Tumor development remains one of the major obstacles following organ transplantation. Immunosuppressive drugs such as cyclosporine and tacrolimus directly contribute to enhanced malignancy, whereas the influence of the novel compound mycophenolate mofetil (MMF) on tumor cell dissemination has not been explored. We therefore investigated the adhesion capacity of colon, pancreas, prostate and kidney carcinoma cell lines to endothelium, as well as their beta1 integrin expression profile before and after MMF treatment. Tumor cell adhesion to endothelial cell monolayers was evaluated in the presence of 0.1 and 1 μM MMF and compared to unstimulated controls. beta1 integrin analysis included alpha1beta1 (CD49a), alpha2beta1 (CD49b), alpha3beta1 (CD49c), alpha4beta1 (CD49d), alpha5beta1 (CD49e), and alpha6beta1 (CD49f) receptors, and was carried out by reverse transcriptase-polymerase chain reaction, confocal microscopy and flow cytometry. Adhesion of the colon carcinoma cell line HT-29 was strongly reduced in the presence of 0.1 μM MMF. This effect was accompanied by down-regulation of alpha3beta1 and alpha6beta1 surface expression and of alpha3beta1 and alpha6beta1 coding mRNA. Adhesion of the prostate tumor cell line DU-145 was blocked dose-dependently by MMF. In contrast to MMF's effects on HT-29 cells, MMF dose-dependently up-regulated alpha1beta1, alpha2beta1, alpha3beta1, and alpha5beta1 on DU-145 tumor cell membranes. We conclude that MMF possesses distinct anti-tumoral properties, particularly in colon and prostate carcinoma cells. Adhesion blockage of HT-29 cells was due to the loss of alpha3beta1 and alpha6beta1 surface expression, which might contribute to a reduced invasive behaviour of this tumor entity. The enhancement of integrin beta1 subtypes observed in DU-145 cells possibly causes re-differentiation towards a low-invasive phenotype.
Pathologic data indicate that human cytomegalovirus (HCMV) infection might be associated with the pathogenesis of several human malignancies. However, no definitive evidence of a causal link between HCMV infection and cancer dissemination has been established to date. This study describes the modulation of the invasive behavior of NCAM-expressing tumor cell lines by HCMV. Neuroblastoma (NB) cells, persistently infected with the HCMV strain AD169 (UKF-NB-4AD169 and MHH-NB-11AD169), were added to endothelial cell monolayers and adhesion and penetration kinetics were measured. The 140- and 180-kDa isoforms of the adhesion receptor NCAM were evaluated by flow cytometry, Western blot, and reverse transcriptionpolymerase chain reaction (RT-PCR). The relevance of NCAM for tumor cell binding was proven by treating NB with NCAM antisense oligonucleotides or NCAM transfection. HCMV infection profoundly increased the number of adherent and penetrated NB, compared to controls. Surface expression of NCAM was significantly lower on UKF-NB-4AD169 and MHH-NB-11AD169, compared to mock-infected cells. Western-blot and RT-PCR demonstrated reduced protein and RNA levels of the 140- and 180-kDa isoform. An inverse correlation between NCAM expression and adhesion capacity of NB has been shown by antisense and transfection experiments. We conclude that HCMV infection leads to downregulation of NCAM receptors, which is associated with enhanced tumor cell invasiveness.
The surgical removal of a primary tumor can result in the rapid growth of metastases. The production of angiogenesis inhibitors by the primary tumor is one mechanism for the inhibition of metastatic tumor growth. The effect of curative radiotherapy to a primary tumor known to make an inhibitor of angiogenesis and the effects on distant metastases has not been studied. We here show that the eradication of a primary Lewis lung carcinoma (LLC-LM), which is known to generate angiostatin, is followed by the rapid growth of metastases that kill the animal within 18 days after the completion of radiation therapy. The right thighs of C57BL/6 mice (n = 25) were injected s.c. with 1 x 10(6) LLC-LM cells. Animals were randomized to one of five groups: no irradiation, 40 Gy in one fraction, 30 Gy in one fraction, 40 Gy in two 20 Gy fractions, or 50 Gy in five 10 Gy fractions. Tumors were clinically eradicated in each treatment group. All of the surviving animals became dyspneic and were killed within 14-18 days after the completion of radiation therapy. Examination of their lungs revealed >46 (range, 46-62) surface metastases in the treated animals compared with 5 (range, 2-8) in the untreated animals. The lung weights had increased from 0.2 g (range, 0.19-0.22 g) in the controls to 0.58 g (range 0.44-0.84) in the experimental animals. The most effective dose regimen was 10 Gy per fraction for five fractions, and serial experiments were conducted with this fractionation scheme. Complete response of the primary tumor was seen in 25 of 35 (71%) mice. The average weight of the lungs in the nonirradiated animals was 0.22 g (range, 0.19-0.24 g) and in the irradiated animals was 0.66 g (range, 0.61-0.70 g). The average number of surface metastases increased from five per lung (range, 2-13) in the control animals to 53 per lung (range, 46-62) in the irradiated animals. Both differences were statistically significant with P < 0.001. If the nontumor-bearing leg was irradiated or the animals were sham-irradiated, no difference in the number of surface metastases or lung weights was observed between the control group and the treated group. Urinary levels of matrix metalloproteinase 2, the enzyme responsible for angiostatin processing in this tumor model, were measured and correlated with the viability and size of the primary tumor. Administration of recombinant angiostatin prevented the growth of the metastases after the treatment of the primary tumor. In this model, the use of radiation to eradicate a primary LLC-LM tumor results in the growth of previously dormant lung metastases and suggests that combining angiogenesis inhibitors with radiation therapy may control distant metastases.
An evidence-based consensus meeting was held with urologists, a pharmacist and a cardiologist to perform a structured benefit-risk analysis of reclassifying tadalafil, a phosphodiesterase type 5 (PDE5) inhibitor for treatment of erectile dysfunction (ED), to be available without prescription in Germany. As per the Brass process endorsed by regulatory authorities, an evidence-based Brass value tree was developed, which identified the incremental benefits and risks that should be considered above the safety and efficacy evidence required for prescription medicines. During the Group Delphi consensus meeting, the expert panel rated the likelihood and clinical impact of each benefit and risk on a scale of 0 (none) to 3 (high). Overall attribute scores were calculated from the product of the mean likelihood and mean clinical impact scores giving a possible score of 0-9. The overall benefit attribute scores ranged from 2.8 to 5.4. The overall risk attribute scores ranged from 0.2 to 2.2 though most were 1.0 or less (3 or more is generally considered to be of concern). On balance, the independent meeting scored the benefits of reclassification of tadalafil higher than the risks and considered the risk mitigation strategies of the packaging label and patient information leaflet (PIL) sufficient.
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