Search

PURPOSE This Provisional Clinical Opinion update presents a clinically pragmatic approach to hepatitis B virus (HBV) screening and management. PROVISIONAL CLINICAL OPINION All patients anticipating systemic anticancer therapy should be tested for HBV by 3 tests—hepatitis B surface antigen (HBsAg), hepatitis B core antibody (anti-HBc) total immunoglobulin (Ig) or IgG, and antibody to hepatitis B surface antigen—but anticancer therapy should not be delayed. Findings of chronic HBV (HBsAg-positive) or past HBV (HBsAg-negative and anti-HBc–positive) infection require HBV reactivation risk assessment. Patients with chronic HBV receiving any systemic anticancer therapy should receive antiviral prophylactic therapy through and for minimum 12 months following anticancer therapy. Hormonal therapy alone should not pose a substantial risk of HBV reactivation in patients with chronic HBV receiving hormonal therapy alone; these patients may follow noncancer HBV monitoring and treatment guidance. Coordination of care with a clinician experienced in HBV management is recommended for patients with chronic HBV to determine HBV monitoring and long-term antiviral therapy after completion of anticancer therapy. Patients with past HBV infection undergoing anticancer therapies associated with a high risk of HBV reactivation, such as anti-CD20 monoclonal antibodies or stem-cell transplantation, should receive antiviral prophylaxis during and for minimum 12 months after anticancer therapy completion, with individualized management thereafter. Careful monitoring may be an alternative if patients and providers can adhere to frequent, consistent follow-up so antiviral therapy may begin at the earliest sign of reactivation. Patients with past HBV undergoing other systemic anticancer therapies not clearly associated with a high risk of HBV reactivation should be monitored with HBsAg and alanine aminotransferase during cancer treatment; antiviral therapy should commence if HBV reactivation occurs. Additional information is available at www.asco.org/supportive-care-guidelines .

Checkpoint inhibitors have become standard therapy for multiple cancers, and their use will increase in the next year as regulatory approvals for additional indications are expected. It is essential for clinicians to be aware of the potential for rare immune-related adverse effects. Here, we report the case of a new diagnosis of myasthenia gravis (MG) after the use of nivolumab for squamous cell carcinoma of the bladder. A review the literature identified 10 cases of MG diagnosed after programmed cell death protein 1 inhibitor therapy. This is the first case, to our knowledge, reported in association with bladder cancer. The precise diagnosis of MG has important implications on management, as treatment with steroids can transiently worsen myasthenia in nearly 50% of cases.

Nivolumab is a standard treatment option in several advanced malignancies, but safety and efficacy are still unknown in patients with human immunodeficiency virus (HIV) infection. We describe a case series of people living with HIV (PLWH) receiving nivolumab in the Veterans Health Administration (VA) and report responses and toxicities. We identified all PLWH who received nivolumab at any VA facility since 2000 in the Corporate Data Warehouse (CDW), which provides nationwide research access to VA electronic medical records. We identified 16 HIV-infected nivolumab recipients. The median number of nivolumab doses received was 6 (range, 1-32). Changes in CD4 count during therapy were variable, with 70% (7/10) of patients experiencing increases. Half of PLWH were treated for non-small-cell lung cancer; 2 for Hodgkin lymphoma (HL), 2 for renal cell carcinoma, and 4 for off-label cancers. For non-small-cell lung cancer, 7 patients had evaluable responses. Although 5 of 7 patients immediately progressed, 1 had a partial response and 1 had stable disease, which were both durable. Two of 16 (14%) PLWH had complete responses; both with HL (2/2 HL, 100%). The prevalence of immune-related adverse effects was 40% overall (6/15); 27% (4/15) had pneumonitis. To our knowledge, this is the largest case series reporting outcomes with nivolumab in PLWH. Outcomes were comparable with those seen in studies of HIV-uninfected patients, and particularly interesting for HL. The reason for the high proportions of immune-related adverse effects is unclear, but needs to be confirmed in larger studies.

Hepatocellular carcinomas (HCC) show resistance to chemotherapy and have blunt response to apoptotic stimuli. HCC cell lines express low levels of the Fas death receptor and are resistant to FasL stimulation, whereas immortalized hepatocytes are sensitive. The variable Fas transcript levels and consistently low Fas protein in HCC cells suggest posttranscriptional regulation of Fas expression. The 3′-untranslated region (UTR) of Fas mRNA was found to interact with the ribonucleoprotein Human Antigen R (HuR) to block mRNA translation. Silencing of HuR in HCC cells increased the levels of cell surface Fas and sensitized HCC cells to FasL. Two AU-rich domains within the 3′-UTR of Fas mRNA were identified as putative HuR-binding sites and were found to mediate the translational regulation in reporter assay. Hydrodynamic transfection of HuR plasmid into mice induced downregulation of Fas expression in livers and established functional resistance to the killing effects of Fas agonist. Human HCC tumor tissues showed significantly higher overall and cytoplasmic HuR staining compared with normal liver tissues, and the high HuR staining score correlated with worse survival of patients with early-stage HCC. Combined, the protumorigenic ribonucleoprotein HuR blocks the translation of Fas mRNA and effectively prevents Fas-mediated apoptosis in HCC, suggesting that targeting HuR would sensitize cells to apoptotic stimuli and reverse tumorigenic properties. Implications: Demonstrating how death receptor signaling pathways are altered during progression of HCC will enable the development of better methods to restore this potent apoptosis mechanism. Mol Cancer Res; 13(5); 809–18. ©2015 AACR.

Purpose: We investigated the effect of the mTOR inhibitor RAD001 (everolimus) on human bladder cancer (BC) cells in vitro and in vivo. Experimental Design: The effect of RAD001 on the growth of UM-UC-3, UM-UC-6, UM-UC-9, and UM-UC-14 BC cells were assessed by crystal violet and [3H]thymidine incorporation assays. Flow cytometric cell-cycle analyses were done to measure the apoptotic cell fraction. Protein synthesis was measured using tritium-labeled leucine incorporation assays. The effects of RAD001 on the mTOR pathway were analyzed by Western blotting. To test the effects of RAD001 in vivo, UM-UC-3, UM-UC-6, and UM-UC-9 cells were subcutaneously implanted into nude mice. Tumor-bearing mice were treated orally with RAD001 or placebo. Tumors were harvested for immunohistochemical analysis. Results: In vitro, RAD001 transiently inhibited BC cell growth in a dose-dependent manner. This effect was augmented by re-treatment of cells after 3 days. UM-UC-14 cells were the most sensitive to RAD001, whereas UM-UC-9 cells were the least sensitive. After re-treatment with RAD001, only sensitive cell lines showed G1-phase arrest, with no evidence of apoptosis. RAD001 significantly inhibited the growth of tumors that were subcutaneously implanted in mice. Inhibition of protein synthesis through the S6K and 4EBP1 pathways seems to be the main mechanism for the RAD001-induced growth inhibition. However, inhibition of angiogenesis was the predominant mechanism of the effect of RAD001 on UM-UC-9 cells. Conclusions: The mTOR inhibitor RAD001 inhibits growth of BC cells in vitro. RAD001 is effective in treating BC tumors in an in vivo nude mouse model despite the heterogeneity of in vitro responses. Clin Cancer Res; 17(9); 2863–73. ©2011 AACR.

Purpose An American Society of Clinical Oncology (ASCO) provisional clinical opinion (PCO) offers timely clinical direction to ASCO's membership following publication or presentation of potentially practice-changing information. This PCO addresses recommendations for chronic hepatitis B virus (HBV) infection screening in patients receiving cytotoxic or immunosuppressive chemotherapy for treatment of malignant diseases. Clinical Context The Centers for Disease Control and Prevention (CDC) issued Recommendations for Identification and Public Health Management of Persons with Chronic Hepatitis B Virus Infection, recommending screening for hepatitis B infection (hepatitis B surface antigen [HBsAg], antihepatitis B core antigen [anti-HBc], and antibodies to HBsAg [anti-HBs]) for “persons receiving cytotoxic or immunosuppressive therapy (eg, chemotherapy for malignant diseases…).” Provisional Clinical Opinion The evidence is insufficient to determine the net benefits and harms of routine screening for chronic HBV infection in individuals with cancer who are about to receive cytotoxic or immunosuppressive therapy or who are already receiving therapy. Individuals with cancer who undergo certain cytotoxic or immunosuppressive therapies and have HBV infection or prior exposure to HBV may be at elevated risk of liver failure from HBV reactivation. As such, HBV screening requires clinical judgment. Physicians may consider screening patients belonging to groups at heightened risk for chronic HBV infection or if highly immunosuppressive therapy is planned. Highly immunosuppressive treatments include, but are not limited to, hematopoietic cell transplantation and regimens including rituximab. Screening based on a high risk of prior HBV exposure or risk of reactivation due to planned therapeutic regimens should include testing for HBsAg as a serologic marker for HBV infection. In some populations, testing for anti-HBc should also be considered. There is no evidence to support serologic testing for anti-HBs in this context. When evidence for chronic HBV infection is found, antiviral therapy before and throughout the course of chemotherapy may be considered to reduce the risk of HBV reactivation, although evidence from controlled trials of this approach is limited. Screening and/or treating HBV infection should not delay the initiation of chemotherapy. NOTE. ASCO's provisional clinical opinions (PCOs) reflect expert consensus based on clinical evidence and literature available at the time they are written, and are intended to assist physicians in clinical decision-making and identify questions and settings for further research. Due to the rapid flow of scientific information in oncology, new evidence may have emerged since the time a PCO was submitted for publication. PCOs are not continually updated and may not reflect the most recent evidence. PCOs address only the topics specifically identified in the PCO and are not applicable to interventions, diseases or stages of disease not specifically identified. PCOs cannot account for individual variation among patients, and cannot be considered inclusive of all proper methods of care or exclusive of other treatments. It is the responsibility of the treating physician or other health care provider, relying on independent experience and knowledge of the patient, to determine the best course of treatment for the patient. Accordingly, adherence to any PCO is voluntary, with the ultimate determination regarding its application to be made by the physician in light of each patient's individual circumstances. ASCO PCOs describe the use of procedures and therapies in clinical practice and cannot be assumed to apply to the use of these interventions in the context of clinical trials. ASCO assumes no responsibility for any injury or damage to persons or property arising out of or related to any use of ASCO's PCOs, or for any errors or omissions.

Transitional cell carcinoma (TCC) of the bladder ranks fourth in incidence of all cancers in the developed world, yet the mechanisms of its origin and progression remain poorly understood. There are also few useful diagnostic or prognostic biomarkers for this disease. We have combined a transgenic mouse model for invasive bladder cancer (UPII-SV40Tag mice) with DNA microarray technology to determine molecular mechanisms involved in early TCC development and to identify new biomarkers for detection, diagnosis, and prognosis of TCC. We have identified genes that are differentially expressed between the bladders of UPII-SV40Tag mice and their age-matched wild-type littermates at 3, 6, 20, and 30 weeks of age. These are ages that correspond to premalignant, carcinoma in situ, and early-stage and later stage invasive TCC, respectively. Our preliminary analysis of the microarray data sets has revealed ∼1,900 unique genes differentially expressed (≥3-fold difference at one or more time points) between wild-type and UPII-SV40Tag urothelium during the time course of tumor development. Among these, there were a high proportion of cell cycle regulatory genes and a proliferation signaling genes that are more strongly expressed in the UPII-SV40Tag bladder urothelium. We show that several of the genes upregulated in UPII-SV40Tag urothelium, including RacGAP1, PCNA, and Hmmr, are expressed at high levels in superficial bladder TCC patient samples. These findings provide insight into the earliest events in the development of bladder TCC as well as identify several promising early-stage biomarkers. Cancer Prev Res; 3(6); 776–86. ©2010 AACR.

Purpose: The study aims to evaluate the efficacy and toxicity of fenretinide in preventing tumor recurrence in patients with transitional cell carcinoma (TCC) of the bladder. Experimental Design: We conducted a multicenter phase III, randomized, placebo-controlled trial of fenretinide (200 mg/day orally for 12 months) in patients with non–muscle-invasive bladder TCC (stages Ta, Tis, or T1) after transurethral resection with or without adjuvant intravesical Bacillus Calmette-Guerin (BCG). Patients received cystoscopic evaluation and bladder cytology every 3 months during the 1-year on study drug and a final evaluation at 15 months. The primary endpoint was time to recurrence. Results: A total of 149 patients were enrolled; 137 were evaluable for recurrence. The risk of recurrence was considered to be “low” in 72% (no prior BCG) and intermediate or high in 32% (prior BCG) of the evaluable patients. Of the lower-risk group, 68% had solitary tumors and 32% had multifocal, low-grade papillary (Ta, grade 1 or grade 2) tumors. The 1-year recurrence rates by Kaplan-Meier estimate were 32.3% (placebo) versus 31.5% (fenretinide; P = 0.88 log-rank test). Fenretinide was well tolerated and had no unexpected toxic effects; only elevated serum triglyceride levels were significantly more frequent on fenretinide (versus placebo). The Data Safety and Monitoring Board recommended study closure at 149 patients (before reaching the accrual goal of 160 patients) because an interim review of the data showed a low likelihood of detecting a difference between the two arms, even if the original accrual goal was met. Conclusions: Although well tolerated, fenretinide did not reduce the time-to-recurrence in patients with Ta, T1, or Tis TCC of the bladder.

Purpose To conduct a phase III trial of adjuvant 13-cis-retinoic acid (13cRA) plus interferon alfa (IFN-α) for preventing tumor recurrence and second primary tumors (SPTs) of skin squamous cell carcinoma (SCC) among patients with aggressive skin SCC. Patients and Methods Sixty-six patients with aggressive skin SCC were randomly assigned to receive either 6 months of combined 13cRA (1 mg/kg/d orally) and IFN-α (3 × 106 U subcutaneously three times per week) or no adjuvant therapy (control group) after SCC surgery and/or radiation. Results At 21.5 months median follow-up, treatment did not improve the time to tumor recurrence and SPT versus control (hazard ratio [HR], 1.13; 95% CI, 0.53 to 2.41), time to tumor recurrence (HR, 1.08; 95% CI, 0.43 to 2.72), or time to SPT (HR, 0.89; 95% CI, 0.27 to 2.93). Adjuvant 13cRA and IFN-α was moderately tolerable; 29% of patients in the treatment arm required dose reductions for grade 3 or 4 toxicities. Conclusion Results of this phase III trial do not support 13cRA plus IFN-α for adjuvant therapy of aggressive skin SCC. With high rates of tumor recurrence and SPTs, patients with aggressive skin SCC continue to have an unmet medical need, with devastating mortality, morbidity, and financial consequences. Promising agents with preclinical and early clinical results relevant to aggressive skin SCC deserve a high priority for future clinical drug development.

Purpose This updated provisional clinical opinion presents a revised opinion based on American Society of Clinical Oncology panel consensus in the context of an evolving database. Context Despite the 2010 provisional clinical opinion recommendation, there is still evidence of suboptimal hepatitis B virus (HBV) screening among patients at high risk for HBV infection or HBV reactivation after chemotherapy. This updated provisional clinical opinion introduces a risk-adaptive strategy to identify and treat patients with HBV infection to reduce their risk of HBV reactivation. Provisional Clinical Opinion Medical providers should screen by testing patients for HBV infection before starting anti-CD20 therapy or hematopoietic cell transplantation. Providers should also screen patients with risk factors for HBV infection. Screening should include both hepatitis B surface antigen (HBsAg) and hepatitis B core antibody (anti-HBc), because reactivation can occur in patients who are HBsAg positive/anti-HBc positive or HBsAg negative/anti-HBc positive. Either total anti-HBc or anti-HBc immunoglobulin G (not immunoglobulin M) test should be used. Clinicians should start antiviral therapy for HBsAg-positive/anti-HBc–positive patients before or contemporaneously with cancer therapy and monitor HBsAg-negative/anti-HBc–positive patients for reactivation with HBV DNA and ALT levels, promptly starting antivirals if reactivation occurs. Clinicians can initiate antivirals for HBsAg-negative/anti-HBc–positive patients anticipating cancer therapies associated with a high risk of reactivation, or they can monitor HBV DNA and ALT levels and initiate on-demand antivirals. For patients who neither have HBV risk factors nor anticipate cancer therapy associated with a high risk of reactivation, current evidence does not support HBV screening before initiation of cancer therapy. Two panel members provided a minority viewpoint, involving a strategy of universal HBsAg and selective anti-HBc testing.

To compare the survival of women and men with transitional cell bladder cancer.We used the Surveillance Epidemiology and End Results database to identify patients aged 35 years or older diagnosed with bladder cancer between 1991 and 2001 actively followed up. We excluded cases diagnosed by autopsy or death certificates and those of unknown race. We used Cox proportional hazard regression to analyze survival in patients with advanced disease.Of the 31,009 patients meeting eligibility criteria, 26.7% were women. Median age at diagnosis for women and men was 72 and 70 years, respectively. Regional disease was diagnosed in 20.3% of white women and 35.5% of African-American women, compared with only 17.6% of white men and 25.9% of African-American men (P.001). Increased age, African-American race, and being female significantly increased the hazard of death (hazard ratio [HR] 1.037, 95% confidence interval [CI] 1,034-1.041; HR 1.402, 95% CI 1.187-1.656; and HR 1.842, 95% CI 1.158-2.931).Women with bladder cancer, particularly African-Americans, have shorter survival. This is partially explained by higher risk of diagnosis with poorly differentiated tumors, advanced stage, and advanced age. Women should be targeted for timely diagnosis.II-2.

An inverse relationship exists between the expression of 15-lipoxygenase-2 (15-LOX-2) and peroxisome proliferator-activated receptor gamma (PPARgamma) in normal prostate epithelial cells (PrECs) compared with their expression in prostate carcinoma cells (PC-3). The reason for this difference, however, is unknown. We hypothesized that this inverse expression partly involves the 15-LOX-2 promoter and 15-S-hydroxyeicosatetraenoic acid (15-(S)-HETE), a product of 15-LOX-2 that binds to PPARgamma. We identified an active steroid nuclear receptor half-site present in the 15-LOX-2 promoter fragment F-5 (-618/+177) that can interact with PPARgamma. After forced expression of wild-type PPARgamma, 15-(S)-HETE (1 microM) decreased F-5 reporter activity in PrECs whereas forced expression of 15-LOX-2 resulted in 15-(S)-HETE production which enhanced F-5 activity in PC-3. In contrast, the expression of dominant-negative PPARgamma reversed the transcriptional activation of F-5 by enhancing it 202-fold in PrEC or suppressing it in PC-3; the effect in PC-3 was positively increased 150-fold in the presence of 15-(S)-HETE (1 microM). Peroxisome proliferator-activated receptor gamma interacted with 15-LOX-2 promoter sequences in pulldown experiments using biotinylated 15-LOX-2 (-560/-596 bp) oligonucleotides. In gelshift analyses PPARgamma and orphan receptor RORalpha were shown to interact with the F-5 fragment in PC-3 cells. These data suggest that crosstalk mechanisms exist between the 15-LOX-2 gene and PPARgamma to counterbalance expression and help explain the inverse relationship of these genes in normal versus cancer cells.

Stromal-epithelial interactions and the bioactive molecules produced by these interactions maintain tissue homeostasis and influence carcinogenesis. Bioactive prostaglandins produced by prostaglandin synthases and secreted by the prostate into seminal plasma are thought to support reproduction, but their endogenous effects on cancer formation remain unresolved. No studies to date have examined prostaglandin enzyme production or prostaglandin metabolism in normal prostate stromal cells. Our results show that lipocalin-type prostaglandin D synthase (L-PGDS) and prostaglandin D2 (PGD2) metabolites produced by normal prostate stromal cells inhibited tumor cell growth through a peroxisome proliferator–activated receptor γ (PPARγ)–dependent mechanism. Enzymatic products of stromal cell L-PGDS included high levels of PGD2 and 15-deoxy-Δ12,14-PGD2 but low levels of 15-deoxy-Δ12,14-prostaglandin J2. These PGD2 metabolites activated the PPARγ ligand-binding domain and the peroxisome proliferator response element reporter systems. Thus, growth suppression of PPARγ-expressing tumor cells by PGD2 metabolites in the prostate microenvironment is likely to be an endogenous mechanism involved in tumor suppression that potentially contributes to the indolence and long latency period of this disease.

Background: Evidence of the chemopreventive effects of the dietary antioxidants α-tocopherol (vitamin E) and l-selenomethionine (selenium) comes from secondary analysis of two phase III clinical trials that found treatment with these antioxidants reduced the incidence of prostate cancer. To determine the effects of selenium and vitamin E in blood and prostate tissue, we undertook a preoperative feasibility study complementary to the currently ongoing Selenium and Vitamin E Cancer Prevention Trial.Methods: Forty-eight patients with clinically localized prostate cancer enrolled on this 2 × 2 factorial design study were randomized to take selenium, vitamin E, both, or placebo for 3 to 6 weeks before prostatectomy. Sera were collected from patients before and after dietary supplementation. Thirty-nine patients were evaluable, and 29 age-matched disease-free men served as controls. Mass profiling of lipophilic serum proteins of lower molecular weight (2-13.5 kDa) was conducted, and mass spectra data were analyzed using custom-designed software.Results: Weighted voting analyses showed a change in sera classification from cancerous to healthy for some patients with prostate cancer after dietary intervention. ANOVA analysis showed significantly different treatment effects on prediction strength changes among the four groups at a 95% confidence level. Eliminating an outlying value and performing post hoc analysis using Fisher's least significant difference method showed that effects in the group treated with the combination were significantly different from those of the other groups.Conclusion: In sera from patients with prostate cancer, selenium and vitamin E combined induced statistically significant proteomic pattern changes associated with prostate cancer–free status.

The inducible form of cyclooxygenase (COX), COX-2, is up-regulated in many epithelial cancers and its prostaglandin products increase proliferation, enhance angiogenesis, and inhibit apoptosis in several tissues. Pharmacologic inhibition and genetic deletion studies showed a marked reduction of tumor development in colon and skin. COX-2 has also been strongly implicated in urinary bladder cancer primarily by studies with nonselective COX- and COX-2-selective inhibitors. We now show that forced expression of COX-2, under the control of a keratin 5 promoter, is sufficient to cause transitional cell hyperplasia (TCH) in 17% and 75% of the heterozygous and homozygous transgenic lines, respectively, in an age-dependent manner. TCH was strongly associated with inflammation, primarily nodules of B lymphocytes; some T cells and macrophage infiltration were also observed. Additionally, transitional cell carcinoma was observed in ∼10% of the K5.COX-2 transgenic mice; no TCH or transitional cell carcinoma was observed in wild-type bladders. Immunohistochemistry for vascular proliferation and vascular endothelial growth factor showed significant increases above that in wild-type urinary bladders. Our results suggest that overexpression of COX-2 is sufficient to cause hyperplasia and carcinomas in the urinary bladder. Therefore, inhibition of COX-2 should continue to be pursued as a potential chemopreventive and therapeutic strategy.

Supported by NIH Specialized Programs of Research Excellence Grant in Genitourinary Cancer CA91846, and National Cancer Institute Grants CA16672, CA066723, and CA85078.

Selective cyclooxygenase (COX)-2-inhibiting and other nonsteroidal anti-inflammatory drugs (NSAIDs) show promise for preventing and treating bladder and prostate cancers. In contrast to the strong NSAID epidemiology in colorectal cancer, the epidemiologic data on NSAIDs and genitourinary (GU) cancers are limited and mixed. However, a substantial body of preclinical in vitro and in vivo animal model data shows consistent NSAID activity in treating, and in some cases preventing, GU cancers and begins to address the mechanisms behind this activity (eg, involving Akt and ERK2 in the prostate). Many preclinical and clinical NSAID studies currently under way are helping to resolve the best type (selective or nonselective COX inhibitors or non-COX inhibitors), dose and duration of NSAID treatment for prevention in the GU setting. Future studies likely will focus on clarifying the NSAID mechanisms behind and developing NSAID combinations for both treating and preventing GU cancers.

Kinases can phosphorylate and regulate androgen receptor activity during prostate cancer progression. In particular, we showed that glycogen synthase kinase-3 beta phosphorylates the androgen receptor, thereby inhibiting androgen receptor-driven transcription. Conversely, the glycogen synthase kinase-3 beta inhibitor lithium chloride suppressed the glycogen synthase kinase-3 beta-mediated phosphorylation of the androgen receptor, thereby enabling androgen receptor-driven transcription to occur. The androgen receptor hinge and ligand-binding domains were important for both the phosphorylation and the inhibition of transcriptional activity of the receptor by glycogen synthase kinase-3 beta. Furthermore, androgen receptor phosphorylation was augmented by LY294002, an indirect inhibitor of protein kinase B/Akt that inhibits glycogen synthase kinase-3 beta. We also showed that the mutation of various phosphorylation sites on glycogen synthase kinase-3 beta affected the ability of these mutants to co-distribute with the androgen receptor in the cell nucleus, also that both glycogen synthase kinase-3beta and androgen receptor proteins can be found in cell nuclei of prostate cancer tissue samples. Because glycogen synthase kinase-3 beta activity is suppressed after the enzyme is phosphorylated by protein kinase B/Akt and Akt activity frequently increases during the progression of prostate cancer, nullification of the glycogen synthase kinase-3 beta-mediated suppression of androgen receptor activity by Akt likely contributes to prostate cancer progression.

Patients who have bladder cancer with superficial disease are at high risk for recurrence but low risk for progression to muscle invasive disease and are regularly monitored with cystoscopy to detect recurrent tumors. This clinical setting (high recurrence rate, excellent surveillance, and low risk of progression in stage) provides an ideal setting for secondary chemoprevention. Vitamins, difluoromethylornithine, and cyclooxygenase inhibitors have demonstrated activity in preclinical models of bladder cancer. Limited clinical data suggests that vitamins and their analogs may be useful for decreasing the rate of tumor recurrence in patients with superficial bladder cancer. Ongoing clinical trials are exploring the roles of fenretinide, difluoromethylornithine, and celecoxib in the treatment of patients with a history of superficial bladder cancer.

Bladder cancer is strongly related to tobacco use and is estimated to cause 54,500 new cancer cases and 11,700 deaths in the United States in 1998. Approximately two thirds of new US cases will be superficial tumors, predominantly low-grade papillary. After standard therapeutic resection (with or without intravesical therapy), the superficial bladder tumor recurrence rate is 30% to 70% within 12 months of resection. Morbidity is substantial, with frequent cystoscopy, recurrence, resections, and possible cystectomy for progression to invasive cancers. Therefore, new approaches, including chemoprevention, are needed. Data suggest that bladder carcinogenesis is a multi-step, multifocal (field effect) process, possibly involving the spread of premalignant clones--all of which are prerequisites for effective chemopreventive approaches. To date, retinoids are the best-studied chemopreventive agents in this site, achieving mixed clinical results (with 13-cis-retinoic acid and etretinate) in superficial bladder tumors. This review includes the epidemiology and biology of bladder carcinogenesis in addition to preclinical and clinical retinoid data, and focuses on the most promising avenue of current retinoid chemoprevention in the bladder: the potent apoptosis-inducing retinoid fenretinide (4-HPR), which currently is in three phase III trials.

Background: The synthetic retinoid N-(4-hydroxyphenyl) retinamide (4HPR) can inhibit the growth of tumor cells. Preliminary results from a clinical trial suggest that 4HPR may reduce ovarian cancer incidence. We examined the growth-inhibitory effects of 4HPR on gynecologic cancer cell lines in vitro and the role of retinoid receptors in modulating this effect. Methods: Twelve human gynecologic cancer cell lines (the ovarian cell lines-A224, AD10, UCI 101, UCI 107, SKOV3, 222, CP70, ML3B, and ML5; the cervical cell lines-HT3 and ME180; and the endometrial cell line-Hec 1A were tested for sensitivity to 4HPR (by assaying cell proliferation rates). Gel electrophoretic analysis of DNA fragmentation was used to measure programmed cell death (apoptosis). Specific retinoid receptor (retinoic acid receptor [RAR] and retinoid X receptor) messenger RNA (mRNA) levels were measured by northern blot hybridization. AD10 cells were stably transfected with human RARβ complementary DNA, and the effect of 4HPR on cell proliferation was examined. Results: 4HPR inhibited the growth of all 12 cell lines, but to varying degrees; Ic 50 values (i.e., concentrations that inhibit proliferation by 50%) ranged from 0.3 to 9 μM. Following 4HPR treatment, ovarian cancer cells that were sensitive to 4HPR (222, CP70, and UCI 101; Ic 50

Resistance to Fas-mediated apoptosis is associated with poor cancer outcomes and chemoresistance. To elucidate potential mechanisms of defective Fas signaling, we screened primary lymphoma cell extracts for Fas-associated proteins that would have the potential to regulate Fas signaling. An activation-resistant Fas complex selectively included nucleolin. We confirmed the presence of nucleolin-Fas complexes in B-cell lymphoma cells and primary tissues, and the absence of such complexes in B-lymphocytes from healthy donors. RNA-binding domain 4 and the glycine/arginine-rich domain of nucleolin were essential for its association with Fas. Nucleolin colocalized with Fas on the surface of B-cell lymphoma cells. Nucleolin knockdown sensitized BJAB cells to Fas ligand (FasL)-induced and Fas agonistic antibody-induced apoptosis through enhanced binding, suggesting that nucleolin blocks the FasL–Fas interaction. Mice transfected with nucleolin were protected from the lethal effects of agonistic anti-mouse Fas antibody (Jo2) and had lower rates of hepatocyte apoptosis, compared with vector and a non-Fas-binding mutant of nucleolin. Our results show that cell surface nucleolin binds Fas, inhibits ligand binding, and thus prevents induction of Fas-mediated apoptosis in B-cell lymphomas and may serve as a new therapeutic target.

Lung cancer is a major cause of mortality in the United States and accounts for the majority of all cancer deaths in both men and women. It is hoped that through broadening our understanding of the mechanisms involved in transformation of bronchial epithelial cells we will be able to improve methods of diagnosis and treatment of this disease, with the ultimate goal of reducing on lung cancer mortality. A knowledge of the molecular mechanisms involved in processes such as cell division and differentiation is paramount to this task, because it is known that aberrant responses to growth factors or cytokines found in the normal cellular milieu can lead to abnormal cell growth and/or transformation. Signals initiated at the cell membrane by tumor promoters, growth factors, or cytokines are transduced from the cell membrane to the nucleus and are, in part, mediated centrally by transcription factors encoded by nuclear protooncogenes. The transcription factors myc, jun, and fos have been characterized in both normal and transformed lung epithelial cells through detailed studies using cell lines. In this manuscript, we review what is known about the expression and regulation of these nuclear protooncogenes in normal and malignant epithelial cells of the lung, and their role in the development of lung cancer.

Context Lung cancer is the leading cause of cancer death. Most patients are diagnosed with advanced disease, resulting in a very low 5-year survival. Screening may reduce the risk of death from lung cancer. Objective To conduct a systematic review of the evidence regarding the benefits and harms of lung cancer screening using low-dose computed tomography (LDCT). A multisociety collaborative initiative (involving the American Cancer Society, American College of Chest Physicians, American Society of Clinical Oncology, and National Comprehensive Cancer Network) was undertaken to create the foundation for development of an evidence-based clinical guideline. Data Sources MEDLINE (Ovid: January 1996 to April 2012), EMBASE (Ovid: January 1996 to April 2012), and the Cochrane Library (April 2012). Study Selection Of 591 citations identified and reviewed, 8 randomized trials and 13 cohort studies of LDCT screening met criteria for inclusion. Primary outcomes were lung cancer mortality and all-cause mortality, and secondary outcomes included nodule detection, invasive procedures, follow-up tests, and smoking cessation. Data Extraction Critical appraisal using predefined criteria was conducted on individual studies and the overall body of evidence. Differences in data extracted by reviewers were adjudicated by consensus. Results Three randomized studies provided evidence on the effect of LDCT screening on lung cancer mortality, of which the National Lung Screening Trial was the most informative, demonstrating that among 53 454 participants enrolled, screening resulted in significantly fewer lung cancer deaths (356 vs 443 deaths; lung cancer−specific mortality, 247 vs 309 events per 100 000 person-years for LDCT and control groups, respectively; relative risk, 0.80; 95% CI, 0.73-0.93; absolute risk reduction, 0.33%; P = .004). The other 2 smaller studies showed no such benefit. In terms of potential harms of LDCT screening, across all trials and cohorts, approximately 20% of individuals in each round of screening had positive results requiring some degree of follow-up, while approximately 1% had lung cancer. There was marked heterogeneity in this finding and in the frequency of follow-up investigations, biopsies, and percentage of surgical procedures performed in patients with benign lesions. Major complications in those with benign conditions were rare. Conclusion Low-dose computed tomography screening may benefit individuals at an increased risk for lung cancer, but uncertainty exists about the potential harms of screening and the generalizability of results.

Significant morbidity and expense result from frequent recurrences of nonmuscle-invasive bladder cancer (NMIBC) after standard treatment, and carcinoma in situ (Tis) is a poor prognostic factor. Predicated on observational and preclinical data strongly supporting cyclooxygenase-2 (COX-2) in the pathogenesis, and the activity of COX-2 inhibitors, in bladder cancer, we conducted a randomized, double-blind, placebo-controlled trial to determine whether celecoxib could reduce the time-to-recurrence (TTR) in NMIBC patients at high risk for recurrence. A total of 146 patients were randomized to celecoxib (200 mg) or placebo orally twice daily for at least 12 months. The average treatment duration was 1.25 years. Primary intent-to-treat analysis revealed celecoxib did not statistically significantly prolong TTR compared with placebo (P = 0.17, log rank) with a median follow-up of 2.49 years. The recurrence-free rate at 12 months with celecoxib was 88% (95% CI: 0.81–0.96) versus 78% (95% CI: 0.69–0.89) with placebo. After controlling for covariates with Cox regression analysis, recurrence rates did not differ between the two study arms (HR = 0.69; 95% CI: 0.37–1.29). However, celecoxib had a marginally significant effect on reducing metachronous recurrences (vs. placebo) with HR of 0.56 (95% CI: 0.3–1.06; P = 0.075). Celecoxib was well tolerated, with similar adverse events and quality-of-life in both arms. Our clinical trial results do not show a clinical benefit for celecoxib in preventing NMIBC recurrence but further investigation of COX-2 inhibitors in this setting is warranted. Cancer Prev Res; 4(10); 1580–9. ©2011 AACR.

In 2001, we reported that mortality may have been higher with isotretinoin (30 mg/d for 3 years) than with placebo in the subgroup of current smokers among the 1,166 patients with definitively resected early-stage non–small cell lung cancer who participated in the randomized, controlled Lung Intergroup Trial. We report the overall and cause (cancer, cardiovascular disease, or other)-specific mortality associated with long-term isotretinoin after an extended median follow-up of 6.2 years that included the capture of cause-of-death data from 428 deceased patients. Overall mortality was 36.7% in each of the two trial arms, about two thirds related to cancer and one third to other or unknown causes. Overall and cancer deaths increased in current smokers in the isotretinoin arm during the treatment and the extended follow-up period. No mortality end point increased among never smokers and former smokers taking isotretinoin, and cancer deaths decreased marginally in this combined subgroup. Isotretinoin also increased deaths from cardiovascular disease in current smokers. The present analysis supports the safety of protracted isotretinoin use in the combined group of never smokers and former smokers, which has important public health implications, for example, for treating acne in young people. The increased mortality in current smokers in this study is further evidence of the multifaceted danger of active smoking. The overall indications of this study have public health implications for treating acne in young people and other uses of retinoids in smokers. Cancer Prev Res; 3(6); 738–44. ©2010 AACR.

We previously showed that low-dose fenretinide (200 mg/d) had limited activity in retinoid-resistant oral leukoplakia (34% response rate) possibly because serum drug levels were insufficient to induce retinoid receptor–independent apoptosis. Therefore, we designed the single-arm phase II trial reported here to investigate whether higher-dose fenretinide would improve leukoplakia response over that of our previous study. Leukoplakia patients received fenretinide (900 mg/m2 twice daily) in four 3-week cycles (1 week on drug followed by 2 weeks off). At week 12, clinical responses were determined and blood samples were collected for serum drug level assessments. A planned interim futility analysis led to early trial closure after the initial 15 (of 25 planned) patients because only 3 (20%) had a partial response (stopping rule: ≤4 responses in first 16 patients). Fenretinide was well tolerated—only one grade 3 adverse event (diarrhea) occurred. Serum fenretinide levels changed from 0 (baseline) to 0.122 ± 0.093 μmol/L (week 12). In correlative in vitro studies, high-dose fenretinide inhibited the growth of head and neck cancer cells more and oral leukoplakia cells less than did lower doses of fenretinide. This result is consistent with our clinical finding that high-dose fenretinide did not improve on the historical response rate of lower-dose fenretinide in our previous oral leukoplakia trial.

Cross-contamination of cell lines is a serious but often unrecognized problem. We describe the authentication of a panel of transitional cell carcinoma cell lines using the short tandem repeat profiling technique to detect cross-contamination.Genomic DNA was isolated from UM-UC-1, UM-UC-2, UM-UC-3 (ATCC), UM-UC-6, UM-UC-9, UM-UC-10, UM-UC-11, UM-UC-13, UM-UC-14, UM-UC-16, T24 and KU7 cell lines. Short tandem repeat loci (D3S1358, D16S539, vWA, FGA, TH01, TPOX, CSF1PO, D5S818, D13S317 and D7S820) and a segment of the X-Y homologous gene amelogenin were co-amplified by polymerase chain reaction. Profiling was done using POP-4TM performance optimized polymer 4 (Applied Biosystems) with an ABI Prism 310 genetic analyzer. DNA sequencing of TP53 and immunohistochemistry for p53 were performed in UM-UC-3 and UM-UC-3-GFP.All cell lines had a unique short tandem repeat profile except UM-UC-2 and T24, which were virtually identical. T24 short tandem repeat profiles matched those of early passage number UM-UC-2. Stable transfection of the green fluorescence protein marker gene did not alter UM-UC-6, UM-UC-14 or KU7 profiles. However, the short tandem repeat profile for UM-UC-3-GFP was different from that of UM-UC-3. DNA sequencing showed a difference in TP53 between UM-UC-3 and UM-UC-3-GFP, confirming that UM-UC-3-GFP is not derived from UM-UC-3.Short tandem repeat profiling provides a unique genetic signature of human cell lines that does not significantly change with passage or green fluorescence protein transduction. Using short tandem repeat profiling we noted that the cell line UM-UC-2 is T24. DNA fingerprinting using short tandem repeat profiling is an easy and reliable tool that can be used to verify cell lines.

Dysregulation of Akt, PTEN, Drg-1, Cx-26, and L-plastin expression appear to be important in the progression of various cancers. Their expression in bladder cancer has not been well characterized. To assess the expression of these genes and their relationship to the outcome of bladder cancer, we used a bladder cancer tissue microarray (TMA) of 251 transitional cell carcinomas. We quantitated immunohistochemical staining of each protein using both automated and manual methods and correlated the expression levels with the clinicopathologic characteristics of the tumor and patient survival. Overall, the results from both automated and manual analyses were similar. We found a significant correlation between the expression of PTEN, Cx-26 and L-plastin with known clinically important pathologic features of bladder cancer (tumor grade, stage, and growth pattern). Aberrant localization patterns of Cx-26 and Drg-1 were observed in bladder tumors. There was also a significant correlation in expression among pAkt, PTEN, and L-plastin. Although the expression of these genes correlated with factors known to be associated with patient outcome, none of them was an independent predictor of progression-free or overall survival.

Objectives Cyclooxygenase 2 (COX-2) is aberrantly expressed in multiple tumor types including bladder cancer and is associated with enhanced growth, resistance to apoptosis, invasion, and angiogenesis. To evaluate the mechanisms through which COX-2 expression alters normal urothelium, we transfected the SV-40 immortalized human urothelial cell line SV-HUC with COX-2. Methods SV-HUC cells were stably transfected with a plasmid containing COX-2 under a CMV promoter. Following isolation of monoclonal transfectants, COX-2 expression was determined by Western and Northern analyses. Prostaglandin E2 (PGE2) in the culture supernatant was measured by ELISA. Cell growth was measured by crystal violet assay. Cellular invasion through Matrigel and anchorage-independent growth in 0.4% agarose were assessed. Tumorigenicity was evaluated by subcutaneous injection of cells in nude mice with and without Matrigel. Results Four of 12 clones stably overexpressing COX-2 at high levels relative to vector-transfected control cells were chosen for further study. Cell growth rates of these 4 clones were higher than vector control cells. PGE2 production was elevated in 3 of these 4 clones, and PGE2 levels correlated significantly with invasion through Matrigel. COX-2-transfected cells did not form colonies in soft agarose or tumors in nude mice. Conclusions Forced COX-2 expression in SV-HUC immortalized urothelial cells contributes to increased PGE2 production and increased invasion through Matrigel. However, it is insufficient to induce malignant transformation.

OBJECTIVES To develop a population-based clinical model of bladder cancer (BC) care costs and identify cost drivers. METHODS We retrospectively reviewed a cohort of 4863 patients with BC identified from the linked Surveillance, Epidemiology and End Results-Medicare database, aged at least 65 years and diagnosed between 1994 and 1996. We collected the records of Medicare reimbursements (a surrogate of costs) through 1998 and classified them into clinically relevant intervals and care types by disease invasiveness to derive the cumulative costs of care. We calculated the incremental resource use costs using sex and age-matched controls from a 5% general population sample and compared similarly matched patients with other cancer (OC). We inflated all costs to 2006 U.S. dollars. RESULTS The annual cost of care for all patients with muscle-invasive BC (MIBC) was $35.72M (95% confidence interval $35.69M to $35.75M), 70% more than the $21.03M (95% confidence interval $21.00M to $21.05M) for patients with non-MIBC. The major cost drivers, regardless of disease stage, were diagnostic/surveillance and complications, accounting for up to 43% and 37% of BC care costs, respectively. Comorbidity-adjusted incremental annual resource costs per patient with MIBC were more than four times greater than those for patients with non-MIBC, similar to those of OC controls (P = 0.490-0.913), except for inpatient (P = 0.002) and hospice (P

Purpose: Epidemiologic and clinical data suggest that selenium could prevent prostate cancer, but it has not been shown that supplemental selenium leads to an increased concentration of selenium in prostate tissue compared with adjacent tissue. Experimental Design: We conducted a randomized, controlled, short-term trial of l-selenomethionine (SeMet) versus observation in men with organ-confined prostate cancer. The primary endpoint was the measurement of selenium concentration in prostate tissue and seminal vesicle (SV). We assessed baseline selenium levels in serum and in toenail specimens (reflecting long-term intake) and post-intervention selenium levels in serum, and in prostate and SV tissues using hydride generation atomic fluorescence spectroscopy. Results: Sixty-six eligible patients were randomly assigned to the SeMet (n = 34) or observation (n = 32) arm; both arms had similar baseline patient characteristics. Baseline serum selenium was similar in the two groups (P = 0.64). Baseline toenail selenium levels were slightly higher in the SeMet group than in the control group (P = 0.07). After the intervention, the mean serum selenium level increased 15% in the SeMet arm and was higher than in the observation arm (P = 0.001). The selenium concentration in prostate tissue was 22% higher in the SeMet arm (n = 26) than in the observation arm (n = 25; 1.80 versus 1.47 ppm; P = 0.003, Wilcoxon rank sum test) and remained significantly higher after adjusting for chronic selenium intake (P = 0.021, ANCOVA). SV selenium concentration was similar in both groups (P = 0.384) and was lower than in prostate tissue. Conclusions: The present study is the first to show that selenium taken as oral supplementation accumulates preferentially in the human prostate gland as opposed to the SV. These findings support the hypothesis that oral selenium supplementation may contribute to the cancer preventive effects of selenium.

Selective COX-2 inhibitors such as celecoxib and NS-398 are being evaluated as chemopreventive and therapeutic agents for bladder and other cancers. We investigated the effects of these nonsteroidal anti-inflammatory agents on a panel of bladder cancer cell lines, and assessed their effects on anchorage-dependent and -independent growth, cell cycle, apoptosis and morphology. The human bladder cancer cell lines UM-UC-1, -3, and -6 were assayed for COX-2 expression by Western analysis using a monoclonal antibody to COX-2. UM-UC-1, -3, and -6 cells were grown in the presence of increasing concentrations of NS-398 and celecoxib, and cell growth was quantitated over 7 days by crystal violet elution. The cell lines were treated with NS-398 and celecoxib for 48 h and analyzed by flow cytometry with propidium iodide staining and Br-dUTP staining for apoptosis. Anchorage-independent growth was assessed using an agarose growth assay. Western analysis demonstrated that COX-2 expression in UM-UC-1, -6, and -3 was high, low, and undetectable, respectively. NS-398 and celecoxib produced dose-dependent growth inhibition of UM-UC-1 and -6. Both NS-398 and celecoxib also inhibited anchorage-dependent and -independent growth of UM-UC-3 in a dose-dependent fashion, despite the low basal expression of COX-2 in this cell line. Cell cycle analyses of UM-UC-1 and -6 revealed a 50% reduction in S-phase in the presence of 100 microM NS-398 whereas a smaller reduction in S-phase was noted in UM-UC-3 cells. Furthermore, treatment with 100 microM celecoxib resulted in significant apoptosis in all three cell lines, which was associated with downregulation of Bcl-2. COX-2 selective inhibitors NS-398 and celecoxib produced dose-dependent growth inhibition of bladder cancer cells associated with a significant reduction in S-phase. Induction of apoptosis in all three cell lines by celecoxib was associated with downregulation of Bcl-2. These changes occur independently of COX-2 expression levels suggesting the presence of a COX-2 independent pathway.

Objectives To estimate the lifetime cost of bladder cancer and the contribution of complications to the total costs. Methods We reviewed the medical records of a retrospective cohort of 208 patients with bladder cancer who registered at our comprehensive cancer center from 1991 to 1999. We multiplied the number of resources used during management of bladder cancer by their unit charges. We converted charges into costs using the Medicare cost-to-charge ratio and inflated these to 2005 U.S. dollars. We estimated future costs by creating two extreme hypothetical scenarios. In the best-case scenario, we assumed patients with superficial disease developed recurrences at the cohort’s mean rate and that patients with muscle-invasive disease were disease free after definitive therapy. Survival was based on the U.S. life expectancy in both cases. In the worst-case scenario, we assumed patients with superficial disease developed muscle-invasive disease and that all patients subsequently died of bladder cancer. Results The average cost of bladder cancer was $65,158 among the cohort patients. Sixty percent of this cost ($39,393) was associated with surveillance and treatment of recurrences, and 30% ($19,811) was attributable to complications. The lifetime cost of bladder cancer was lower for the worst-case scenario ($99,270) than for the best-case scenario ($120,684). However, a greater proportion of the costs were attributable to complications with the worst-case scenario (43%, $42,290) compared with the best (28%, $34,169). Conclusions The management of bladder cancer and its associated complications results in a major economic burden. More cost-effective surveillance strategies and approaches for preventing complications are crucial to minimizing the disease’s clinical and economic consequences.

15-Lipoxygenase-2 (15-LOX-2) synthesizes 15-S-hydroxyeicosatetraenoic acid (15-S-HETE), an endogenous ligand for the nuclear receptor, peroxisome proliferator-activated receptor-gamma (PPAR-gamma). Several studies have described an inverse relationship between 15-LOX-2 and PPAR-gamma expression in normal versus tumor samples. To systematically determine if this is a ubiquitous phenomenon, we used a variety of epithelial and nonepithelial cells and some tissues to further evaluate the extent of this inverse relationship. The levels of mRNA or protein were measured by reverse transcriptase polymerase chain reaction or Western gray level intensity, whereas distribution was determined by in situ hybridization or immunofluorescence. 15-S-HETE was measured by liquid chromatography/tandem mass spectrometry. Normal epithelial cells/samples generally expressed high levels of 15-LOX-2 along with the enzyme product 15-S-HETE, but both levels were reduced in cancer cells/samples. In contrast, most cancer cells expressed high levels of PPAR-gamma mRNA and protein, which were absent from normal epithelial cells. Overall, the inverse relationship between these two genes was primarily restricted to epithelial samples. Forced expression of PPAR-gamma reduced 15-LOX-2 protein levels in normal cells, whereas forced expression of 15-LOX-2 in tumor cells suppressed PPAR-gamma protein levels. These results suggest that feedback mechanisms may contribute to the loss of 15-LOX-2 pathway components, which coincide with an increase in PPAR-gamma in many epithelial cancers.

Cell lines have become an essential component for the investigation of cancer. We have developed a panel of cell lines derived from human urothelial cancers and we describe some of their important characteristics.Ten human urothelial cancer cell lines were characterized by their growth in athymic nude mice, CAR expression and their susceptibility to adenoviral mediated transfer of the green fluorescence protein gene. TP53 mutation status and immunochemical analysis of p53, pRB and p16 were also examined.Five cell lines rapidly produced tumors in athymic nude mice. Two cell lines produced tumors in 1 month, 1 produced them in 3 months and 2 were nontumorigenic. The cell lines varied in CAR expression and in their susceptibility to adenoviral mediated gene transduction. There was no direct correlation between CAR expression and susceptibility to adenoviral mediated gene transduction. Seven cell lines had TP53 mutations, of which 2 had large deletions and did not express p53 protein by immunostaining. All cell lines expressed abnormal pRB by immunochemical analysis (3 had no staining and 7 had homogenously strong staining) and 8 did not express p16 (7 showed homogeneously strong pRB staining).Our panel of 10 human urothelial cell lines differed in genetic alterations, growth in nude mice, susceptibility to adenoviral mediated gene transduction, and expression of p53, p16 and pRB. The availability of various urothelial cancer cell lines with differing genotypic and phenotypic features will facilitate further research into bladder cancer.

Recent studies have found that cyclooxygenase-2 (COX-2) protein expression was low and inducible with cytokines in prostate cancer cells (in the absence of serum) and that, in contrast, COX-2 expression was high in normal prostate epithelial cells (EC). Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) was expressed at high levels in the prostate cancer cell line PC-3 but not in ECs. In contrast to previous findings by others, PPAR-gamma ligands did not induce PPAR-gamma expression in EC or PC-3. The present study examined the relationship between PPAR-gamma and COX-2 expression patterns in EC and PC-3 in the presence and absence of serum and/or the PPAR-gamma agonist 15-deoxy-Delta12,14-prostaglandin J(2) (15d-PGJ(2)). We also evaluated the effects that the forced expression of PPAR-gamma1 and PPAR-gamma2 had on COX-2 in ECs. We found that expression of PPAR-gamma and COX-2 protein was inversely correlated in ECs and PC-3. Low COX-2 expression in PC-3 was up-regulated by serum, and 15d-PGJ(2) blocked serum-induced COX-2 expression and activity in a dose-dependent manner. 15d-PGJ(2) had no effect on COX-2 expression in ECs or PPAR-gamma expression in either cell type. However, forced expression of PPAR-gamma1 or PPAR-gamma2 in ECs suppressed the high level of endogenous COX-2. This effect was not isoform specific and was augmented by 15d-PGJ(2). The present study showed that PPAR-gamma activation can be an important regulator of COX-2 in prostate cells and may be an important target for prostate cancer chemoprevention.

Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) is being studied intensively for its role in carcinogenesis and in mediating the effects of prostate cancer treatment and prevention drugs. Prostate cancers express abundant and higher constitutive levels of PPAR-gamma than do normal prostate cells and are growth inhibited by ligand activation of PPAR-gamma. However, little is known about the role of PPARs in tumorigenesis or in normal prostate epithelial cells (EC). We examined the expression, phosphorylation patterns, and functions of the human PPAR (hPPAR)-gamma1 and hPPAR-gamma2 isoforms in normal prostate ECs to determine if activation of the receptor is sufficient for PPAR-gamma ligand activity in prostate cells. We found that ECs did not express either PPAR-gamma1 or PPAR-gamma2 protein and were not sensitive to growth inhibition by the PPAR-gamma ligand 15-deoxy-Delta12,14-prostaglandin J(2) (15d-PGJ(2)). In contrast, prostate cancer cells (PC-3), which express PPAR-gamma1 receptor isoform, are growth inhibited by PPAR-gamma ligand. Forced expression of hPPAR-gamma1 or hPPAR-gamma2 made ECs sensitive to 15d-PGJ(2) and led to reduced cellular viability. The direct repeat-1 promoter containing PPAR response elements was transactivated in ECs expressing exogenous PPAR-gamma1 or PPAR-gamma2, indicating that either isoform can be active in these cells. 15-Lipoxygenase-2, expressed at high levels in ECs, was down-regulated by transfecting PPAR-gamma expression construct (either gamma1 or gamma2 isoform) into ECs. Addition of PPAR-gamma ligand 15-hydroxyeicosatetraenoic acid in the presence of PPAR-gamma expression caused further down-regulation of 15-lipoxygenase-2. Our data illustrate that a PPAR-gamma ligand (15d-PGJ(2)) activates PPAR-gamma1 and selectively induces cell death in human prostate cancer cells but not in normal prostate ECs. These findings have important implications for the development of PPAR-gamma-targeting agents that prevent or treat prostate cancer and spare normal prostate cells.

Background The incidence of second primary lung cancers (SPLC) after resection of nonsmall cell lung cancer (NSCLC) is estimated to be 1% to 4% per patient year. The overall effect of SPLC on survival after resection of stage I NSCLC is unknown. Here we report the incidence, management, and outcome of SPLC in a large prospective cohort of patients who underwent careful follow-up. Methods National Cancer Institute Intergroup Trial NCI #I91-0001 examined the effectiveness of isotretinoin A for chemoprevention of second primary tumors, the primary endpoint in that trial. Prospective data from patients randomly assigned to the placebo arm were analyzed. Results Five hundred sixty-nine patients underwent complete resection of pathologic stage I NSCLC. The median follow-up was 5.9 years. Second primary tumors developed in 88 (15%) patients. Of these, 49 (56%) were SPLC (incidence = 1.99/100 patient-years), with a median interval from initial surgery of 4.2 years. Second primary lung cancer never developed in patients who had never smoked (n = 44, p = 0.046; never versus ever smokers). Current smokers had a higher incidence of SPLC than former smokers (hazard ratio=1.91, p = 0.03). Age, sex, stage, histology, tumor location and initial surgery had no effect on SPLC development. Despite semiannual follow-up with chest radiographs, 12 (24%) patients had metastatic disease at the time of diagnosis of SPLC. Surgical resection was performed in 31 (63%) SPLC patients. Median survival was 4.1 years in those who underwent surgery and 1.4 years in those who did not ( p = 0.003). Overall SPLC-related mortality in the original cohort was 3.7%. Conclusions Patients who undergo surgery for SPLC can achieve prolonged survival. Despite close follow-up however many patients with SPLC present with advanced disease. That indicates a need for continued lifelong postoperative surveillance.

Bladder tumors from Egyptian patients with a high prevalence of bilharziasis were immunohistochemically analyzed for the expression of cytokeratin 20 (CK20), AN43, prostaglandin dehydrogenase (PGDH), and cyclooxygenase-2 (COX-2). The tumors included 26 transitional cell carcinomas (TCC), 10 squamous cell carcinomas (SCC) and 2 tumors of mixed TCC/SCC histology. Of the 28 TCC tumors, 21 (75%) expressed CK20 and 25 (89%) expressed AN43. CK20 was not expressed in any of the 10 SCC tumors and AN43 was expressed in 2 of them. PGDH was expressed in 18 (64%) of the 28 tumors with TCC histology and 1 of the 10 SCC. A subset of 21 tumors (16 TCC and 5 SCC) was tested for COX-2 expression. COX-2 was detected in 69% of the 16 TCC tumors examined but was not seen in the SCC tumors. As tumors increased in stage, the expression of these proteins changed. CK20, AN43 and PGDH decreased but COX-2 expression increased in higher stage tumors. The histologic phenotype of these cancers is reflected in their expression of these proteins and is modified further as tumors progress in stage.

Centrosomal abnormalities have been implicated in chromosomal segregation aberrations that result from the formation of multipolar mitotic spindles and lead to aneuploidy. Aneuploidy is a characteristic of neoplasia and underlies the development and progression of bladder cancer. Therefore, centrosomal abnormality may play a key role in urothelial tumor transformation. The purpose of our investigation was to determine whether centrosomal abnormalities are present in malignant urothelial cells, define the relationship between centrosomal abnormalities and aneuploidy and determine whether the presence of centrosomal abnormalities might be a potential diagnostic marker for bladder cancer. Bladder wash specimens obtained from patients with and without a history of urothelial carcinoma were analyzed for centrosomal abnormalities using an immunoassay with a γ-tubulin antibody. FISH with centromeric probes for chromosomes 4 and 9 and DNA ploidy image analysis were performed to detect aneuploidy. Defective centrosomes were found in 40 of 45 bladder wash specimens from patients with bladder cancer but in none of the 10 samples from patients without it. A large percentage (69%) of grade 1 tumors were positive for centrosomal abnormalities, and these abnormalities were increasing in numbers and size in grade 2 (93%) and grade 3 (100%) specimens. Centrosomal abnormalities and numerical chromosomal aberrations frequently appeared concomitantly in the same malignant cells. All of the specimens showing aneuploidy also exhibited centrosomal abnormalities: centrosomal defects and aneuploidy occurred together in 80% of malignant bladder tumors, with an especially high percentage in higher-grade tumors. The overall positivity of centrosomal abnormalities was higher than that of aneuploidy (88% vs. 80%), especially in grade 1 tumors (69% vs. 46%), whereas aneuploidy was strongly associated with grade 2 and grade 3 tumors. Centrosomal abnormalities are common in bladder cancer, even in low-grade tumors, and strongly associated with cancer grade and aneuploidy, especially in high-grade neoplasms. Centrosomal abnormalities appear to be intrinsic to aneuploidy and tumorigenesis and may be potential markers for early detection of bladder cancer. © 2003 Wiley-Liss, Inc.

The synthetic retinoid N-(4-hydroxyphenyl)retinamide [4-HPR (or fenretinide)] has preclinical and clinical preventive activity in breast carcinogenesis. 4-HPR and its metabolites have been shown to accumulate in the mammary tissue of rodents. We assessed levels of 4-HPR and its major metabolite, N-(4-methoxyphenyl)retinamide (4-MPR), in plasma and in normal and neoplastic breast tissue obtained from women treated with 4-HPR.We randomly assigned 14 women with suspected or very recently diagnosed breast cancer to receive 100, 200, or 300 mg of 4-HPR daily for 3-12 days before scheduled biopsy, lumpectomy, or mastectomy. Using high-performance liquid chromatography, we measured post-4-HPR-treatment concentrations of 4-HPR and 4-MPR in plasma and breast tissue obtained during surgery.Breast tissue and plasma retinamide (4-HPR plus 4-MPR) concentrations increased significantly with short-term oral administration of 4-HPR. Retinamide levels increased in a linear and dose-related fashion in plasma, whereas they peaked and plateaued at 200 mg/day in breast tissue. The total retinamide concentration in breast tissue exceeded that in plasma at each 4-HPR dose. The highest mean tissue:plasma retinamide ratios were achieved at 200 mg/day: 639.5 +/- 253.8 to 190.6 +/- 91.9 ng/ml (4.8:1) for 4-HPR and 594.4 +/- 201.9 to 130.5 +/- 37.8 ng/ml (6.6:1) for 4-MPR. Plasma retinol levels decreased in association with increasing 4-HPR doses. Two patients experienced grade 1 toxicity at the 300 mg/day dose.These findings indicate that retinamides preferentially accumulate in human breast tissue (versus plasma). 4-HPR tissue concentrations at 200 mg/d were equivalent to those that inhibit growth and induce apoptosis of breast cancer cells in vitro. Previous clinical and correlative laboratory results suggest that 4-HPR may reduce risk in premenopausal women, who are more prone (than are postmenopausal women) to estrogen receptor (ER)-negative breast cancer development. The present results and previous data (including in vitro 4-HPR activity against ER-negative breast cancer) support further study of 4-HPR in the setting of premenopausal/ER-negative breast cancer prevention.

We have previously demonstrated decreased Jun/AP-1 activity in the breast cancer cell line MCF-7 when compared to normal or immortalized mammary epithelial cells. In this paper, we overexpress Jun in MCF-7 cells (MCF7Jun) and demonstrate that it results in diverse biologic and biochemical changes, which mimic those seen clinically in breast cancer. Overexpression of Jun causes significant alterations in the composition of AP-1, decreased junB and increased fra-1 expression and results in an increased biologic aggressiveness. MCF7Jun cells exhibit increased cellular motility, increased expression of a matrix degrading enzyme MMP-9, increased in vitro chemoinvasion and tumor formation in nude mice in the absence of exogenous estrogens. Furthermore, MCF7Jun cells are unresponsive to the growth stimulating effects of estrogen and growth inhibitory effects of tamoxifen. Analysis of the estrogen receptor (ER) expression and activity showed that the MCF7Jun cells have no detectable ER. MCF-7 cells overexpressing mutant forms of cJun were responsive to the growth stimulatory effects of estrogen indicating that full-length cJun is required to acquire the estrogen-independent phenotype in breast cancer cells.

Background: Isolation of the first neoplastic acquired immunodeficiency syndrome-related Kaposi's sarcoma (KS) cell line (KS Y-1) has furthered understanding of the pathogenesis of KS. Studies with KS Y-1 cells have indicated that inhibition of KS cell proliferation occurs in early pregnancy in mice and after treatment with certain commercial preparations of human chorionic gonadotropin (hCG, a pregnancy hormone purified from urine). The activity of the commercial preparations has been attributed to an hCG-associated factor(s) (HAF). While several clinical benefits of HAF are clearly evident, the basis for its anti-KS properties remains unknown. We investigated the apoptosis-inducing effects of HAF and the expression of apoptosis-related proteins in KS cells. Methods: KS Y-1 and KS SLK cells were treated with clinical-grade crude preparations of hCG, recombinant hCG, or urine fractions exhibiting anti-KS activity and then examined for features of apoptosis. Levels of proteins associated with apoptosis were monitored by western blot analysis, and cell DNA content was assessed by flow cytometry. Tumors induced in mice by inoculation of KS Y-1 cells were treated with preparations of hCG, and the tumors were examined for cell morphology and also for DNA fragmentation by use of the terminal deoxynucleotidyl transferase-mediated digoxigenin-deoxyuridine triphosphate nick-end-labeling (TUNEL) assay. Results: The HAF present in some preparations of hCG and in urine fractions has the ability to induce apoptosis in KS cells in vitro and in vivo. HAF-triggered apoptosis was preceded by increased levels of the apoptosis-related proteins c-Myc and c-Rel and cell accumulation in G 0 /G 1 phase of the cell cycle. KS Y-1 cells transfected with a c-Myc complementary DNA showed elevated rates of apoptosis. Conclusion: The anti-KS activity of HAF appears to induce apoptosis. Such activity suggests a role for HAF in pregnancy-related regulation of cell death.

In an attempt to characterize the molecular alterations of cervical adenocarcinoma, we analyzed 32 paraffin-embedded specimens for the presence of K-ras mutations, p53 overexpression, p16 and Rb protein expression, and the presence of HPV 16 and 18 DNA. Overall 25/32 (78%) of the tumors displayed an abnormality in at least one of these analyses. K-ras mutations were detected by PCR amplification and RFLP analysis in 3 tumors, including 2 at codon 12 and 1 at codon 61. p53 overexpression determined by immunohistochemistry was demonstrated with80% of tumor nuclei staining in 4 cases, 10-15% of nuclei staining in 3 cases, and1% of nuclei staining in 5 cases. The pattern of staining was diffuse in 6 cases, focal in 1 case, and scattered in 5 cases. Analysis of p16 protein expression in 23 specimens revealed 1 tumor with abnormal staining, while Rb protein expression was determined to be normal in all 25 tumors tested. HPV DNA, detected by PCR with type-specific primers, was found in 16 tumors (50%), including 7 (22%) with HPV 16 and 9 (28%) with HPV 18. There was no correlation among these abnormalities except that the presence of HPV and strong p53 overexpression (80% tumor nuclei staining) were mutually exclusive events. Clinical correlation demonstrated that p53 overexpression involving the majority of tumor cell nuclei is characteristic of advanced stage disease, while HPV positivity and activated ras genes are associated with early stage disease.