Antibody to keyhole limpet hemocyanin labels retinal horizontal cells in some amphibians, but not in others

The intact rat adrenal gland in short-term (3-h) organ culture may be amenable for the identification of factors involved in regulating adrenal cell apoptosis under defined conditions. In this model, culturing in the absence of trophic support (basal; control) triggered apoptosis in the intact rat adrenal gland; oligonucleosome formation, a measure of apoptosis, was 56.4-fold greater than that of glands snap-frozen at the start of incubation. Angiotensin II (Ang II) (100 nM) enhanced apoptosis by 67% over control. By contrast, adrenocorticotropin (ACTH) (100 nM) attenuated basal apoptosis by 59% and antagonized the enhanced apoptosis induced by Ang II back to the control level. Quartering of the glands enhanced basal oligonucleosome formation 182.2% greater than that of intact glands. Interestingly, quartering of the glands abolished the influences of Ang II and ACTH on apoptotic DNA fragmentation, but did not alter ACTH-induced corticosterone secretion. These data suggest that some level of gross adrenal structural information or compartmentalization, sufficiently disrupted by quartering, is required for the hormonal modulation of adrenal cell survival.     

Cross reacting antibodies against keyhole limpet haemocyanin may interfere with the diagnostics of acute schistosomiasis

The number of individuals catching schistosomiasis has increased with the popularity of 'primitive tourism' in Africa. Highly immunogenic material originating from the intestine of intravascular adult schistosomes gives rise to an antibody response making possible early identification of infected individuals using serology. Antibodies against gut associated antigens (anti-GAA), detected by indirect immunofluorescence microscopy employing sections of adult worms as antigen may occur before the onset of egg production. In the present study we show that this well known schistosomiasis-specific anti-GAA staining reaction can be confused with a similar staining reaction with ducts of both male and female worms. Antibodies with duct reactivity were seen in sera both from schistosomiasis-patients and patients with some other invasive worm infections. Cross reactive anti-duct antibodies appear to have different specificity. One cross reactive antibody reacted with antigenic epitopes present in keyhole limpet haemocyanin (KLH). Anti-duct reactivity could be inhibited by absorption with KLH. This was most obvious in the trichinellosis patient sera.     

Immunotherapy of cancer with antibody

A long-term suppression of a transplanted solid tumor that has been growing in a syngeneic animal can be achieved by the administration of antibody against the tumor. The susceptibility of such growing tumor cells to antibody treatment is similar to that of a comparable number of freshly injected tumor cells.     

Keyhole limpet hemocyanin: structural and functional characterization of two different subunits and multimers

Keyhole limpet hemocyanin (KLH), the large respiratory glycoprotein from the primitive gastropod mollusc, Megathura crenulata, is a potent immunogen used classically as a carrier protein for haptens and more recently in human vaccines and for immunotherapy of bladder cancer. Two KLH isoforms were identified and isolated by high-performance anion exchange chromatography. Subsequent analyses disclosed that these isoforms--designated KLH-A and KLH-B--were composed of distinct subunits that differed in primary structure, molecular weight (KLH-A was 449,000 and KLH-B was 392,000), polymerization/reassociation characteristics, and O2-binding constants (KLH-A had a P50 of 7.32 and KLH-B had a P50 of 2.46). Both subunits appear to be composed of eight oxygen binding domains, and reassociate in solution only with like subunits. These results support the concept that structural and functional heterogeneity is a common feature of molluscan hemocyanins, and provide a rational basis for studying and optimizing the immunostimulatory properties of KLH.     

Immunotherapy of metastatic kidney cancer

Interleukin 2 (IL2), like Interferon alpha (IFN), is active in metastatic renal cancer, considered to be a chemoresistant cancer. 20 to 30% of objective responses, including 5 to 10% of complete remissions are reported with various protocols of IL2 administration. The considerable toxicity is now well controlled, allowing treatments to be administered in standard wards or even on an outpatient basis by subcutaneous injection. IFN, generally given as long-term treatment, achieved average response rates of between 15 and 20%. Although IL2 and IFN have been granted Product Marketing Authorization in France, the modalities of optimal administration, the place of the combination of IL2-Interferon alpha and the factors predictive of response to treatment still remain unclear.     

Immunotherapy of head and neck cancer

Understanding the mechanisms responsible for photodamage to the skin is most important for dermatology. 3-D cultures have been used as tools to mimic the in vivo situation for several years. We irradiated such a system containing human dermal fibroblasts cultured in collagen gels, a well-known model considered to be a dermal equivalent, which reproduces the interaction between cells and the surrounding extracellular matrix. The effects of solar irradiation (315-800 nm) on the steady-state levels of the mRNAs of extracellular matrix components (type I and III collagens) and their degrading enzymes (interstitial collagenase, MMP-1 and stromelysin 1, MMP-3) were measured. Exposure to low levels of solar radiation (0-10 J cm-2 in the UVA, i.e. suberythemal UVA doses) caused a transient decrease in type I procollagen mRNA, an increase in MMP-mRNA, and no change in type III procollagen mRNA steady-state levels. These results describe the early changes in the connective tissue of the skin following exposure to low-level solar stimulation, and may help explain the long-term changes in photodamaged skin.     

Volume of reaction by the Archibald ultracentrifuge method (lobster hemocyanin)

Samples of lobster hemocyanin (Homarus americanus) under conditions of reversible reaction between whole (25 S) and half (17 S) molecules have been subjected to accurately known nitrogen pressures in analytical ultracentrifuge cells. A modified pressurization chamber of the type developed by Schumaker and colleagues has been constructed for this purpose. The molecular weight was then determined at the top (liquid-gas) meniscus, by means of the Archibald method. The logarithmic dependence upon pressure of the derived equilibrium constant then gave directly the volume of reaction. Experiments were performed in veronal-citrate buffers at pH 8, where the molar volume of formation of whole (dodecameric) molecules from half molecules appears to be negative, and at pH 8.46 in veronal-citrate buffer in the presence of 0.003 molar free calcium ion, where the molar volume of formation was estimated to be + 390 cm3/mole. In glycine-sodium hydroxide buffer at pH 9.6 containing 0.0047 molar free calcium, the molar volume of formation of whole molecules was estimated to be +120 +/- 70 cm3, corresponding to an estimated difference in partial specific volume between whole molecules and half molecules of only 1.3 (10)-4cm3/gram. The correctness of the sign of this value in glycine buffer has been verified by pressure-jump light-scattering experiments.     

Evaluation of c-ras oncogene product (p21) in superficial bladder cancer

OBJECTIVES: The aim of our study is the evaluation of the prognostic importance of p21 protein in superficial bladder cancer. METHODS: One hundred and fourteen patients with an initial diagnosis of monofocal bladder cancer (stage Ta-T1) following TUR were investigated. On the tissue removed by TUR, besides the usual pathological evaluation, an immuno-histochemical investigation was carried out in order to ascertain the presence of c-ras oncogene product (protein p21). The actuarial curves concerning the time free from the first recurrence were computed, comparing different subgroups in regard to protein p21 presence, grade and stage of the tumour. RESULTS: The analysis of the results shows the importance of tumour stage as a predictor of recurrence, as well as that of the presence of c-ras products. This last factor increases the risk of recurrence almost 2-fold, in the same time lag, for c-ras-positive patients (p < 0.001). The prognostic significance of c-ras is independent of stage. CONCLUSION: Our data underline the possibility of acquiring important information on the prognosis of superficial bladder cancer patients, pointing out the significance of c-ras oncogene product.     

Immunotherapy in metastatic cancer of the kidney

Evidence accumulated over the last 15 years has clearly demonstrated that metastatic renal cell cancer is an excellent model of the effect of immunotherapy for the treatment of cancer. To date, at least 2 cytokines, interleukin-2 and alpha-interferon have been found to be effective. Objective response is obtained in 15 to 30% of the patients treated with interleukin-2 and 10 to 30% with interferon. Complete response can be achieved in 5% of the cases. Clinically, the best results are seen in patients in good general health and lung metastasis. Complete response for more than 5 years is often observed. Combination protocols with both cytokines and other combinations with infusion of activated lymphocytes have not shown to be more effective than one cytokine alone. It may be possible to obtain higher response rates by combining cytokines with chemotherapy protocols. Surgery still has a role to play however, particularly in patients with an isolate accessible metastasis. Among the perspectives for new immunotherapies, interleukin-12, a strong stimulator of the natural killer population is in phase II trials. Other possibilities include the use of selective populations of lymphocytes as adoptive immunotherapy or combinations using immunotherapy and surgery. Despite the enthusiasm generated by these new techniques, it is imperative to continue rigorous clinical trials in order to develop immunotherapy as a reliable routine treatment.     

Sensitization of human bladder cancer cells to Fas-mediated cytotoxicity by cis-diamminedichloroplatinum (II)

PURPOSE: The resistance of bladder cancer to anticancer chemotherapeutic drugs is a major problem. Several immunotherapeutic approaches have been developed to treat drug-resistant tumor cells. The Fas antigen (Fas)-Fas ligand pathway is involved in cytotoxic T lymphocyte and natural killer cell-mediated cytotoxicity. Like the Fas ligand, anti-Fas monoclonal antibody (mAb) induces apoptosis in tumor cells expressing Fas. Several anticancer drugs also mediate apoptosis and may share with Fas common intracellular pathways leading to cell killing. We reasoned that treatment of drug-resistant cancer cells with a combination of anti-Fas mAb and drugs might overcome their resistance. This study has investigated whether anticancer drugs synergize with anti-Fas mAb in cytotoxicity against bladder cancer cells. MATERIALS AND METHODS: Cytotoxicity was determined by a 1-day microculture tetrazolium dye assay. Synergy was assessed by isobolographic analysis. RESULTS: Treatment of the T24 human bladder cancer cell line with anti-Fas mAb in combination with 5-fluorouracil, mitomycin C or methotrexate did not overcome resistance to these agents. However, treatment of T24 tumor cells with a combination of anti-Fas mAb and cisdiamminedichloroplatinum (II) (CDDP) resulted in a synergistic cytotoxic effect. In addition, the CDDP-resistant T24 line (T24/CDDP) was sensitive to treatment with a combination of anti-Fas mAb and CDDP. Synergy by combination of anti-Fas mAb and CDDP was also achieved in three other bladder cancer lines and four freshly derived human bladder cancer cells. The combination of anti-Fas mAb and carboplatin also resulted in a synergistic cytotoxic effect on T24 cells; however, the combination of anti-Fas mAb and trans-diamminedichloroplatinum (II) resulted in an additive cytotoxic effect. Treatment with CDDP enhanced the expression of Fas on T24 cells. The synergy achieved in cytotoxicity with anti-Fas mAb and CDDP was also achieved in apoptosis. Incubation of T24 cells with anti-Fas mAb increased the intracellular accumulation of CDDP. Treatment of freshly isolated bladder cancer cells with CDDP enhanced their susceptibility to lysis by autologous lymphocytes. CONCLUSIONS: This study demonstrates that combination treatment of bladder cancer cells with anti-Fas mAb and CDDP overcomes their resistance. Synergy was achieved with established CDDP-resistant bladder cancer cells and freshly isolated bladder cancer cells. In addition, the sensitization required low concentrations of CDDP, thus supporting the potential in vivo application of combination of CDDP and immunotherapy in the treatment of CDDP- and/or immunotherapy-resistant bladder cancer.     

Immunotherapy of cancer with "immune" RNA. A preliminary report

A phase I clinical trial of immunotherapy with "Immune" RNA was undertaken fifteen months ago. Twenty-six cancer patients were treated with RNA extracted from the lymphoid organs of sheep immunized with either autologous tumor cells or allogeneic tumor cells of the same histologic type. Eighteen patients had gross disease and eight had minimum residual disease. RNA was administered weekly, intradermally, at doses up to 9 mg/week without any significant local or systemic toxicity. Four patients improved, thirteen achieved stability of disease or possible improvement, seven were treatment failures, and two are indeterminate. Lymphocyte-mediated cytotoxicity to allogeneic tumor target cells of the same histologic type were monitored in eleven patients. In seven patients, cytotoxicity increased after "Immune" RNA therapy; no change was observed in three patients; a decrease was noted in one patient. There appeared to be a possible correlation between cytotoxicity assessed in vitro and clinical response. There is some evidence that these responses may be specific for the particular tumor used to immunize the RNA donor.     

Genetics of bladder cancer

Bladder cancer manifests many different forms, ranging from superficial to aggressive muscle invasion, which suggests that various genetic alterations are responsible. Several attempts have been made to establish correlations between specific genetic alterations and various grades of the disease. Numerous types of chromosomal abnormalities have been observed, involving [1p, 1q, 2q, 3p, 4p, 5q, i(5p), +7, +8, 8p, 9p, 9q, 10q23-25, 11p, 11q, +11, 13q, 14q, 17p, 18q, 21q, and Y]. In addition, p53 mutations and loss of heterozygosity on various chromosomes have recently begun to shed light on the molecular pathways of transitional cell carcinomas of the bladder. We have begun to focus on specific genomic sites (especially 9q), although the heterogeneity of the disease and the variable presentation suggests divergent progression pathways. When the genetic basis of bladder cancer is fully understood, new diagnostic and therapeutic strategies will be developed, which in turn may promote better clinical management by pathologists and urologists.     

Predicting prognosis in patients with superficial bladder cancer

Bladder cancer is the most common urologic malignancy and is expected to affect approximately 54,000 people in 1998. Superficial bladder tumors (Tis, Ta, and T1 lesions) account for approximately 70% to 80% of these malignancies. Although many advances have been made in the management of patients with superficial bladder cancer, such disease often recurs and can progress, leading to death. It is therefore imperative to find an accurate method of identifying patients at risk for disease progression. Many recent investigations have been conducted to determine whether new biological markers will help predict disease progression and, to a lesser extent, tumor response to treatment. These new markers include DNA ploidy, S-phase, certain monoclonal antibodies, the p53 (alias TP53) tumor-suppressor gene, the retinoblastoma (Rb) gene, cell adhesion molecules, and angiogenesis. It is hoped that such prognostic indicators, coupled with cytoscopic and pathologic characteristics of the tumor, will lead to selective aggressive treatment of patients at high risk for progression while sparing low-risk patients from unnecessary procedures.     

The binding of Cd(II) to the hemocyanin of the Mediterranean crab Carcinus maenas

In idiopathic dilated cardiomyopathy (IDC), the relation between the signal-averaged electrocardiogram and ventricular tachycardia (VT) remains unclear. In this study, conventional time domain and frequency domain analyses (2-dimensional, spectral temporal mapping and spectral turbulence analysis) of the signal-averaged electrocardiogram were performed in 64 patients with IDC. Eight patients had a history of symptomatic sustained VT and an additional 24 had nonsustained VT recorded during ambulatory electrocardiography. Conventional time domain analysis, using the 25 and 40 Hz filter, and spectral temporal mapping, detected late potentials within the terminal QRS in 8 (13%), 14 (22%) and 18 (28%) patients, respectively. Late potentials were seen more often in patients with than without VT, and in patients with sustained versus nonsustained VT, but these differences were not significant. The predictive accuracy of these techniques in detecting either form of VT were: sensitivity, 22, 25 and 31%; specificity, 97, 81 and 75%; and overall predictive value, 59, 53 and 50%, respectively. Two-dimensional frequency domain analysis of the signal-averaged electrocardiogram revealed a higher energy and area ratio in patients with than without VT (entire QRS), and in patients with sustained versus nonsustained VT (entire QRS and terminal QRS). Spectral turbulence analysis was abnormal in 24 patients (39%), but no differences were observed between patients with and without VT. During follow-up (mean duration 18 +/- 14 months), 5 patients had arrhythmic events (3 died suddenly, 1 had aborted sudden death and 1 developed sustained VT).(ABSTRACT TRUNCATED AT 250 WORDS)     

Nephrogenic metaplasia (adenomatoid tumors) of bladder

Two cases of rare nephrogenic metaplasia of the bladder (adenomatoid bladder tumor) support the theory that these tumors originate as an atypical metaplastic response of urothelium to chronic infection or irritation. That these may also represent mesonephric or mesothelial choristomas is considered. Although benign, they may recur. Therapy should consist of antibiotics and transurethral resection or fulguration.     

The pathogenesis of bladder cancer

Carcinoma of the urinary bladder appears to arise, in at least some cases, from carcinoma in situ developing in a field of atypical epithelial proliferation. There is both a spatial and temporal relationship between invasive and in situ bladder cancer, although the exact relationship between the noninvasive flat and papillary types of tumors is not known. Prenoeplastic bladder lesions are defined as irreversible, although not necessarily progressive, and an experimental animal model of the disease has been developed. The appearance of pleomorphic microvilli on the luminal surface of epithelial cells of the urinary bladders of Fischer rats is correlated with the irreversibility of hyperplastic epithelial lesions induced by feeding N-[4-(5-nitro-2-furyl)-2-thiazolyl] formamide (FANFT) to the test animals. This alteration can be visualized by scanning electron microscopy of cytologic and histologic preparations.     

Organ preservation with radiochemotherapy in locally advanced bladder cancer

Bladder cancer is frequent in Western countries and predominantly affects males (ratio: 3:1). In 15-25% of cases there is muscular wall invasion. Treatment of > T1 tumors is radical cystectomy with or without preoperative radiotherapy. In T2 there is 60% survival at 5 years, 40% in T3. Exclusive radiotherapy used to prevent radical cystectomy has lower survival rates in T2 (30-40% at 5 years) as well as in T3 (20% at 5 years). Recently, concomitant radiotherapy and chemotherapy has been introduced again to prevent demolitive surgery. Results are similar or slightly superior than those of surgery alone. In our experience with radiotherapy 180 cGy daily for a total dose of 64 Gy in combination with fluorouracil in locally advanced tumors 40% bladder preservation was achieved.     

Selective bladder preservation by combination treatment of invasive bladder cancer

BACKGROUND: For patients with invasive bladder cancer the usual recommended treatment is radical cystectomy, although transurethral resection of the tumor, systemic chemotherapy, and radiotherapy are each effective in some patients. We sought to determine whether these treatments in combination might be as effective as radical cystectomy and thus might allow the bladder to be preserved and the cancer cured. METHODS: We enrolled 53 consecutive patients with muscle-invading bladder cancer (stages T2 through T4, NXM0) in a trial of transurethral surgery, combination chemotherapy, and irradiation (4000 cGy) with concurrent cisplatin administration. Urologic evaluation of the tumor response directed further therapy: radical cystectomy in the 8 patients who had incomplete responses, additional chemotherapy and radiotherapy (6480 cGy) in the 34 patients who had complete responses or who were unsuited for cystectomy, and alternative care in the 11 patients who could not tolerate either irradiation or chemotherapy. RESULTS: After a median follow-up of 48 months, 24 of the 53 patients (45 percent) were alive and free of detectable tumor. In 31 patients (58 percent) the bladder was free of invasive tumor and functioning well, even though in 9 (17 percent) a superficial tumor recurred and required further transurethral surgery and intravesical drug therapy. Of the 28 patients who had complete responses after initial treatment, 89 percent had functioning tumor-free bladders. CONCLUSIONS: Conservative combination treatment may be an acceptable alternative to immediate cystectomy in selected patients with bladder cancer, although a randomized clinical trial that included a group for simultaneous comparison would be required to produce definitive results.