Expression of an activated mammalian target of rapamycin in adenocarcinoma of the cervix: A potential biomarker and molecular target therapy

Alterations of the Akt/mTOR pathway have been observed in numerous types of cancer, thus this pathway represents an exciting new target for molecular therapeutics. We investigated the expression of activated Akt (p-Akt) and mTOR (p-mTOR) in patients with adenocarcinoma of the cervix and the involvement of the p-Akt/p-mTOR pathway in response to combination of inhibitor agents, rapamycin and LY294002, with conventional therapy, cisplatin, in vitro. Immunohistochemistry analysis of p-Akt and p-mTOR was conducted in 26 patients with adenocarcinoma of the cervix. Western blot analysis was performed to determine the protein expression involved in response to chemotherapy in cervical cancer cell lines. The results showed that p-Akt and p-mTOR were identified in 50% and 53.8% of adenocarcinoma of the cervix. The expression of p-mTOR was a significant independent marker for prognosis. A significant correlation between p-Akt and p-mTOR was observed. There was no correlation between their expressions with any of clinicopathological factors. In the in vitro study, cisplatin at CPI(50) targets both the apoptosis and survival pathway by activating the caspase-cascade; inhibiting Akt, mTOR, p70S6K, and 4EBP1. Combination of rapamycin with cisplatin induced synergistic interaction. On the other hand, combination with LY294002 resulted in either synergistic or antagonistic effect depending on the doses given. Rapamycin pretreatment potentiated cisplatin-induced apoptosis cell death and enhanced blocking of the survival pathway. Overall, the expression of p-mTOR is a significant prognostic marker of adenocarcinoma of the cervix and a potential molecular target for the treatment of cervical cancer. Inhibition of the mTOR pathway contributes to cisplatin-induced apoptosis in cervical cancer cell lines.     

Cancer‐associated fibroblasts educate normal fibroblasts to facilitate cancer cell spreading and T‐cell suppression

In some tumors, a small number of cancer cells are scattered in a large fibrotic stroma. Here, we demonstrate a novel mechanism for expansion of pro‐tumor fibroblasts via cancer‐associated fibroblast (CAF)‐mediated education of normal fibroblasts (NFs). When NFs were incubated with conditioned medium from CAFs, the resulting CAF‐educated fibroblasts (CEFs) generated reactive oxygen species, which induced NF‐κB‐mediated expression of inflammatory cytokines and the extracellular matrix protein asporin (ASPN), while expression of a common CAF marker gene, α‐SMA, was not increased. ASPN further increased CEF expression of downstream molecules, including indoleamine 2,3‐dioxygenase 1 (IDO‐1), kynureninase (KYNU), and pregnancy‐associated plasma protein‐A (PAPP‐A). These CEFs induce cytocidal effects against CD8⁺ T cells and IGF‐I activation in cancer cells. CEFs were generated without cancer cells by the direct mixture of NFs and CAFs in mouse xenografts, and once CEFs were generated, they sequentially educated NFs, leading to continuous generation of CEFs. In diffuse‐type gastric cancers, ASPN^high/IDO‐1^high/KYNU^high/α‐SMA⁻ CEFs were located at the distal invading front. These CEFs expanded in the fibrotic stroma and caused dissemination of cancer cells. ASPN may therefore be a key molecule in facilitating tumor spreading and T‐cell suppression.     

A phase III randomized study comparing oral doxifluridine and oral 5-fluorouracil after curative resection of gastric cancer

To investigate the usefulness of oral doxifluridine (5'-DFUR), an active intermediate metabolite of capecitabine (XELODA), in gastric cancer patients after curative resection, we conducted a phase III randomized controlled study to compare oral 5'-DFUR and oral 5-fluorouracil (5-FU). 485 gastric cancer patients with Stage II or III operative findings at curative resection were registered and administered 5'-DFUR (460 mg/m2/day, daily, for two years) or 5-FU (115 mg/m2/day, daily, for the same period). Although no differences in overall survival or disease-free survival were detected, subset analysis showed 5'-DFUR was more effective in reducing peritoneal recurrence than 5-FU (p = 0.047), and in patients with Stage III or stage IIIb (histologic findings) in the 5'-DFUR group had more favorable disease-free survival curves and survival curves than the 5-FU group with similar stages.     

Salivary chromogranin A as a measure of stress response to noise

Effects of noise on the secretion of salivary chromogranin A (CgA), which is considered to be a substitute measure of catecholamines, were investigated in a laboratory experiment. This study included 20 male subjects with normal hearing; their ages ranged from 21 to 24 years. Prior to the experiment, the subjects were asked to answer a questionnaire containing the 28-item General Health Questionnaire (GHQ-28) and Weinstein's noise sensitivity scale. White noise at 90 dB was presented to the subjects for 15 min with 15-minute-rest periods before and after noise exposure. It was shown that salivary CgA levels increased significantly during noise exposure and decreased immediately after it (Friedman's test, p = 0.001, two tailed). This result suggests that salivary CgA can be used to measure the stress response to noise. Furthermore, individual differences in the change in salivary CgA levels were discussed in relation to the subjective responses of the participants to the questionnaire. Some subjects showed prolonged elevation in the salivary CgA levels and the others showed immediate recovery or no effects. These individual differences correlated with the score on the somatic symptoms in GHQ-28; this implies that the score on the somatic symptoms in GHQ-28 could be a measure of physiological sensitivity to noise.     

Antithrombotic effects of KBT-3022, a novel antiplatelet agent,in an arterial thrombosis model in the guinea-pig

We examined the antithrombotic effect of KBT-3022, a novel antiplatelet agent, on photochemically induced arterial thrombosis in the guinea-pig saphenous artery, and compared its effect with that of aspirin and that of ticlopidine. Pretreatment with KBT-3022 [0.1 mg (0.2 μmole), 0.3 mg (0.7 μmole), 1 mg (2.2 μmole) and 3 mg (6.7 μmole)/kg, p.o.] 3 h prior to thrombosis induction prolonged the time required to achieve the thrombotic occlusion and inhibited the collagen (low concentration, 0.2 μg/ml; high concentration, 1 μg/ml)-induced platelet aggregation in whole blood ex vivo, each effect being concentration-dependent. The effects were tested of ticlopidine [30 mg (99.9 μmole), 100 mg (333.1 μmole) and 300 mg (999.2 μmole)/kg, p.o.] and aspirin [10 mg (55.5 μmole), 30 mg (166.5 μmole) and 100 mg (555.1 μmole)/kg, p.o.]. Both drugs prolonged the time to occlusion (but only at their highest concentration), and also inhibited the collagen (low concentration)- induced platelet aggregation in whole blood ex vivo. Aspirin [100 mg (555.1 μmole)/kg, p.o.], but not ticlopidine (at any concentration), inhibited the collagen (high concentration)-induced platelet aggregation. A good correlation was found between the antithrombotic and antiplatelet-aggregating effects of both KBT-3022 and aspirin, but not of ticlopidine. The antithrombotic potency of KBT-3022 was 300 times that of aspirin and 1000 times that of ticlopidine. These observations suggest that KBT-3022 may be clinically effective against platelet-rich thrombosis. Drug Dev. Res. 40:217–222, 1997. © 1997 Wiley-Liss, Inc.     

Gene transfer of secreted-type modified interleukin-18 gene to B16F10 melanoma cells suppresses in vivo tumor growth through inhibition of tumor vessel formation

Interleukin-18 is a novel cytokine identified as a strong inducer of interferon-gamma. Interleukin-18 has been shown to have similar bioactivities to interleukin-12 and to have antitumor efficacy in experimental models. In this study, we investigated whether the introduction of the interleukin-18 gene to B16F10 melanoma cells can induce antitumor response or not. Before the transfection, we modified the interleukin-18 gene to enable transfected tumor cells to secrete bioactive interleukin-18, because interleukin-18 does not have a signal sequence and requires processing by the interleukin-1 converting enzyme to attain the mature form. We found that B16 melanoma cells transduced with hybrid cDNA consisting of the interferon-beta signal sequence and mature interleukin-18 sequence, but not native interleukin-18, secreted a large amount of interleukin-18 and exhibited retarded tumor growth when injected in syngeneic mice. The antitumor effect was mostly abrogated by administration of anti-interferon-gamma antibody, but was not affected by in vivo depletion of CD8+ T cells or natural killer cells. Histologic analysis revealed that vascularization was markedly reduced and that necrosis was extensively induced in interleukin-18-secreting B16F10 melanoma (B16/IL18) tissues, whereas abundant tumor vessel formation was observed in B16/IL18 tissues of interferon-gamma-neutralized mice. We also found that chemokines, interferon-inducible protein-10 and monokine induced by interferon-gamma, were produced in B16/IL18 tissues and that the expression of both chemokines was dependent on that of interferon-gamma in the tumor tissues. Further, we showed that B16 melanoma cells secreted both chemokines in response to interferon-gamma. In addition, the expression of angiogenin, an angiogenic factor of melanoma, in B16 melanoma cells was reduced by interferon-gamma treatment. These results indicate that gene transfer of secreted-type interleukin-18 to B16F10 melanoma cells is a useful method of triggering an antitumor response without any systemic adverse effects and that the antitumor efficacy is mainly mediated by antiangiogenic activity, which is possibly involved in at least two dynamic changes induced by interferon-gamma inside B16 melanoma cells: the upregulation of antiangiogenic chemokines, interferon-inducible protein-10 and monokine induced by interferon-gamma, and the downregulation of angiogenic factor, angiogenin.     

Characterization of dermatitis arising spontaneously in DS-Nh mice maintained under conventional conditions: another possible model for atopic dermatitis

DS-Nh (DS Nh/+) mice spontaneously develop dermatitis when they are housed in a conventional environment. In this study, we analyzed the clinical and histopathological features of dermatitis in DS-Nh mice, which is characterized by erythema, edema, and erosion on the face, neck, chest and flexor surfaces of their forelegs with marked scratching behavior. Histopathological examination, including immunohistochemistry, revealed that inflammatory cells consisting of mast cells, eosinophils, CD4-positive T cell-dominant lymphocytes and CD11b-positive macrophages infiltrated the skin lesions. The cytokine production pattern of inflammatory cells in a lesional skin tissue was shifted to the Th2-type (IL-4) rather than the Th1 type (IFN-gamma). Serum IgE levels were elevated and correlated with the severity of the clinical skin conditions. These skin symptoms were observed in association with a colonization of Staphylococcus aureus. Similar clinical and histopathological symptoms were inducible with repeated percutaneous immunization of heat-killed S. aureus on the back of SPF DS-Nh mice. These results suggest that the spontaneous dermatitis that occurs in conventionally raised DS-Nh mice is comparable to a certain type of human atopic dermatitis (AD), which is associated with S. aureus, a recognized environmental factor. Thus, we consider that DS-Nh mice offer a useful model for investigating the pathogenesis of AD and for developing new therapeutic approaches or drugs for treating AD.     

A case of ileoileal intussusception caused by metastatic pedunculated tumor of cutaneous angiosarcoma

Cutaneous angiosarcoma is a rare aggressive vascular tumor that occurs in elderly patients and is usually located on the head and face. Metastases often develop in the cervical lymph nodes, lungs, bone, liver and spleen. There have been no reports of ileoileal intussusception due to metastatic tumor from cutaneous angiosarcoma. We reported a case of cutaneous angiosarcoma in a 67-year-old Japanese male accompanied with ileoileal intussusception due to metastatic angiosarcoma. We assume that the metastatic tumor in the small intestine was metastasized hematogeneously from cutaneous angiosarcoma, resulting in the formation of nodules and the rapid growth of a pedunculated tumor as a forerunner of the ileoileal intessusception.     

Tegafur-induced photosensitivity — evaluation of provocation by UVB irradiation

Dermatology, A 75-year-old Japanese man with photodistributed erythematous lichen planus like eruptions visited the dermatology clinic of Kobe University in November 1994. He had had an operation for gastric cancer in July 1992, and thereafter he had been taking 600 mg/d of tegafur. In July 1994, he was exposed to the sun for 2 h and the following day noticed an itchy rash in the areas exposed to the sun. He consulted his surgeon and stopped i taking the tegafur In October. Thereafter his eruption gradually improved. A biopsy J specimen taken on the initial visit to our hospital, 22 days after the cessation of tegafur, revealed perivascular collections of mononuclear cells in the upper dermis and slight liquefaction degeneration of the epidermal basal cells with some civatte bodies. Immunofluorescent staining showed no deposits of immunoglobulins or complements. Photosensitivity studies were performed with a bank of 7 fluorescent sunlamps (Toshiba FL20SE) emitting 280–370 nm (nnainly UVB energy, peaKing at 305 nm) and a bank ot 14 fluorescent black lights (Toshiba FL32SBL) emitting 300–420 nm (mainly UVA energy peaking at 365 nm). His minimal erythema doses (MEDs) of 58.5 mJ/cm² in UVB and of large dose of UVA over 12.6 J/cm² were in the normal range. Patch tests and two sets of photopatch tests were made with 5% tegafur and 5-fluorouracil (5-FU) in white petrolatum using Finn chambers. One set was exposed to 6.3 J/cm² of UVA, and a second set to a suberythemal dose of UVB, 40 mJ/cm², after 24 h of closed patch tests. Twenty four hours after UV irradiation for photopatch tests, and 48 h after the initial patch for patch tests, the reaction was evaluated. No abnormal reactions in patch and photopatch tests were detected. Tegafur was readministered at a dose of 200 mg every eight hours. After a total dose of 2400 mg (day 4), the skin of the back was exposed to UVB (6–120 mJ/cm²) and UVA (2–12.6 J/cm²) two hours after the last oral intake of tegafur. No decrease in MED or any abnormal reaction was observed. After a total of 3600 mg (day 6), a new skin site on the back was exposed to 3 MED of UVB. The next day (day 7, after a total dose of 4200 mg), 3 MED of UVB was irradiated on the same site and 5 MED of UVB was irradiated at a new site. Lastly, after a total dose of 4800 mg (day 8), 3 MED and 5 MED of UVB was again irradiated on the same sites that were irradiated on the previous day, and 10 MED UVB and 21 J/cm² UVA were irradiated at new individual sites. Eight days after the last irradiation (day 16, after 9600 mg of drug intake) the 10 MED UVB irradiated site revealed miliar-sized papules with a faint red hue after sunburn reaction. Simultaneously, an edematous erythema recurred on the face, neck, upper back, and hands, where the rash had previously been, without exposure to UVB or UVA. These tests were conducted while the patient was hospitalized and he was very careful not to be exposed to the sun, even through glass. The biopsy specimens of 10 MED irradiated sites and the flare-up lesion of his upper back revealed liquefaction degeneration of the epidermal basal cells with civatte bodies. Immunofluorescent staining study showed IgM, IgA, and C4 deposition of the civatte bodies in the flare-up lesion which had not been exposed to any UV irradiation.