significant overall change was noted for any of the in vitro lymphocyte function tests. DNCB skin test ratings, peripheral leukocyte counts and absolute lymphocyte counts did not change significantly during the study. Lymphocytes from a second group of patients were tested before, at 24 and 48 hours after HDM therapy. In vitro lymphocyte function tests were significantly depressed 24 hours following treatment, but returned to pretreatment levels at 48 hours. Long-term adjuvant ADR and HDM therapy does not appear to alter in vivo and in vitro CMI, although transient depressions were noted. The preservation of intact CMI may account in part, for the effectiveness of these agents as surgical adjuvants.     

In vivo experiments, in vitro assays in experimental oncology: establishment of LA 795 Vv-Vt

In vivo experiment, in vitro assays (in vivo-in vitro system) is a kind of experimental technique which is different from both in vitro cell line and tumor line. The new model can be grown and studied either as an animal tumor or a cell culture. The specimen could be assayed for cell survival in vitro. This model can be used to study the response of malignant cells to the treatment in a precision and depth that is impossible in tumors grown in vivo. LA 795 Vv-Vt is the first in vivo-in vitro system developed in China. It was established with lung adenocarcinoma of T-739 mouse. The tumor cells could grow freely both in vivo and in vitro with a plating efficiency of 20%. Characteristics of LA 795 Vv-Vt tumor and cell in culture were described and compared. In order to estimate the radiation response of different doses, cell survival curves of tumors irradiated under oxic and hypoxic conditions were drawn and compared. The oxygen enhance ratio was 2.98. The experiments indicate that in vivo-in vitro system has a good dependent relation in dose and effect and is worth extensive application.     

8-Cl-adenosine-induced inhibition of colorectal cancer growth in vitro and in vivo

The cAMP analogue 8-Cl-cAMP induces apoptosis and inhibits proliferation of a wide variety of malignancies in vitro and in vivo with relatively little toxicity. The antitumor effects of this compound are thought to involve its ability to modulate type I protein kinase A (PKAI). However, a nontoxic metabolite of 8-Cl-cAMP, 8-Cl-adenosine, with no known activity against PKAI, exerts growth inhibitory effects in breast, ovary, pancreas, and colorectal cancer cells in vitro and accumulates in xenografted tumors after 8-Cl-cAMP treatment in vivo. To characterize further the antitumor effects of 8-Cl-adenosine in colorectal cancer, we examined its effects on cell growth in vitro (cell number, ³H-thymidine incorporation, and soft agar colony formation) using the isogenically matched colorectal cancer cell lines HCT116, HCT116-E6 (p53-depleted), and 80S14 (p21^WAF1/Cip1-null). 8-Cl-adenosine inhibited cell growth by 89%, 74%, and 79%, respectively in HCT116, HCT116-E6, and 80S14 cells after a 72-hour exposure. Growth inhibition coincided with DNA endoreduplication and subsequent apoptosis. Furthermore, nontoxic doses of 8-Cl-adenosine administered i.p. twice weekly for 4 weeks to athymic mice suppressed growth of HCT116-derived xenografts by 50%. These results show that 8-Cl-adenosine exerts antitumor activity against colorectal cancer independent of p53 and p21^WAF1/Cip1.     

天花粉蛋白抑制乳腺癌生长的实验研究

目的 探讨天花粉蛋白(TCS)对MDA-MB-231和MCF-7乳腺癌细胞及乳腺癌裸鼠移植瘤的影响.方法 四唑盐比色法(MTT)检测TCS处理前后细胞的增殖情况;流式细胞术(FCM)检测TCS处理前后细胞周期变化和凋亡率;将MDA-MB-231细胞接种于裸鼠,成瘤后分为对照组、TCS组和盐水组.检测各组裸鼠移植瘤体积、瘤重和肝、肾功能、血常规变化.结果 随着TCS作用时间延长、浓度增大、MDA-MB-231和MCF-7细胞生长受到明显抑制;TCS处理后细胞阻滞于G0/G1期,并可检测到凋亡峰;TCS组裸鼠移植瘤体积和瘤重较对照组明显缩小,TCS对荷瘤裸鼠肾功能、血常规无明显影响,可引起谷丙转氨酶轻度升高.结论 TCS可抑制MDA-MB-231和MCF-7乳腺癌细胞增殖和乳腺癌裸鼠移植瘤生长.     

Association of carmustine with a lipid emulsion: in vitro,in vivo and preliminary studies in cancer patients

PURPOSE: We had previously shown in acute leukemia and in breast and ovary carcinoma patients that a cholesterol-rich emulsion (LDE) that binds to receptors for low-density lipoprotein (LDL) may concentrate in neoplastic tissues. In this study, the potential of LDE as a carrier for anticancer drugs was investigated. METHODS: LDE was associated with carmustine, and the cytotoxicity of the LDE-carmustine complex was studied in a neoplastic cell line and its biodistribution was studied in mice. The plasma kinetics of the complex and its uptake by tumor and normal tissue were determined in cancer patients. Finally, an exploratory clinical study to determine the toxicity profile of LDE-carmustine at escalating dose levels was conducted in 42 advanced cancer patients refractory to conventional chemotherapy. RESULTS: Carmustine formed a stable association with LDE. The pharmacological action of carmustine, as tested in cancer cells, was not diminished by association with LDE compared with the free drug and was indeed mediated by the LDL receptor. The biodistribution in mice and plasma kinetics in patients of the emulsion were not changed by association of the drug. The uptake of LDE-carmustine by tumor was severalfold greater than the uptake by the corresponding normal tissue. Finally, patients treated with LDE-carmustine showed negligible side effects even at very high dose levels. CONCLUSIONS: Association with LDE preserves the cytotoxicity of carmustine and markedly diminishes its side effects.     

NRAGE suppresses metastasis of melanoma and pancreatic cancer in vitro and in vivo

We previously reported that human NRAGE could significantly alter the cellular skeleton and inhibit cell–cell adhesion, suggesting that human NRGAE play a potential role in cellular motility. Here, we report overexpression of human NRAGE in PANC-1 and B16-Bl6 cells could significantly suppress the metastasis of these cells in vitro and in vivo. Consistently, PANC-1 with stable silencing of NRAGE by RNA interference, exhibits a more metastatic phenotype than the native cell. Expression of epithelial proteins, including E-cadherin and β-catenin is down regulated in siRNA-NRAGE PANC-1 cells. Further studies find that overexpression of human NRAGE suppresses the mRNA expression and activity of MMP2 significantly. Summary, our studies indicate for the first time that NRAGE could suppress metastasis of melanoma and pancreatic cancer probably through downregulation of MMP-2.     

Correlation of in vivo malignancy with in vitro properties of human-mouse hybrid cells.

The in vivo tumorigenicity of malignant mouse-nonmalignant human somatic cell hybrids was correlated with the in vitro characteristics. The renal adenocarcinoma mouse cell line RAG and the normal, diploid human cell line Wl-38 were used as the fusion parents. Five independent RAG Wl 38 hybrid clones were tested for concanavalin A (Con A) agglutination patterns, in vitro invasiveness, and tumor formation in immunosuppressed mice. Tests on the parental lines showed that RAG cells agglutinated at much lower levels of Con A than the Wl-38 cells. RAG cells overgrew Wl-38 cells in the in vitro invasiveness assay. RAG cells readily formed tumors in untreated adult or young immunosuppressed mice, whereas the Wl-38 cells did not. The five hybrid clones were all similar, since they had Con A agglutination levels intermediate to those of both parents, though patterns were more "tumor-like", overgrew the Wl-38 cells in the invasiveness assay, and formed tumors in immunosuppressed mice.     

Correlation between physical performance and fatigue in cancer patients

Background: Fatigue and a reduction in physical ability are common andoften severe problems of cancer patients regardless of disease stage andmodality of treatment. However, while physical performance can be assessedobjectively with laboratory tests, fatigue is a subjective phenomenon whoseperception is influenced by past experience and expectations for the future.Patients and methods: To evaluate the correlation between fatigue andphysical impairment, we assessed maximal physical performance with atreadmill test, and mental state with two questionnaires, the Profile ofMood States (POMS) and the Symptom Check List (SCL-90-R), in a successiveseries of 78 cancer patients with solid tumors or hematologicalmalignancies.Results: A weak association between fatigue and maximal physicalperformance was found (r = –0.30; P < 0.01). However, intensity offatigue showed a strong correlation with several indicators of psychologicaldistress such as depression (r = 0.68), somatization (r = 0.64) and anxiety(r = 0.63; P for all <0.001). Furthermore, patients with lower levels ofphysical performance had significantly higher scores for depression (P =0.005), somatization (P = 0.03) and anxiety (P = 0.08), and significantlylower scores for vigor (P = 0.05) than their counterparts whose physicalperformance was higher.Conclusions: We conclude that fatigue in cancer patients may be related tomood disturbance but appears to be independent of physical performance.Moreover, low physical performance can be viewed as an independent predictorof mental distress in cancer patients.     

Somatostatin in medullary thyroid cancer. In vitro and in vivo studies

The authors evaluated the presence of somatostatin (SRIF) in the plasma and in the tumor tissue of a total of 22 patients with medullary thyroid cancer (MTC) and studied the effect of exogenous SRIF administration on basal and pentagastrin (PG)-stimulated plasma calcitonin (CT) and carcinoembryonic antigen (CEA) levels. Mean plasma SRIF concentrations were significantly higher than those found in normal controls, with five of 15 patients having plasma SRIF levels above the mean + 2 SD of normal controls. High immunoreactive SRIF concentrations were found in the extract of three tumor tissues but not in one follicular thyroid cancer or in one toxic diffuse goiter. By immunoperoxidase staining seven of 11 (63.6%) primary MTC and five of 13 (38.5%) metastases expressed SRIF antigen in a low number of cells and with a weak degree of staining. As expected, CT was expressed in almost 100% of the cases with positivity in most of the cells and strong degree of staining. Patients with positive SRIF staining in the primary tumor had longer survival than SRIF negative patients. Infusion of synthetic SRIF (11 micrograms/minute/45 minutes) produced a significant reduction of plasma CT (but not CEA) levels in 12 of the 15 patients submitted to this test. Maximal percent decrease of plasma CT ranged from 10.8% to 72.7% of the basal value and was usually observed between 30 and 45 minutes from the beginning of the infusion. When infused together with the injection of PG, SRIF was able to significantly (P less than 0.05) inhibit the PG-induced CT release in five of six patients tested. These results demonstrate the following: SRIF is present in a few cells of many primary MTC and less frequently in their metastases; tentatively, the expression of SRIF antigen in the tumor seems to be associated with longer survival; increased SRIF concentrations are found in the plasma of some patients with metastatic involvement; and treatment with exogenous SRIF reduces the basal and PG-induced CT (but not CEA) release from the tumor.     

Cytokine production of lung cancer cell lines: Correlation between their production and the inflammatory/immunological responses both in vivo and in vitro

Cytokines produced by tumor cells may have various effects on antitumor immune responses and tumor growth. In the present study, the cytokine production of 31 lung cancer cell lines was evaluated, while any correlation with the histological type, the induction of tumor-specific cytotoxic T lymphocytes (CTL) in vitro, and angiogenesis and the infiltration of inflammatory cells in tumor tissues were also examined. Production of interleukin (IL)-1alpha, IL-1beta, IL-4, IL-6, IL-8, IL-10, tumor necrosis factor (TNF)-alpha, granulocyte macrophage colony stimulating factor (GM-CSF), granulocyte colony stimulating factor, transforming growth factor (TGF)-beta and vascular endothelial growth factor (VEGF) in the culture supernatant was measured using enzyme-linked immunosorbent assay. Each cytokine was produced in a substantial number of the tumor cell lines. In particular, IL-6, IL-8, TGF-beta and VEGF were produced in 18 (55%), 29 (94%), 31 (100%) and 28 (90%) of 31 cell lines, respectively. However, neither IL-4 nor TNF-alpha was produced at all by any tumor cell line. TGF-beta production was significantly higher in adenocarcinoma than in squamous cell carcinoma (P = 0.03). Immunohistochemical staining revealed the magnitude of macrophage infiltration, and angiogenesis in surgically resected tumor tissue specimens correlated well with GM-CSF and IL-8 production from the corresponding cell lines. Among six lung cancer cell lines, CTL were induced in the three lung cancer cell lines that produced a lower amount of TGF-beta (<100 pg/mL). These findings suggested that TGF-beta produced by tumor cells could inhibit the induction of CTL in vitro. The present results suggest that the production of various cytokines from tumor cells might exert various paracrine effects both in vivo and in vitro.     

Cardamonin reduces chemotherapy-enriched breast cancer stem-like cells in vitro and in vivo

The failure of cytotoxic chemotherapy in breast cancers has been closely associated with the presence of drug resistant cancer stem cells (CSCs). Thus, screening for small molecules that selectively inhibit growth of CSCs may offer great promise for cancer control, particularly in combination with chemotherapy. In this report, we provide the first demonstration that cardamonin, a small molecule, selectively inhibits breast CSCs that have been enriched by chemotherapeutic drugs. In addition, cardamonin also sufficiently prevents the enrichment of CSCs when simultaneously used with chemotherapeutic drugs. Specifically, cardamonin effectively abolishes chemotherapeutic drug-induced up-regulation of IL-6, IL-8 and MCP-1 and activation of NF-κB/IKBα and Stat3. Furthermore, in a xenograft mouse model, co-administration of cardamonin and the chemotherapeutic drug doxorubicin significantly retards tumor growth and simultaneously decreases CSC pools in vivo. Since cardamonin has been found in some herbs, this work suggests a potential new approach for the effective treatment of breast CSCs by administration of cardamonin either concurrent with or after chemotherapeutic drugs.