Copland, John A.

Abstract Bladder cancer is the 5th most commonly diagnosed cancer in the United States. Because superficial bladder cancer (SBC) recurs in about 60% of cases, standard treatment involves tumor resection followed by intravesical therapies to prevent recurrence, as well as lifelong periodic cystoscopy. There is an urgent need for better intravesical therapies to prevent both the recurrence and progression of SBC. Recent evidence has demonstrated that glycogen synthase kinase – 3 beta (GSK3β) plays an important role in pancreatic, ovarian and colon cancers; however, to date the possible role of this important kinase has not been examined in bladder cancer. We propose that GSK3β plays a cancer promoting role in bladder cancer and that inhibition of GSK3β will reduce bladder cancer growth. The GSK3β inhibitor, AR-A014418, has been demonstrated to reduce tumor growth in the previously cited cancers. AR-A014418 reduced cell proliferation of three bladder cancer cell lines (HT1376, T24 and UM-UC-3) with EC50 values of 17.5μM for HT1376, 8μM for T24, and 10μM for UM-UC-3 cells. The effect of AR-A014418 on downstream targets in these cell lines was evaluated by Western blots which indicated an increase in apoptosis in these cells in response to drug treatment as demonstrated by the presence by both PARP and caspase 3 cleavage. The NFkB downstream target XIAP also exhibited reduced protein levels after drug treatment. The HT1376 cell line was transduced with luciferase containing lentivirus particles to generate a luciferase expressing cell line, HT1376-LUC to use for both ectopic and orthotopic models of bladder cancer growth. The efficacy of AR-A014418 in reducing bladder cancer tumor growth was tested in an ectopic mouse model. HT1376-LUC cells mixed with matrigel were transplanted into the right flank of 50 female nude mice. After one week, mice containing tumors with an average volume of 52 mm3 were randomized into four treatment groups of 10 mice each. Mice were treated twice weekly via IP injection with placebo, 1, 5 or 7mg/kg AR-A014418. Tumor sizes were measured and imaged for luciferase activity once weekly. Tumor growth was inhibited by both 5mg/kg and 7mg/kg doses by 19 days of treatment (p < 0.05, Student's T test). Together, the in vitro and in vivo data suggest that GSK3β is a novel target for bladder cancer treatment. Future studies using an orthotopic model of bladder cancer are planned. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1671. doi:10.1158/1538-7445.AM2011-1671

Abstract Background: Stage IVC (metastatic) anaplastic thyroid cancer (ATC) has a median overall survival (OS) of only 1.9 months. Single agent paclitaxel has modest activity in ATC. In pre-clinical studies, CS-7017 inhibited ATC proliferation through a novel mechanism–activation of PPAR-γ, followed by sequential upregulation of RhoB, then p21. Moreover, CS-7017 augmented the proapoptotic effects of paclitaxel. Based on these results, a multicenter study was conducted to determine if CS-7017+paclitaxel might prove safe and effective in advanced ATC. Design: A Phase 1 study was conducted to determine the Phase 2 dose. In Phase 1, patients received CS-7017 orally BID for one week (run-in phase), followed by a 3 h iv infusion of paclitaxel (175 mg/m2) every 3 weeks in combination with BID CS-7017. CS-7017 doses were escalated (0.15, 0.3, or 0.5 mg). Tissue biopsies were obtained at baseline and at one and three weeks for immunohistochemistry of PPAR-γ responsive proteins; serum CS-7017 PK studies were also performed. Results: Nineteen patients were enrolled, but 4 were not dosed (3 due to progression, 1 ineligible). Seven participated in CS-7017 dose levels 1a/1b (0.15 mg BID), six in dose level 2 (0.3 mg BID), and two in dose level 3 (run-in phase only–0.5 mg BID). Demographics: Of the 15 treated patients, 10 (67%) were women. Median age was 59 years (range: 43-82) Efficacy: Of 15 patients receiving drug, one had a confirmed partial response (PR) lasting from Day 69 to Day 175, and eight had stable disease (SD) as their best response. Median Time to Progression in 7 patients at Dose Level 1 was 49 days, but 70 days in Dose Level 2 (43% prolongation); corresponding median survival was 99 (0.15 mg BID) vs. 140 days (0.3 mg BID, 42% increase). Median peak CS-7017 blood level was 8.6 ng/mL (range: 5.1 to 13.7) for Level 1 and 22.0 ng/mL (17.0 to 31.5) for Dose Level 2. Adverse Events: Ten patients had AEs ≥ Grade 3, with two (anemia and localized edema) related to CS-7017. Thirteen events of fluid retention/edema were reported in 8 patients, with only 2 events of CTCAE grade ≥ 3. Eight patients had ≥ one SAE, with one (anemia) due to CS-7017. One SAE (anaphylactic reaction) was related to paclitaxel. No dose limiting toxicity was observed. Immunohistochemistry: Biopsies were available on 7 patients. PPAR-γ, RXR-α, RhoB were present in all; Angiopoietin-like 4 was induced by CS-7017. Conclusions: Combination therapy with CS-7017 and paclitaxel was tolerated and has biologic activity in patients with metastatic anaplastic thyroid carcinoma, with initial exploratory data suggesting a dose-dependent improvement in time to progression and survival in response to escalating CS-7017 dosage. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1278. doi:10.1158/1538-7445.AM2011-1278

The development of subspecialty tumor groups for uncommon malignancies represents an effective approach to building experience, increasing patient volumes and referrals, and fostering development of increased therapeutic options and clinical trials for patients afflicted with otherwise historically neglected cancers.

Abstract Background: Treatment of aromatase refractory metastatic breast cancer (MBC) is difficult and challenging. Estrogen receptor (ER) resistance causes enhanced expression of the vascular endothelial growth factor (VEGF).Several studies have shown that the ER interacts with the VEGF pathway and is an important mechanism of resistance. Therefore we embarked on a phase II study of fulvestrant, a complete ER suppressor and bevacizumab, a well studied VEGF monoclonal antibody in aromatase refractory MBC patients. Methods: A single stage phase II study with an interim analysis of fulvestrant and bevacizumab was conducted with these objectives: 6 month progression-free survival rate (PFS), tumor response, toxicity, and overall survival. Regimen: fulvestrant 250 mg day1 and 15 (cycle 1) then day 1 (cycle 2 and beyond) and bevacizumab 10mg/kg days 1 and 15 of each 4 weeks is a cycle. Results: At the time of interim analysis, 11/20 evaluable patients achieved 3-month progression-free survival status while remaining on treatment for at least 3 months, not meeting the protocol specified efficacy requirements and thus halting accrual. 36 patients were enrolled from September 2007-December 2008; 33 patients were evaluable. Number of prior metastatic chemotherapy regimens: 0 in 26 patients and 1 in 7 patients. 22 (67%) patients received prior hormonal therapy in the metastatic setting. 18 (55%) had measurable disease. A median of 6 cycles (range 1-19) were administered. 12/33 evaluable patients (95% CI:20-55%) achieved 6-month progression-free survival status while remaining on treatment for a least 6 months. Among 18 patients with measurable disease, 2 (11%) patients CI:1.4-35%) had a confirmed tumor reponse (both PR). Additionally, 2 patients had stable disease for greater than 6 months, for a clinical benefit rate of 22%. Median follow up was 8.5 months (range 1.7-17.5 months). Median progression-free survival was 6.2 months (95% CI:5.4-10.1 months). The 6 -month overall survival rate was 84.8% (95% CI 73.5-98%). The median dose level administered was 250 mg for fulvestrant and 10 mg/kg for bevacizumab for cycles 1-19. The most common grade 3/4 adverse events (AEs) were hypertension 2 (6%), headache pain 2 (6%), and confusion 2 (6%). There was 1 grade 5 central nervous system hemorrhage. 13 (39%) patients experienced a grade 3 non-hematologic AE and 4 (12%) experience a grade 4+ non-hematologic AE. Conclusion: Fulvestrant/bevacizumab is safe and tolerable. Although this regimen did not meet its statistical endpoint, 22% of evaluable patients with aromatase refractory diesease achieved clinical benefit with minimal toxicity. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4096.

Abstract Disparities in clinical outcomes of breast cancer have been described among different racial and ethnic groups in the U.S. Convincing data exist showing that Latina women have a lower incidence of breast cancer but a higher breast cancer–related mortality rate compared with white women. Noticeable differences in breast cancer incidence are present even within different Latina subsets with a higher incidence in second- and third-generation women compared with foreign born. An increasing amount of data exists pointing to significant differences in the genetics and biology of breast cancer in Latinas as a significant contributor to the higher mortality, including a higher incidence of triple-negative breast cancers (which do not overexpress HER-2 protein and are negative for estrogen receptors and progesterone receptors). Other social and environmental factors are likely to play a significant role as well, including a lower rate of screening mammography, variable access to medical care, among others. Recent data are inconclusive regarding differences among racial/ethnic groups in the response to chemotherapy. Data on racial/ethnic variations in the pharmacogenomics of chemotherapy, endocrine treatments, and toxicity are more limited, with some data suggesting differences in frequencies of polymorphisms of genes involved in the metabolism of some of these agents. Further studies are needed on this subject.

Abstract Anaplastic thyroid carcinoma (ATC) is a highly aggressive form of the disease for which new therapeutic options are desperately needed. Previously, we showed that the high-affinity peroxisome proliferator–activated receptor γ (PPARγ) agonist, RS5444, inhibits cell proliferation of ATC cells via induction of the cyclin-dependent kinase inhibitor p21WAF1/CIP1 (p21). We show here that up-regulation of RhoB is a critical step in PPARγ-mediated activation of p21-induced cell stasis. Using multiple independently derived ATC cell lines, we found that treatment with RS5444 leads to the up-regulation of RhoB and subsequent activation of p21, and that silencing of RhoB by RNAi blocks the ability of RS5444 to induce p21 and to inhibit cell proliferation. Our results show that transcriptional regulation of RhoB by the nuclear transcription factor PPARγ is responsible for the induction of p21 mRNA and protein. We further implicate RhoB as a key signaling effector for the growth inhibition of ATC, as treatment with a histone deacetylase inhibitor shown to increase RhoB expression in lung cancer cells caused the up-regulation of RhoB in ATC cells accompanied by increased expression of p21 and inhibition of cell proliferation; this effect occurred even in ATC cells that were unresponsive to RS5444 due to a lack of expression of PPARγ. Our results implicate RhoB as a novel intermediate in critical signaling pathways and as an additional target for therapeutic intervention in ATC. [Cancer Res 2009;69(4):1536–44]

Abstract Purpose: p27 localization and expression has prognostic and predictive value in cancer. Little is known regarding expression patterns of p27 in renal cell carcinoma (RCC) or how p27 participates in disease progression or response to therapy. Experimental Design: RCC-derived cell lines, primary tumors, and normal renal epithelial cells were analyzed for p27 expression, phosphorylation (T157 of the NLS), and subcellular localization. RCC-derived cell lines were treated with phosphatidylinositol 3-kinase (PI3K) and mammalian target of rapamycin (mTOR) inhibitors and effects on p27 localization were assessed. The potential contribution of cytoplasmic p27 to resistance to apoptosis was also evaluated. Results: p27 was elevated in tumors compared with matched controls, and cytoplasmic mislocalization of p27 was associated with increasing tumor grade. Cytoplasmic localization of p27 correlated with phosphorylation at T157, an AKT phosphorylation site in the p27 NLS. In RCC cell lines, activated PI3K/AKT signaling was accompanied by mislocalization of p27. AKT activation and phosphorylation of p27 was associated with resistance to apoptosis, and small interfering RNA knockdown of p27 or relocalization to the nucleus increased apoptosis in RCC cells. Treatment with the PI3K inhibitors LY294002 or wortmannin resulted in nuclear relocalization of p27, whereas mTOR inhibition by rapamycin did not. Conclusions: In RCC, p27 is phosphorylated at T157 of the NLS, with increasing tumor grade associated with cytoplasmic p27. PI3K inhibition (which reduces AKT activity) reduces T157 phosphorylation and induces nuclear relocalization of p27, whereas mTOR inhibition does not. Clinical testing of these findings may provide a rational approach for use of mTOR and PI3K/AKT pathway inhibitors in patients with RCC.

p120 catenin regulates the activity of the Rho family guanosine triphosphatases (including RhoA and Rac1) in an adhesion-dependent manner. Through this action, p120 promotes a sessile cellular phenotype when associated with epithelial cadherin (E-cadherin) or a motile phenotype when associated with mesenchymal cadherins. In this study, we show that p120 also exerts significant and diametrically opposing effects on tumor cell growth depending on E-cadherin expression. Endogenous p120 acts to stabilize E-cadherin complexes and to actively promote the tumor-suppressive function of E-cadherin, potently inhibiting Ras activation. Upon E-cadherin loss during tumor progression, the negative regulation of Ras is relieved; under these conditions, endogenous p120 promotes transformed cell growth both in vitro and in vivo by activating a Rac1–mitogen-activated protein kinase signaling pathway normally activated by the adhesion of cells to the extracellular matrix. These data indicate that both E-cadherin and p120 are important regulators of tumor cell growth and imply roles for both proteins in chemoresistance and targeted therapeutics.