A novel variant of glutamine: fructose-6-phosphate amidotransferase-1 (GFAT1) mRNA is selectively expressed in striated muscle.

Glutamine:fructose-6-phosphate amidotransferase(GFAT) is the rate-limiting enzyme of the hexosamine synthesis pathway. Products of this pathway have been implicated in insulin resistance and glucose toxicity. GFAT1 is ubiquitous, whereas GFAT2 is expressed mainly in the central nervous system. In the course of developing a competitive reverse transcriptase-polymerase chain reaction assay, we noted that GFAT1 cDNA from muscle but not from other tissues migrated as a doublet. Subsequent cloning and sequencing revealed two GFAT1 mRNAs in both mouse and human skeletal muscles. The novel GFAT1 mRNA (GFAT1Alt [muscle selective variant of GFAT1]) is likely a splice variant. It is identical to GFAT1 except for a 48 or 54 bp insert in the mouse and human, respectively, at nucleotide position 686 of the coding sequence, resulting in a 16 or 18 amino acid insert at position 229 of the protein. GFAT1Alt is the predominant GFAT1 mRNA in mouse hindlimb muscle, is weakly expressed in the heart, and is undetectable in the brain, liver, kidney, lung, intestine, spleen, and 3T3-L1 adipocytes. In humans, it is strongly expressed in skeletal muscle but not in the brain. GFAT1 and GFAT1Alt expressed by recombinant adenovirus infection in COS-7 cells displayed robust enzyme activity and kinetic differences. The apparent K(m) of GFAT1Alt for fructose-6-phosphate was approximately twofold higher than that of GFAT1, whereas K(i) for UDP-N-acetylglucosamine was approximately fivefold lower. Muscle insulin resistance is a hallmark and predictor of type 2 diabetes. Variations in the expression of GFAT isoforms in muscle may contribute to predisposition to insulin resistance.

A cell-based drug delivery system for lung targeting: I. Preparation and pharmacokinetics.

A drug-loaded tumor cell (DLTC) system has been developed for lung metastasis-targeting drug delivery. Doxorubicin was loaded into B16-F10 murine melanoma cells (96 microg/10(6) cells). The loading process led to the death of all the carrier cells. The diameter of DLTC was 15.03+/-2.36 microm (mean +/- SD). The amount and rate of doxorubicin being released from the DLTC mainly depended on the drug loading and carrier cell concentration. Over a 6-month storage in phosphate buffered saline (PBS) at 4 degrees C, the decrease in intracellular drug concentration and the carrier cell number were less than 25% and 5%, respectively. After a bolus injection of 30 microg doxorubicin in either DLTC form or free solution into the mice tail veins, drug deposit in the lung from DLTC was 3.6-fold of that achieved by free drug solution. The latter resulted in higher drug content in liver and spleen. Extensive trypsinization of DLTC reduced its lung targeting effect by 30%, and the density of surface adhesion molecule GM3 on DLTC surface by 25%. In conclusion, this DLTC system demonstrated a lung-targeting activity that may be partially attributed to its specific surface characteristics.

A cell-based drug delivery system for lung targeting: II. Therapeutic activities on B16-F10 melanoma in mouse lungs.

The in vitro and in vivo anticancer activities of doxorubicin-loaded B16-F10 murine melanoma cells (DLTC) were evaluated. DLTC showed similar growth-inhibitory effects against live B16-F10 cells with doxorubicin solution in cell culture system, with the IC50 of 0.11 microM and 0.17 microM, respectively. However, DLTC demonstrated higher effectiveness than the free solution in treating mouse lung cancer caused by live B16-F10 cells. Syngeneic C57BL mice were inoculated intravenously with live B16-F10 cells first, and then received daily treatment of intravenous injections of doxorubicin in either DLTC or free solution form. Compared with the control group treated with phosphate-buffered saline, DLTC eradicated almost all the lung cancer colonies (>99%), while the free solution form reduced the colonies by 61%, when the treatment was given at an early stage. If the treatment started after the establishment of micrometastatic colonies in the mouse lungs, DLTC and free solution treatment resulted in 85% and 30% cancer reduction, respectively. Additional experiments demonstrated that the reduction of lung cancer colonies by DLTC was related to the initial treatment time: the earlier the treatment, the greater the effect. In conclusion, DLTC showed better therapeutic outcomes than free solution form in treating lung cancer of our animal model.

Keyhole limpet hemocyanin immunotherapy of bladder cancer: laboratory and clinical studies.

BACKGROUND: Since the serendipitous observation by Olsson in 1974 that patients immunized with 5 mg of keyhole limpet hemocyanin (KLH) had a marked reduction in recurrence of superficial bladder cancer, multiple laboratory and clinical studies have confirmed the efficacy of KLH immunotherapy. RESULTS: In 1981, we reported that KLH immunotherapy reduced tumor growth and prolonged survival in the MBT-2 murine model of transitional cell carcinoma (TCC), and in 1988, Jurincic and co-workers demonstrated that KLH was superior to mitomycin C chemotherapy in preventing bladder tumor recurrence. Subsequent studies using Immucothel (Biosyn), crude KLH, and endotoxin-free KLH confirmed the efficacy of KLH immunotherapy in the MBT-2 murine bladder cancer model (p < 0.05), and resulted in up to 100% survival. CONCLUSIONS: To evaluate the efficacy of KLH immunotherapy in patients, a multicenter clinical trial was performed. Sixty-four patients with CIS or residual stage T(a), T(1) TCC, or both were enrolled in a phase I-II trial of escalating doses of weekly KLH given intravesically for 6 weeks. Patients were followed with cystoscopic examination, urine cytology, and bladder biopsy. Complete response was seen in 50% of patients with CIS, 20% of patients with residual T(a), T(1) TCC, and 33% of patients with both CIS and residual T(a), T(1) TCC. Responses occurred at all doses tested: 0.4, 2, 10 and 50 mg. No significant difference in response according to dose was noted, but optimal overall complete response was seen with a dose of only 2 mg. The toxicity of KLH is minimal. KLH appears to be a safe and highly effective immunotherapy for superficial bladder cancer.

Quinolone antibiotics: a potential adjunct to intravesical chemotherapy for bladder cancer.

OBJECTIVES: Despite complete transurethral resection of superficial bladder tumors, the recurrence rate averages 88% at 15 years. Intravesical chemotherapy decreases the recurrence rate, particularly if given immediately after tumor resection. Anticancer drugs such as doxorubicin target topoisomerase II as do the quinolone antibiotics. We evaluated two fluoroquinolones independently and in combination with doxorubicin for cytotoxic effects against bladder cancer cells in vitro. METHODS: Three human transitional carcinoma cell lines, T24 (grade I), HTB9 (grade II), and TccSup (grade IV), were exposed to either ciprofloxacin or ofloxacin in concentrations ranging from 0 (control) to 1000 microg/mL for 24, 48, and 96 hours. In a separate experiment, a 30% cytotoxic dose (IC30) of doxorubicin was applied to the cell cultures for 1 hour and washed off, followed by exposure to ciprofloxacin or ofloxacin for 48 and 96 hours. Cytotoxicity was evaluated using the MTT colorimetric assay. RESULTS: At 96 hours, significant cytotoxicity (P <0.05) for ciprofloxacin was seen starting at 12.5 microg/mL (HTB9, TccSup) and 50 microg/mL (T24) and for ofloxacin at 12.5 microg/mL (HTB9) and 50 microg/mL (TccSup, T24). Maximum cytotoxicity with ciprofloxacin was 95.4+/-0.4% (HTB9, 400 microg/mL) and with ofloxacin was 95.2+/-0.3% (HTB9, 800 microg/mL). Exposure to doxorubicin (IC30, 1 hour) resulted in cell kill rates of 30.9+/-5.2% (T24), 50.7+/-2.7% (HTB9), and 25.4+/-10.6% (TccSup). The addition of as little as 25 microg/mL of ciprofloxacin increased kill rates to 78.5+/-1.2% (T24), 61.2+/-1.6% (HTB9), and 74.2+/-2.4% (TccSup); P < 0.05 relative to doxorubicin alone. Similarly, 50 microg/mL of ofloxacin significantly increased kill rates to 81.8+/-1.6% (T24), 63.3+/-2.5% (HTB9), and 67.8+/-2.0% (TccSup). Both drugs showed even greater synergism at higher concentrations. CONCLUSIONS: Ciprofloxacin and ofloxacin exhibit significant time- and dose-dependent cytotoxicity against transitional carcinoma cells and significantly enhance the cytotoxicity of doxorubicin. These effects occur at concentrations achievable in the urine of patients after oral administration. This suggests that quinolone antibiotics might be useful as an adjunct to intravesical chemotherapy and might reduce seeding of cancer cells after transurethral resection of bladder tumors.

Exogenous 17beta-estradiol blocks alpha and mu but not pi class glutathione S-transferase immunoreactivity in epithelium of Syrian hamster vas deferens.

Members of the glutathione S-transferase (GST) family of detoxification enzymes play a role in chemotherapy resistance in certain cancers but have not been directly implicated as agents whose absence may predispose tissues to hormonally induced tumorigenesis. Here we report the development of a polyclonal antiserum to a hamster mu class GST, and immunohistochemical analysis of alpha, mu, and pi class GSTs to study the effects of hormone treatment on their expression in reproductive tract tissues of male golden Syrian hamsters. These animals develop leiomyosarcomas in the vas deferens after treatment with testosterone propionate (TP) and 17beta-estradiol (E2). High levels of all three GST classes were detected throughout the reproductive tract epithelium of control animals. In 100% of the experimental animals, 4 weeks of treatment either with E2 alone, or with E2 plus TP promoted a complete loss of immunostaining for alpha and mu class GSTs, but not for pi class GSTs, only in the epithelial lining of the vas deferens. In contrast, treatment with TP alone resulted in moderate hyperplasia of smooth muscle in the proximal vas deferens, with no cellular atypia and no changes in immunoreactivity of any of the GST classes. The consistent and site-specific nature of these results strongly suggests a functional role for GSTs in hormonally induced carcinogenic process. (J Histochem Cytochem 47:91-98, 1999)

Novel retroviral sequences are expressed in the epididymis and uterus of Syrian hamsters.

Combined androgen and oestrogen treatment of male or female Syrian hamsters results via an unknown mechanism in the formation of leiomyosarcomas in the reproductive tract. We have examined the possibility that retroviral gene expression may play a role in tumorigenesis. Evidence of virus-like particles in epididymis and seminal fluid is shown in electron micrographs. We identified expressed retroviral sequences by using RT-PCR to amplify a conserved retroviral reverse transcriptase coding region in RNA isolated from epididymis, testis, clarified seminal fluid and uterus. Phylogenetic analysis allowed us to classify the sequences into two distinct groups: (1) mammalian type-C viruses, having similarity to Moloney murine leukaemia virus, feline leukaemia virus and gibbon ape leukaemia virus amongst others; (2) a mixed ABCD group containing, for example, Chinese hamster and murine intracisternal A-particle virus sequences, mouse mammary tumour virus and human and simian retroviral sequences. The presence of putative full-length retrovirus related to mammalian type-C viruses in the epididymis and uterus was confirmed by Northern blot analysis. However, steroid treatment did not alter retroviral RNA levels in the epididymis or in a uterine tumour relative to untreated uterus. In summary, Syrian hamster reproductive tissues were found to express unique retroviral sequences; however, their role, if any, in hormonal carcinogenesis remains unresolved.

Photodynamic therapy (PDT) in the treatment of patients with resistant superficial bladder cancer: a long-term experience.

INTRODUCTION AND OBJECTIVE: Photodynamic therapy (PDT) combines a photosensitizer such as Photofrin with red laser light (630 nm) to destroy cancer cells. Investigators have reported effectiveness of PDT in the management of patients with recurrent superficial bladder cancer. We retrospectively reviewed our experience in 58 patients to assess the long-term role of PDT in the management of resistant superficial transitional cell carcinoma (TCC) including Ta, T1, and refractory carcinoma in situ (CIS) of the urinary bladder. MATERIALS AND METHODS: All 58 patients had failed at least one course of standard intravesical therapy or had contraindication for intravesical chemo- or immunotherapy. Patients with malignancy present (Ta-T1/Grade I-III, CIS) were accepted for ablative PDT. Patients undergoing prophylactic PDT after complete resection were confirmed to be tumor-free by cystoscopy and bladder was cytology before PDT. Post-PDT evaluations included weekly telephone contact to assess acute adverse reactions and assessment of efficacy and bladder toxicity at three months and quarterly thereafter. RESULTS: These 58 patients underwent a single PDT treatment with 2.0 or 1.5 mg/kg of Photofrin and 10-60 J/cm2 light (630 nm). At three months, complete response rates were 84% and 75% for residual resistant papillary TCC and refractory CIS respectively; and 90% of patients treated prophylactically had not had recurrences. At a median followup of 50 months (range 9-110), 59% (34/58) of the responders are alive, with 31/34 still disease-free. CONCLUSION: PDT using 1.5 mg/kg of Photofrin and 15 J/cm2 of light (630 nm) should be considered a safe and effective treatment for refractory CIS or recurrent papillary TCC.

Glutathione S-transferase in hormonal carcinogenesis.

Treatment with testosterone propionate (TP) and diethylstilbestrol (DES) or TP and estradiol (E2) for 8-9 months causes development of leiomyosarcomas in the vas deferens or uterus of Golden Syrian hamsters at a frequency of 100%. In males, treatment with estrogens alone results in renal tumors, fatal within 6 months. No leiomyosarcomas have been detected after treatment with estrogens alone, perhaps due to this high mortality rate. In tissue culture, treatment with the glucocorticoid (GC) triamcinolone acetonide (TA) results in an increased expression of a mu-class glutathione S-transferase (GST) (hGSTYBX). We have characterized this induction as a secondary response, i.e. requiring new protein synthesis. Here we describe homologies to known transcription factor-binding sites in the hGSTYBX promoter which may be involved in the induction event. hGSTYBX is a member of a superfamily of detoxification enzymes, induced by genotoxic compounds and reactive oxygen species (ROS). We also describe the effects of several known inducers of other GST family members on hGSTYBX. While implicated in many chemotherapeutic-resistant tumors, GST enzymes have not yet been characterized as a functional agent in hormonal carcinogenesis. This latter possibility is the focus of our investigations. To study the effects of hormone treatment on GST levels in vivo, we have developed a polyclonal antibody to hGSTYBX, and conducted immunohistochemistry on tissues from control and treated animals. Treatment with TP and E2 causes a loss of hGSTYBX expression in the epithelium of the vas deferens. We hypothesize that the loss of this protective enzyme leaves the cells vulnerable to the genotoxic effects of estrogen or estrogenic metabolites.

Liposomal delivery of alpha-Interferon to murine bladder tumor cells via transferrin receptor-mediated endocytosis.

The role of transferrin receptor-mediated endocytosis in promoting murine bladder tumor cell (MBT2) uptake of liposomes and the antiproliferative effect of liposome-entrapped alpha-interferon (alpha-IFN) against MBT2 were investigated. Liposomes (0.11 microm) were prepared using phosphatidylcholine and phosphatidylserine in a molar ratio of 7:3, with or without surface conjugation of transferrin-polylysine (TFPL). The uptake of plain liposomes (without TFPL) by MBT2 was less than 5% after incubation for 48 h. In contrast, cell uptake of TFPL-liposomes was markedly enhanced by TFPL in a dose-dependent manner and reached plateau levels in 24 h. This increase was partially blocked by the addition of free transferrin, suggesting that the uptake process involves transferrin receptor-mediated endocytosis. The antiproliferative activity of alpha-IFN (100-200 U/well), delivered via plain liposomes, measured against blank liposome control, was in the range of 25-35%, which was similar to that of free alpha-IFN. In comparison, inhibition of cell proliferation by same concentrations of alpha-IFN delivered by TFPL-liposomes was 90-100%. These results show a strong correlation between antiproliferative activity and the uptake of liposomes by the tumor cells, indicating that TFPL-liposomes promote intracellular delivery of alpha-IFN and enhance the effect of alpha-IFN against MBT2 cell growth. The potential cytotoxicity of drug-free liposomes was also investigated. Liposomes containing various concentrations of TFPL showed significant dose- and time-dependent antiproliferative activity against MBT2. This effect maybe attributed to lipidosis and/or the destruction of intracellular cycle of iron transport.

Allium sativum (garlic) treatment for murine transitional cell carcinoma.

BACKGROUND: Currently, immunotherapy with Bacillus Calmette-Guerin (BCG) is the most effective treatment for superficial bladder carcinoma, but treatment-related toxicity may limit its use in some patients. Alternative treatments are needed for patients who fail to respond to BCG immunotherapy. Allium sativum (AS), or garlic, is known to have a broad range of biologic activities, including immune stimulation and reported antitumor activity. For these reasons, the authors conducted a series of experiments designed to explore the possible therapeutic effects of AS in the MBT2 murine bladder carcinoma model. METHODS: C3H/HeN mice were randomized prior to initiation of each experimental protocol. Mice received 1 x 10(3) MBT2 cells in 0.1 mL RPMI-1640, administered subcutaneously into the right thigh, on Day 0 of the experiment. AS was injected at the site of tumor transplantation on Day 1 and at 2- to 7-day intervals up to Day 28. To evaluate the effects of oral AS in this model, treatment was initiated 30 days prior to tumor inoculation and continued for 30 days after tumor inoculation. Animals in all experiments were followed for tumor incidence, tumor growth, and survival. RESULTS: In the initial experiments, subcutaneous AS significantly reduced tumor volume compared with the saline control (P < 0.05). Unfortunately, treatment-related death was also observed, requiring reduction in the total dose of AS. Animals that received 5 weekly immunizations of AS (5 mg, 5 mg, 1 mg, 1 mg, and 1 mg; cumulative dose = 13 mg) had significantly reduced tumor incidence, tumor growth, and increased survival when compared with animals that received the saline control. No treatment-related deaths were observed with this treatment schedule. To determine whether systemic AS administration might be effective, orally administered AS was tested at doses of 5 mg, 50 mg, and 500 mg per 100 mL of drinking water. Mice that received 50 mg oral AS had significant reductions in tumor volume (P < 0.05) when compared with animals that received the saline control, and mice that received 500 mg oral AS had significant reductions in both tumor volume and mortality (P < 0.05). CONCLUSIONS: The significant antitumor efficacy of subcutaneous and oral AS warrants further investigation and suggests that AS may provide a new and effective form of therapy for transitional cell carcinoma of the bladder.

Sequential whole bladder photodynamic therapy treatments: A preclinical study.

We postulated that sequential whole bladder photodynamic therapy (WBPDT) treatments with a low WBPDT dose would result in improved safety profile and good local tumor control. However, the drawback with such a proposal is the potential cumulative effect of sequential WBPDT treatments on bladder function. We designed this preclinical study to determine the safety of sequential WBPDT treatments. Six female dogs underwent a single WBPDT treatment comprising 1.5 mg/kg of Photofrin® and 15 J/cm(2) of light. Four dogs received a second treatment; and three dogs received three treatments. Pre- and post-WBPDT evaluations included cystoscopy and saline cystometry at baseline, 1 week, and 12 weeks. Gross and histopathologic analysis of cystectomy specimens occurred at 1 and 12 weeks. A single photodynamic therapy (PDT) treatment induced average bladder capacity losses of 11% (0-33) and 0% at post-WBPDT weeks 1 and 12, respectively. A second sequential WBPDT treatment caused average bladder capacity losses of 36% (0-57%) and 17% (2-24%) at weeks 1 and 12, respectively. Three sequential WBPDT treatments induced average bladder capacity losses of 22% (0-42) and 0% at weeks 1 and 12, respectively. Full recovery in bladder capacity occurred in all cases except after the second sequential treatment, which induced a persistent bladder capacity loss of 17% at 12 weeks. Histopathologic analysis of cystectomy specimens revealed a focal discernible injury to the superficial muscle in only one of the dogs that received three treatments. We conclude that sequential WBPDT treatments using low dose PDT Photofrin® (1.5 mg/kg and light ≤15 J/cm(2)) is safe, and we recommend using this low WBPDT dose in clinical investigation.

Accuracy of three-dimensional linear and angular estimates obtained with the Ariel Performance Analysis System.

Computer-assisted video motion analysis is a method of evaluating human kinematics that offers promise both for research and for clinical application. This study determined the upper limits of accuracy and consistency of linear and angular measures obtained using the Ariel Performance Analysis System. Reference standards included a meter stick and a universal 360 degrees goniometer. Average mean error observed for reconstruction of absolute point estimates was found to be less than 3.5 mm. Mean error estimate for three-dimensional (3D) reconstruction of a linear standard was found to be 1.4 mm (SD 0.30). Average mean angular error observed for 3D reconstruction of goniometer settings 10 degrees to 170 degrees was found to be 0.26 degrees (mean SD 0.21). System users are cautioned that some increased error associated with software derivation of joint angles exists as angles approach 180 degrees, use of wide-angle lens accessories introduces a systematic field-dependent bias; and planar rotation introduces some (< 2 degrees) random error.

Drug therapy of bacillus Calmette-Guérin sepsis.

Intravesical bacillus Calmette-Guérin (BCG) is widely used for the treatment of transitional cell carcinoma of the bladder. Although it is usually well tolerated, sepsis can occur, which has resulted in at least eight deaths [3]. The survival of Connaught BCG-infected mice treated with single and combination antibiotic and steroid therapy was evaluated. Triple-drug therapy with isoniazid, rifampin, and prednisolone resulted in 53% survival compared with 25% survival in the control group (P = 0.0209). A survival of only 10.5% was observed with treatment using prednisolone alone. This survival was worse than that of the control group (25%), and approached statistical significance (P = 0.0669). Our data suggest that BCG sepsis probably has components of both a hypersensitivity reaction and bacterial sepsis; they support the current use of combination antibiotic and steroid therapy for treatment of BCG sepsis in humans, but argue against treatment with steroids alone.

Megadose vitamins in bladder cancer: a double-blind clinical trial.

Epidemiological and laboratory studies suggest that vitamin supplements may be helpful in the prevention of some cancers but clinical trials to date have failed to demonstrate protection with naturally occurring vitamins. Without substantiation of the highly touted benefits of vitamins, few physicians who care for cancer patients have recommended their use. A total of 65 patients with biopsy confirmed transitional cell carcinoma of the bladder enrolled in a randomized comparison of intravesical bacillus Calmette-Guerin (BCG) with or without percutaneous administration was also randomized by closed envelope to therapy with multiple vitamins in the recommended daily allowance (RDA) versus RDA multivitamins plus 40,000 units vitamin A, 100 mg. vitamin B6, 2,000 mg. vitamin C, 400 units vitamin E and 90 mg. zinc. The addition of percutaneous BCG did not significantly lessen tumor recurrence but recurrence after 10 months was markedly reduced in patients receiving megadose vitamins. The 5-year estimates of tumor recurrence are 91% in the RDA arm and 41% in the megadose arm (p = 0.0014, Mantel-Cox). Overall recurrence was 24 of 30 patients (80%) in the RDA arm and 14 of 35 (40%) in the high dose arm (p = 0.0011, 2-tailed Fisher's exact test). Megadose vitamins A, B6, C and E plus zinc decrease bladder tumor recurrence in patients receiving BCG immunotherapy. Further research will be required to identify which ingredient(s) provide this protection.

Immunotherapy of murine bladder cancer with keyhole limpet hemocyanin (KLH).

Keyhole limpet hemocyanin (KLH) is a potent immunogen that is being evaluated as an immunotherapeutic alternative to BCG in the treatment of bladder cancer. In the mouse bladder tumor model (MBT2) intralesional KLH significantly reduced tumor incidence, growth rate, and mortality and exhibited antitumor activity similar to that achievable with BCG. Endotoxin contamination of KLH was not responsible for the antitumor activity, although endotoxin alone was shown to have anti-tumor activity in this animal model. Keyhole limpet hemocyanin is both safe and effective in the MBT2 model, and is an immunomodulator to consider for clinical trials.

Keyhole limpet hemocyanin immunotherapy of murine bladder cancer.

The current treatment of choice for superficial bladder cancer, bacillus Calmette-Guérin, has significant adverse side effects. We have compared two alternative immunotherapies--crude keyhole limpet hemocyanin (KLH) and Immucothel, a KLH modified for clinical use (Biosyn)--in an intralesional mouse model of bladder cancer (MBT2). Crude KLH required either immunization before tumor transplant or frequent intralesional therapy after transplantation to be effective. In addition, Immucothel required pre-immunization to be effective, and increasing the frequency and dosage of post-transplant immunization was not effective without pre-immunization. Preliminary investigations into the KLH-induced anti-tumor mechanism(s) suggest that natural killer cell activity may be involved. Both crude KLH and Immucothel appear to be effective immunotherapies of use in the treatment of transitional cell carcinoma.

Antibiotic and steroid therapy of massive systemic bacillus Calmette-Guerin toxicity.

Intravesical bacillus Calmette-Guerin (BCG) is the most effective treatment of carcinoma in situ available today and is superior to chemotherapy in the prevention of bladder tumor recurrence. While therapy is generally well tolerated, serious and even life threatening toxicity can occur. Treatment options for serious infection include isoniazid, rifampin, ethambutol, and cycloserine, but shock may also be secondary to hypersensitivity and require the addition of corticosteroids. The morbidity and mortality of systemically BCG-infected mice treated with single and combined antimicrobial and/or corticosteroid therapies was evaluated. BCG immunized mice were unable to survive doses of BCG which were uniformly tolerated in naive mice. The addition of cycloserine increased survival in mice treated with isoniazid and rifampin, but optimal survival was achieved with isoniazid, rifampin, and prednisolone. These experimental results support the previously reported clinical success of isoniazid, rifampin and prednisolone in patients with septic BCG reactions.

Immunotherapy of murine transitional cell carcinoma of the bladder using alpha and gamma interferon in combination with other forms of immunotherapy.

BCG immunotherapy is very effective in the treatment of superficial transitional cell carcinoma of the bladder, but its significant toxicity may limit its use in some patients. In an effort to find less toxic and potentially more effective treatments we investigated the possible immunotherapeutic potential of combinations of Alpha Interferon (1000 IU) and Gamma Interferon (500 IU) with bacillus Calmette Guerin (BCG) (10(7) cfu), Interleukin-2 (4000 IU), and Keyhole Limpet Hemocyanin (50 micrograms.) in the MBT2 murine bladder cancer model. Significant reductions (p less than 0.05) in tumor incidence relative to the saline control, 83%, Day 35) was observed in groups receiving alpha interferon (42%), Keyhole limpet hemocyanin (42%), interleukin-2 (25%), alpha interferon + Keyhole limpet hemocyanin (17%), alpha interferon + interleukin-2 (33%), gamma interferon + BCG (42%), and gamma interferon + interleukin-2 (17%). All treatment groups with the exception of the group receiving gamma interferon had significantly reduced tumor volume (p less than 0.05) relative to the saline control. Combination treatment groups were significantly more effective than single agent treatments (p = 0.0057). The exhibited anti-tumor effect of these immunotherapeutic agents alone and in combination suggest that they may prove to be effective forms of immunotherapy for transitional cell carcinoma of the bladder.

Treatment of murine transitional cell carcinoma with intralesional interleukin 2 and murine interferon gamma.

The antitumor effect of intralesionally administered recombinant interleukin-2, alone or in combination with recombinant interferon gamma was studied in murine transitional cell carcinoma, MBT2. In the initial prophylactic model treatment was started at day one at the site of tumor inoculation. Maximal and significant reduction in tumor volume occurred in groups receiving 4,000 units of recombinant interleukin 2 and 10(7) colony forming units Bacillus Calmette Guerin (p less than 0.00001 vs saline control). In the same experiment, a reduction in tumor incidence and increase in survival occurred in groups receiving 4,000 units of recombinant interleukin 2, 1,000 units of recombinant interleukin 2 plus 2,000 units of recombinant interferon gamma, as well as 10(7) colony forming units Bacillus Calmette Guerin relative to saline control (p less than 0.005). The dose-response effect of recombinant interleukin 2 alone was also tested in a model of an established transitional cell carcinoma. Intralesional injection treatments were initiated after tumors were palpable. Reduction in tumor volume was observed in the group receiving 8,000 units of recombinant interleukin 2 (p = 0.01 vs saline control), but no significant advantage in survival was noted.