131I-anticarcinoembryonic antigen therapy of LS174T human colon adenocarcinoma spheroids

LS174T human colon adenocarcinoma multicell spheroids were used to study the radiobiological aspects of radioimmunotherapy. The spheroids were incubated in 131I-anticarcinoembryonic antigen (B7) at an antibody concentration of 0.5 microgram/ml and at 131I concentrations of 2.5 and 7.5 microCi/ml. After incubation times of 90 h, clonogenic cells per spheroid were reduced by 1400-fold and 23-fold at the high and low 131I concentrations, respectively. 131I Nonspecific antibody (PX63) resulted in 2- and 1.2-fold reductions. Spheroid diameter was not significantly affected by therapy but histological examination revealed that there had been a significant reduction in the cell density, particularly near the spheroid surface. Using a theoretical model to estimate radiation dose, a radiation survival curve was constructed. The resulting curve was somewhat concave suggesting the presence of a resistant population of cells. It is likely that this observation is primarily due to the fact that the inner cells received a lower dose than the outer cells. A population of radiobiologically hypoxic cells in the inner portion of the spheroids may also have contributed to the decreasing slope of the curve as well as ongoing cell division leading to new cells which receive a lower radiation dose per cell cycle. Because of the ability to estimate radiation dose for a given biological effect, these types of experiments may allow predictions of the efficacy of radiolabeled antibody therapy for micrometastatic disease.     

Multifactorial resistance in LS174T human colon carcinoma cells selected with doxorubicin.

A series of doxorubicin-resistant variants of the human LS174T colon carcinoma cell line was generated by stepwise selection. These variants also exhibited increased resistance to vinblastine, etoposide, cis-platinum, and melphalan, suggesting that resistance was multifactorial. The parental LS174T cell line and 3 resistant variants were examined for over-expression of P-glycoprotein, changes in total cellular glutathione content, and the level of topoisomerase-II expression. Changes in all of these parameters were observed in the doxorubicin-selectants, along with a marked shift in the intracellular distribution of doxorubicin. P-glycoprotein RNA and protein levels were increased 2- to 3-fold in the resistant variants, while total glutathione levels increased 1.4- to 2.1-fold. Treatment with DL-buthionine-[S,R]-sulfoximine, an inhibitor of glutathione biosynthesis, was able to reverse resistance to cis-platinum and melphalan in these variants, but had little effect on doxorubicin resistance. Immunoblot analysis of cell extracts indicated that the level of DNA topoisomerase II (EC 5.99.1.3) in the doxorubicin-resistant LS174T cells was decreased by approximately 50% compared with the parental cell line. Doxorubicin was mainly localized to the cytoplasm in resistant cells, while in the parent line it was mostly found in the nucleus. This constellation of changes suggests that selection with doxorubicin activated several mechanisms of resistance involving drug transport, metabolism, and ability to reach nuclear target sites.     

Proton relaxation times and interstitial fluid pressure in human melanoma xenografts.

The interstitial fluid pressure (IFP) and the proton spin-lattice and spin-spin relaxation times (T1 and T2) of some experimental tumours have been shown to be related to tumour water content. These observations have led to the hypothesis that magnetic resonance imaging (MRI) might be a clinically useful non-invasive method for assessment of tumour IFP. The purpose of the work reported here was to examine the general validity of this hypothesis. R-18 human melanoma xenografts grown intradermally in Balb/c nu/nu mice were used as the tumour model system. Median T1 and T2 were determined by spin-echo MRI using a 1.5-T clinical whole-body tomograph. IFP was measured using the wick-in-needle technique. No correlation was found between tumour IFP and fractional tumour water content. Moreover, there was no correlation between median T1 or T2 and IFP, suggesting that proton T1 and T2 values determined by MRI cannot be used clinically to assess tumour IFP and thereby to predict the uptake of macromolecular therapeutic agents.     

多西环素抑制大肠癌细胞侵袭的实验研究

目的观察多西环素（Doxycycline）对人大肠癌LS174T细胞体外侵袭的作用。方法以人工重组基底膜（Matrigel）体外侵袭实验观察药物对细胞体外侵袭能力的影响;SDS-聚丙烯酰胺凝胶电泳法（明胶酶谱法）观察对金属蛋白酶分泌的影响。结果多西环素抑制LS174T大肠癌细胞体外侵袭,在多西环素浓度为5mg／L,10mg／L,20mg／L时,每100倍视野穿过的细胞数分别为（101．3±6．7）个,（83．7±5．7）个和（42．0±2．0）个,与对照组有显著差异,抑制率分别为17．9％、32．0％和65．9％,呈现剂量依赖性;在5mg／L的低浓度下多西环素即可抑制基质金属蛋白酶-2（MMP-2）的分泌。结论多西环素能够抑制人大肠癌LS174T细胞的侵袭,在低于抗菌浓度下即可抑制基质金属蛋白酶-2的分泌,具有成为抗肿瘤侵袭药物的潜力。     

Biodistribution, pharmacokinetic, and imaging studies with 186Re-labeled NR-LU-10 whole antibody in LS174T colonic tumor-bearing mice

Biodistribution, pharmacokinetic, and radioimaging studies were performed with 186Re-labeled NR-LU-10 whole antibody in athymic nude mice bearing the LS174T tumor growing either s.c. or in an experimental hepatic metastasis model. NR-LU-10 is an IgG2b murine monoclonal antibody (MAb) that reacts with virtually all human tumors of epithelial origin. NR-BC-1, a IgG2b murine MAb that reacts with normal human B-cell and B malignancies, was used as an isotype-matched control. These MAbs were radiolabeled with 186Re (3.7-day physical half-life; 1.07-MeV beta particle and 137-keV gamma, 9% abundance) by a preformed chelate approach by using the triamide thiolate ligand system. 186Re-labeled NR-LU-10 (50 microCi) was injected into nude mice bearing LS174T tumors growing s.c. Biodistribution studies revealed that the LS174T tumor retained the highest concentration of 186Re-labeled NR-LU-10 (5.3% injected dose/g) at day 6. The tumor:blood ratio ranged from 0.1:1 to 10.8:1 by day 6, the last day of analysis. In contrast the tumor:blood ratio of 186Re-labeled NR-BC-1, the isotype-matched MAb control, was 1:1 on day 6. Pharmacokinetic analysis indicated that the t1/2 beta of NR-LU-10 for blood and other tissues ranged from 21 to 25 h, while the t1/2 beta for the LS174T tumor averaged 52 h. The area under the curve for tumor compared to blood was 2.8- to 5.7-fold higher than the area under the curve for all other tissues and organs. The mean residence time for NR-LU-10 in blood and all other organs ranged from 23 to 26 h, while the mean residence time for NR-LU-10 in the LS174T tumor was 72 h. Scintigraphic images revealed selective uptake of the 186Re-labeled NR-LU-10, but not of the 186Re-labeled NR-BC-1, at the LS174T tumor site. Studies in an experimental model of hepatic metastasis revealed a similar selective pattern of 186Re-labeled NR-LU-10 accumulation. Scintigraphic images of the LS174T tumor growing within the athymic nude mouse liver were obtained. The biodistribution, pharmacokinetic, and scintigraphic image results suggest that 186Re-labeled NR-LU-10 shows promise as a therapeutic agent for gastrointestinal cancer.     

Growth inhibition, enhancement of intercellular adhesion, and increased expression of carcinoembryonic antigen by overexpression of phosphoinositides-specific phospholipase C beta 1 in LS174T human colon adenocarcinoma cell line.

By using a retrovirus-derived system we generated derivatives of the human colon adenocarcinoma cell line LS174T (ATCC CL 188) that stably overexpress a full-length cDNA encoding the beta 1 isoform of bovine phosphoinositides-specific phospholipase C (PI-PLC). This was confirmed by the elevated levels of catalytic activity to release phosphoinositides from phosphatidylinositol (PI-PLC) or phosphatidylinositol-bis-phosphate (PIP2-PLC), and the enhanced expressions of messenger RNA and protein. PI-PLC beta 1 overexpresser clones grew to form cell clumps floating in liquid medium, whereas the pMV7-introduced control clones displayed morphologic characteristics that were very similar to those of the parent LS174T cell line. Three individual PI-PLC beta 1 overexpresser cell lines displayed increased doubling time (18.0 h, 21.5 h, and 23.8 h) when compared with 4 individual pMV7-introduced control cell lines (13.1 h, 10.7 h, 12.9 h, and 9.3 h). Anchorage-independent growth ability in soft agar medium was dramatically suppressed by overexpression of PLC beta 1, and the ability of PLC-overproducer clones to form aggregates when cultured in liquid medium was dramatically enhanced when compared with that of pMV7-introduced control clones. Tumorigenicity of PLC beta 1-overproducers was much weaker than that of vector-transduced control clones. The spontaneous release of carcinoembryonic antigen from PLC beta 1-overproducer clones was much higher than that from pMV7 control clones. The ability of PLC beta 1-overproducer clones to form aggregates during suspension culture was much stronger than that of the control clones. These results provide the first evidence that elevated levels of endogenous PI-PLC beta 1 suppress tumor cell growth, but enhance the ability to form cell aggregates and to release carcinoembryonic antigen, an intercellular adhesion molecule.     

Tributyrin-induced differentiation promotes apoptosis of LS 174T colon cancer cells in vitro.

Tributyrin (glyceryl tributyrate, TB) is known to induce malignant cells to differentiate followed by arrest of cell growth and death via apoptosis. We investigated the effects of TB on the distribution of cell cycle phases, differentiation as measured by alkaline phosphatase activity (ALP), and apoptosis of LS 174T colon cancer cells expressed by morphological changes, externalization of phosphatidylserine and stimulation of various caspases. TB (0.6 mM) reduced the proliferation by a 5-fold decrease of tumor cells in the S-phase and 1.3-fold increase in the G2/M-phase of cell cycle after 24 h of incubation. The ALP activity was enhanced in a dose-dependent manner up to 180-fold by 1 mM TB. Apoptosis was seen only above 0.6 mM TB (5-fold increase). Studies with caspase inhibitors revealed that TB mediated cell death was linked to up-regulation of caspases 3 and 8. Our results indicate that TB-induced differentiation promotes apoptosis in LS 174T cells and may explain the mode of action of TB finally resulting in an arrest of tumor cell growth.     

Effect of human natural killer and gammadelta T cells on the growth of human autologous melanoma xenografts in SCID mice

Natural killer (NK) cells were first identified for their ability to kill tumor cells of different origin in vitro. Similarly, gammadelta T lymphocytes display strong cytotoxic activity against various tumor cell lines. However, the ability of both the NK and gammadelta cells to mediate natural immune response against human malignant tumors in vivo is still poorly defined. Severe combined immunodeficient (SCID) mice have been successfully engrafted with human tumors. In this study, the antitumor effect of local as well as of systemic treatments based on NK cells or Vdelta1 or Vdelta2 gamma/delta T lymphocytes against autologous melanoma cells was investigated in vivo. The results show that all three of the populations were effective in preventing growth of autologous human melanomas when both tumor and lymphoid cells were s.c. inoculated at the same site. However, when lymphoid cells were infused i.v., only NK cells and Vdelta1 gamma/delta T lymphocytes could either prevent or inhibit the s.c. growth of autologous melanoma. Accordingly, both NK cells and Vdelta1 gammadelta T lymphocytes could be detected at the s.c. tumor site. In contrast, Vdelta2 gammadelta T lymphocytes were only detectable in the spleen of the SCID mice. Moreover, NK cells maintained their inhibitory effect on tumor growth even after discontinuation of the treatment. Indeed they were present at the tumor site for a longer period. These data support the possibility to exploit NK cells and Vdelta1 gammadelta T lymphocytes in tumor immunotherapy. Moreover, our study emphasizes the usefulness of human tumor/SCID mouse models for preclinical evaluation of immunotherapy protocols against human tumors.     

Radiobiological hypoxia, oxygen tension, interstitial fluid pressure and relative viable tumour area in two human squamous cell carcinomas in nude mice during fractionated radiotherapy

Very little is known about the correlation between the radiobiological hypoxic fraction (rHF) and other measures of tumour oxygenation during fractionated irradiation. In the present study the rHF is determined in untreated human FaDu and GL squamous cell carcinoma in nude mice and in tumours irradiated with 10 fractions in 2 weeks and 20 fractions in 4 weeks, using tumour control as the experimental endpoint. The results were compared with measurements of the pO2, the interstitial fluid pressure (IFP) and the relative viable tumour area. In FaDu tumours the radiobiological hypoxic fractions (rHFs) before and during irradiation were not statistically different from 100%. Depending on the assumptions made for D0, the rHFs of GL tumours were between 0.2 and 4% or 30 and 53%. The median pO2 values were 2.8 mmHg for untreated FaDu tumours and 0.2 mmHg for GL tumours (p < 0.001). The median IFP values were 2.6 mmHg in FaDu and 5.3 mmHg in GL tumours (p = 0.01). No important changes in the pO2 and IFP values were observed during fractionated irradiation. The relative viable tumour area during irradiation decreased by 83% in FaDu tumours (p = 0.002) and by 54% in GL tumours (p = 0.003). It is concluded that differences in rHF exist between FaDu and GL tumours before and during fractionated irradiation and that these differences are not reflected by pO2 and IFP values and the relative viable tumour area.     

Comparison of metabolically labeled mucins of LS174T human colon cancer cells in tissue culture and xenograft

Colon cancer cells in culture synthesize and secrete mucin glycoproteins, which carry a number of cancer-associated antigens. However, the structures and mechanisms of biosynthetic processing are not well understood. Mucins synthesized and secreted by LS174T human colon cancer cells were compared to those in LS174T xenografts in athymic mice. Mucins radiolabeled with glucosamine or sulfate were purified by gel filtration and cesium chloride density gradient centrifugation. The mucins were of high molecular weight and were resistant to chondroitinase ABC, hyaluronidase and HNO2 treatment. They were, however, susceptible to pronase digestion and mild alkaline treatment. Using radiochemical precursors, the cellular mucin was shown to contain fucose, galactose, N-acetylgalactosamine, N-acetylglucosamine, N-acetylneuraminic acid, and sulfate. Oligosaccharides released by beta-elimination had N-acetylgalactosaminitol as the reduced amino sugar and also unreduced galactosamine, indicating that there is N-acetyl-galactosamine O-glycosidically attached to protein core and also peripheral N-acetyl-galactosamine not directly linked to protein. DEAE-cellulose chromatography of mucins showed two major peaks with both intracellular and secreted mucins, but xenograft mucins also had more acidic components. Sulfate-labeled mucins were shifted to less acidic peaks by neuraminidase digestion, which indicates that the same mucin molecules are both sialylated and sulfated. We conclude that the intracellular mucins of cultured colon cancer cells, those secreted into the medium, and those in nude mouse xenografts are chemically similar, but differ in sialic acid and sulfate content. This experimental model system, LS174T cells maintained in culture and as nude mouse xenografts, may be useful for further biosynthetic and structural studies of colon cancer mucin.     

右旋柠烯对人结肠癌LS174T细胞p-ERK和p-Akts表达的影响

目的研究右旋柠烯(D-limonene)对人结肠癌细胞(LS174T)中信号传导分子ERK1/2、Akt及其磷酸化蛋白(p-ERK和p-Akts蛋白)表达的影响。方法 采用四甲基偶氮唑盐(MTT)方法检测右旋柠烯对LS174T细胞增殖的抑制作用;采用Western blot方法检测不同浓度右旋柠烯对LS174T细胞中ERK1/2、Akt及其磷酸化蛋白表达情况的影响。结果 右旋柠烯作用LS174T细胞48h后,呈剂量依赖性抑制细胞增殖;Western blot结果显示右旋柠烯对细胞中ERK1/2、Akt蛋白的表达没有明显影响,但显著下调p-ERK1/2蛋白和p-Akts的表达。结论 右旋柠烯抑制LS174T细胞增殖可能与其抑制p-ERK1/2和p-Akts表达有关。     

Growth of human lymphoma cells in SCID mice.

Two EBV-negative human lymphoma cell lines raised in this laboratory and peripheral blood cells of a patient with large cell lymphoma in leukemic phase were injected intravenously or intraperitoneally into C.B.17 SCID mice. One line (OCI-LY18) was derived from the pleural fluid of a patient with a large cell, immunoblastic malignant lymphoma. Cells of this line are of B cell origin and characterized by multiple rearrangements of the JH locus. The second line (OCI-LY17) was grown from the peripheral blood of a patient with a large cell lymphoma of T-cell phenotype which has characteristic rearrangement of the T-cell receptor beta chain. The cells directly obtained from a patient with large cell non-cleaved malignant lymphoma were of B-cell origin. Animals carrying OCI-LY18 developed large tumor masses within 6-8 weeks of inoculation. The tumors were detected in the intestine, mesentery, retroperitoneum, lymph nodes, spleen, lung and kidney. The masses resembled the primary tumor with respect to histological appearance and immunological phenotype. It was possible to generate secondary cell lines from the tumors found in the inoculated SCID mice. Injection of one of these secondary cell lines into SCID mice resulted in the rapid development of lymphoma and as few as 10 cells were sufficient to establish the disease in the inoculated animals. In contrast cells of OCI-LY17 produced small tumor aggregates that did not appear to progress over time and did not cause death of the animals.(ABSTRACT TRUNCATED AT 250 WORDS)     

Improved engraftment of human acute myeloid leukemia progenitor cells in beta 2-microglobulin-deficient NOD/SCID mice and in NOD/SCID mice transgenic for human growth factors.

Primitive malignant progenitors defined as nonobese diabetic/severe combined immunodeficient (NOD/SCID) leukemia-initiating cells or NOD/SL-IC from patients with acute myeloid leukemia (AML) can be detected and quantitated in sublethally irradiated NOD/SCID mice. However, there is variability in the levels of bone marrow (BM) engraftment obtained after intravenous injection of cells from different AML samples. In the current study, AML cell engraftment in standard NOD/SCID mice was compared to that obtained with NOD/SCID mice transgenic for the human growth factor genes Steel factor (SF), interleukin-3 (IL-3) and granulocyte macrophage-colony-stimulating factor (GM-CSF) (N/S-S/GM/3) as well as beta 2 microglobulin-null NOD/SCID (N/S-beta 2m(-/-)) mice. Three of the eight AML samples that failed to engraft in standard NOD/SCID animals showed easily detectable and up to 70-fold increased in the number of leukemic cells in BM 8-12 weeks post-transplantation in each of the N/S-beta 2m(-/-) and N/S-S/GM/3 mouse strains. In two of the four AML samples studied at limiting dilution, the frequency of NOD/SL-IC detected was increased six- and seven-fold. Thus, in these novel mouse strains a broader spectrum of AML patient samples can be evaluated for their progenitor content and potentially studied for their response to innovative therapeutics in vivo.     

Homing and survival of thymidine kinase-transduced human T cells in NOD/SCID mice

The herpes simplex virus thymidine kinase (HSV-tk) gene conferring ganciclovir (GCV)-specific sensitivity to transduced cells might control Graft-versus-Leukemia (GvL)/Graft-versus-Host Disease (GvHD). Human T lymphocytes were engineered with an LSN-tk retroviral vector encoding tk and neomycin resistance (NeoR) genes. A total of 80 x 10(6) tk(+) lymphocytes were injected intraperitoneally in NOD-SCID mice. Engraftment was evaluated by human CD45(+)/CD3(+) cytofluorimetric analysis and NeoR-based polymerase chain reaction (PCR) on peripheral blood, bone marrow, liver, thymus, and spleen on day +5. After 14 days, GCV (10 mg/kg daily) cytofluorimetric analysis and PCR were repeated (day +19). Immunohistological studies with anti-CD3 monoclonal antibody followed by alkaline phosphatase and monoclonal anti-alkaline phosphatase staining were performed on spleen and liver at the same time points. Human CD45(+)/CD3(+) cells were engrafted in all tissues on day +5 according to cytofluorimetry, immunohistology, and PCR. Lymphocytes "homed" to the white pulp T-cell area and to the red pulp; liver localization is prevalently at the periportal area. After GCV (day +19), cytofluorimetry and immunohistology showed very few CD3(+) cells. PCR identified the transgene in 22% tissue samples (positive only in thymus and spleen). GvHD did not occur in any animal. These data demonstrate elevated doses of human-transduced CD3(+) cells engraft in NOD/SCID mice; after GCV, very few CD3(+) cells can be detected and those that escape treatment can be found in the thymus and in the spleen on day +19. Lack of full response to GCV may account for cases of GvHD in patients receiving tk-transduced T lymphocytes.     

Reduction of interstitial fluid pressure after TNF-alpha treatment of three human melanoma xenografts.

Tumour necrosis factor-alpha (TNF-alpha) reduced the interstitial fluid pressure (IFP) to 54-64% (P < 0.05) and the mean arterial blood pressure (MABP) to 70% (P < 0.01) of control values after 5 h in three human melanoma tumour lines transplanted to nude mice.     

Addition of bromelain and acetylcysteine to gemcitabine potentiates tumor inhibition in vivo in human colon cancer cell line LS174T

The combinations of Bromelain and Acetylcysteine (BromAc^®) with cytotoxics such as Gemcitabine, 5-Fluorouracil or Oxaliplatin have shown a dramatic reduction in IC50 values in a variety of cancers, including colon cancer, suggesting the possibility of effective treatment without undesired side effects. In the current study, we investigated whether a similar effect is present in vivo using the colorectal cell line LS174T. Animals after acclimatization were randomized and allocated equally in the groups for the different studies (safety, dose-escalation, and efficacy). Drugs were delivered by the intraperitoneal route and animals were monitored for wellbeing. Separately, an efficacy study was conducted with intraperitoneal drug delivery after intraperitoneal tumor induction. At the termination of the experiment, tumors and other tissues were collected for evaluation. BromAc^® was safe when delivered intraperitoneally in a rat model at the concentrations used. Subsequent investigations of these adjuvants in combination with Gemcitabine, Oxaliplatin, and 5-Fluorouracil in mice were also proven to be safe. Preliminary efficacy studies with Oxaliplatin and 5-Fluorouracil on tumor growth (LS174T) were negative. Gemcitabine was assessed with BromAc^® showing an almost 71% tumor inhibition compared to controls. This in vivo study indicates that Gemcitabine at 2 mg/kg in combination with BromAc^® 3 mg/300 mg/Kg was effective and safe, supporting its potential for future clinical application.     

Uptake of IgG in osteosarcoma correlates inversely with interstitial fluid pressure, but not with interstitial constituents.

The uptake of therapeutic macromolecules in solid tumours is assumed to be hindered by the heterogeneous vascular network, the high interstitial fluid pressure, and the extracellular matrix. To study the impact of these factors, we measured the uptake of fluorochrome-labelled IgG using confocal laser scanning microscopy, interstitial fluid pressure by the 'wick-in-needle' technique, vascular structure by stereological analysis, and the content of the extracellular matrix constituents collagen, sulfated glycosaminoglycans and hyaluronan by colourimetric assays. The impact of the microenvironment on these factors was studied using osteosarcomas implanted either subcutaneously or orthotopically around the femur in athymic mice. The uptake of IgG was found to correlate inversely with the interstitial fluid pressure and the tumour volume in orthotopic, but not subcutaneous tumours. No correlation was found between IgG uptake and the level of any of the extracellular matrix constituents. The content of both collagen and glycosaminoglycans depended on the site of tumour growth. The orthotopic tumours had a higher vascular density than the subcutaneous tumours, as the vascular surface and length were 2-3-fold higher. The data indicate that the interstitial fluid pressure is a dominant factor in controlling the uptake of macromolecules in solid tumours; and the site of tumour growth is important for the uptake of macromolecules in small tumours, extracellular matrix content and vascularization.     

The growth and metastasis of human, HER-2/neu-overexpressing tumor cell lines in male SCID mice

HER-2/neu is overexpressed on a variety of human adenocarcinomas and overexpression has been associated with a poor prognosis. For this reason, HER-2 has become an attractive target for immunotherapy. To facilitate testing of anti-HER-2-monoclonal antibodies (MAbs) and immunotoxins (ITs), we have evaluated the in vivo growth and metastatic spread of three HER-2-overexpressing human breast cancer cell lines (BT474, MDA-MB-453 and HCC1954) and one ovarian cancer cell line (SKOV3.ip1) in pre-irradiated male SCID mice using subcutaneous (s.c.), intravenous (i.v.) and intraperitoneal (i.p.) routes of injection. All the cell lines tested grew as s.c. tumors and the growth of BT474 and MDA-MB-453 cells after s.c. injection was improved by co-inoculation with Matrigel. Metastases to the lungs were detectable by PCR or histopathology after s.c. injection of BT474 and to a much lesser extent after s.c. injection of HCC1954, MD-MB-453 and SKOV3.ip1cells. I.P. injection of HCC1954 and SKOV3.ip1 cells produced fatal ascites while i.v. injection of SKOV3.ip1, but not BT474 or MDA-MB-453 cells, resulted in infiltration of lungs and death within 9–11 weeks.     

Microdialysis of 5-S-cysteinyldopa from interstitial fluid in cutaneous human melanoma transplanted to athymic mice.

Microdialysis was investigated as a tool for the determination of the extracellular concentration of the pigment metabolite 5-S-cysteinyldopa in human melanoma transplanted to athymic mice. Histology of the tumour with the microdialysis probes in situ showed no tissue damage. With probes equipped with polycarbonate membranes (20 kD) extraction (relative recovery) was approximately 50% at pH 4.0 and flow rates of 1 microliter/min, but at pH 7.0 recoveries were markedly lower, particularly from serum. In a first series of human melanomas transplanted to athymic mice low concentrations of 5-S-cysteinyldopa were detected in only two out of ten dialysates and were not detected in the other eight. Utilizing devices constructed for comparison of membrane characteristics in vitro we found about 4-fold higher recoveries with cuprophane and polyamide membranes than with polycarbonate membranes. Therefore newly constructed microdialysis probes (CMA/11) with cuprophane membranes were tested in vitro and gave recoveries of 38-48% from Ringer-Acetate solutions and 22-31% from serum, and the pH effects were low. When these probes were utilized in a second series of melanomas transplanted to athymic mice, 5-S-cysteinyldopa could easily be quantified in 10/10 experiments. A steady-state level of the dialysate 5-S-cysteinyldopa concentration was reached after 45 min.     

Inhibition of growth of human nasopharyngeal cancer xenografts in SCID mice by arsenic trioxide

AIMS AND BACKGROUND: It is known that arsenic trioxide (As2O3) can induce clinical remission in patients suffering from acute promyelocytic leukemia. It has been suggested that the agent might also be effective against other malignancies. This study was done to explore the efficacy of As2O3 in the treatment of human nasopharyngeal cancer xenografts in SCID (severe combined immunodeficiency) mice. METHODS: Human nasopharyngeal cancer cells from the CSNE-1 cell line were implanted subcutaneously into SCID mice to produce tumors. The tumor inhibitory rate in vivo was assessed after intraperitoneal administration of As2O3. Histopathological changes in the tumor tissues and the toxicity of As2O3 to the liver, heart and kidneys of the host mice were also investigated. RESULTS: At doses of 1 mg/kg and 5 mg/kg As2O3 induced apoptosis in nasopharyngeal carcinoma cells. At 5 mg/kg As2O3 also induced cancer cell differentiation, it reduced the PCNA expression, and inhibited tumor growth. The tumor growth inhibitory rate in this experimental group was 76.02%. No nephrotoxicity was observed histologically at these dose levels but some pathological changes in liver and cardiac tissues were found. As2O3 proved lethal to the SCID mice at a dose of 10 mg/kg. CONCLUSION: As2O3 has an inhibitory effect on human nasopharyngeal carcinoma xenografts in SCID mice. The mechanism of antitumor activity may be due, at least in part, to the induction of apoptosis and differentiation in cancer cells.