Dutasteride monotherapy in men with serologic relapse following radical therapy for adenocarcinoma of the prostate: a pilot study.

OBJECTIVE: The objective of this study was to assess the effect of dutasteride on serum prostate specific antigen (PSA) levels in men with serologic relapse following radical prostatectomy and/or radiation therapy for clinically localized adenocarcinoma of the prostate. METHODS: A prospective, single institution, IRB approved trial was conducted. Entry criteria required that all participants have serologic disease relapse only with serum PSA levels between 0.4 and 10.0 ng/ml. Enrolled participants were treated with 0.5 mg dutasteride daily. The primary endpoints were serum PSA level and clinical recurrence. The rate of durable decline in PSA was assessed according to the recommendations of the Prostate-Specific Antigen Working Group. RESULTS: Thirty-five patients provided informed consent and participated in the present study. At a median follow-up duration of 27 months (range, 4-42 months), 46% of enrolled men had a serum PSA decrease of greater than 10%, and 25% had a serum PSA decrease of greater than 50% (P < 0.001). Pre-study PSA doubling time (PSADT) (≥12 months vs. <12 months), and Gleason score (≤6 vs. ≥7) were associated with a better response to dutasteride, but only PSADT was statistically significant (P < 0.001). Thirty percent of patients experienced PSA progression (increase in serum PSA of greater than 50%). Two (6%) patients developed bone metastasis. No patient was removed from the study for drug-related toxicity. CONCLUSIONS: In the present pilot study, treatment with dutasteride resulted in a significant decrease in serum PSA in men with serologic relapse following radical treatment for adenocarcinoma of the prostate. These data appear to suggest that dutasteride may delay or prevent progression of prostate cancer in some men with biochemical relapse after radical therapy. These findings require confirmation in the setting of a larger, longer trial.

Dissection of platelet and myeloid cell defects by conditional targeting of the beta3-integrin subunit.

The purpose of this work was to determine platelet and myeloid cell-specific requirements for beta3-containing integrins in hemostasis, bone resorption, and tumor growth. LoxP-flanked mice were generated to study the conditional deletion of beta3-integrin in platelets [knockout in platelets (KOP)] and myeloid cells [knockout in myeloid (KOM)]. Using the beta3KOP and beta3KOM strains of mice, we studied the role of beta3-integrin in hemostasis, bone resorption, and subcutaneous tumor growth. Tissue-specific deletion of platelet beta3-integrins in beta3KOP mice did not affect bone mass but resulted in a severe bleeding phenotype. No growth difference of tumor xenografts or in neoangiogenesis were found in beta3KOP mice, in contrast to the defects observed in germline beta3(-/-) mice. Conditional deletion of myeloid beta3-integrins in beta3KOM mice resulted in osteopetrosis but had no effect on hemostasis or mortality. Tumor growth in beta3KOM mice was increased and accompanied by decreased macrophage infiltration, without increase in blood vessel number. Platelet beta3-integrin deficiency was sufficient to disrupt hemostasis but had no effect on bone mass or tumor growth. Myeloid-specific beta3-integrin deletion was sufficient to perturb bone mass and enhance tumor growth due to reduced macrophage infiltration in the tumors. These results suggest that beta3-integrins have cell-specific roles in complex biological processes.-Morgan, E. A., Schneider, J. G., Baroni, T. E., Uluçkan, O., Heller, E., Hurchla, M. A., Deng, H., Floyd, D., Berdy, A., Prior, J. L., Piwnica-Worms, D., Teitelbaum, S. L., Ross, F. P., Weilbaecher, K. N. Dissection of platelet and myeloid cell defects by conditional targeting of the beta3-integrin subunit.

Genome-wide analysis identifies interleukin-10 mRNA as target of tristetraprolin.

Tristetraprolin (TTP) is an RNA-binding protein required for the rapid degradation of mRNAs containing AU-rich elements. Targets regulated by TTP include the mRNAs encoding tumor necrosis factor-alpha, granulocyte-macrophage colony-stimulating factor, interleukin-2 (IL-2), and immediate early response 3. To identify novel target mRNAs of TTP in macrophages, we used a genome-wide approach that combines RNA immunoprecipitation and microarray analysis. A list was compiled of 137 mRNAs that are associated with TTP with an estimated accuracy on the order of 90%. Sequence analysis revealed a highly significant enrichment of AU-rich element motifs, with AUUUA pentamers present in 96% and UUAUUUAUU nonamers present in 44% of TTP-associated mRNAs. We further show that IL-10 is a novel target regulated by TTP. IL-10 mRNA levels were found to be elevated because of a reduced decay rate in primary macrophages from TTP(-/-) mice. Our study demonstrates the importance of experimental approaches for identifying targets of RNA-binding proteins.

Inactive full-length p53 mutants lacking dominant wild-type p53 inhibition highlight loss of heterozygosity as an important aspect of p53 status in human cancers.

Over 1000 different mutants of the tumor suppressor protein p53 with one amino acid change in the core domain have been reported in human cancers. In mouse knock-in models, two frequent mutants displayed loss of wild-type (wt) p53 function, inhibition of wt p53 and wt p53-independent gain of function. The remaining mutants have been systematically characterized for loss of wt p53 function, but not other phenotypes. We report the concomitant assessment of loss of function and interference with wt p53 using URA3-based p53 yeast and confirmatory mammalian assays. We studied 76 mutants representing 54% of over 15 000 reported missense core domain mutations. The majority showed the expected complete loss of wt p53 function and dominant p53 inhibition. A few infrequent p53 mutants had wt p53-like activity. Remarkably, one-third showed no interference with wt p53 despite loss of wt p53 function at 37 degrees C. Half of this group consisted of temperature-sensitive p53 mutants, but the other half was surprisingly made up of mutants with complete loss of wt p53 function. Our findings illustrate the diverse behavior of p53 mutants and mechanisms of malignant transformation by p53 mutants. The identification of full-length p53 mutants without dominant inhibition of wt p53 highlights the importance of determining the status of the wt p53 allele in human cancers, in particular in the context of clinical studies. In the case of p53 mutants with no or weak dominant p53 inhibition, presence of the wt allele may indicate a good prognosis cancer, whereas loss of heterozygosity may spell an aggressive, therapy-resistant cancer.

Ribonomic and short hairpin RNA gene silencing methods to explore functional gene programs associated with tumor growth arrest.

In this chapter, we present an approach using genomic and ribonomic profiling to investigate functional gene programs in a tumor growth model. To reach this goal, ribonomic profiling was combined with RNA interference in a tumor dormancy model. Strategies merging functional genomic technologies are outlined for the identification of novel posttranscriptionally regulated targets of p38 to show that they are functionally linked to the induction or interruption of cellular growth in cancer. In the first section of this chapter, we describe a method for the detection of mRNA subsets associated with RNA-binding proteins such as hnRNP A1 using (1) immunopurification of mRNA-protein complexes, from either whole cell lysates or subcellular fractions and (2) gene expression arrays to find those mRNAs bound to hnRNP A1. In the second section, short hairpin RNA technology was used to create a library of shRNAs that target p38 induced mRNAs expression libraries are utilized to "knockdown" the genes identified in the first section. Finally, this library of gene candidates is evaluated in vivo to address their functional role in the induction or maintenance of dormancy.

A global suppressor motif for p53 cancer mutants.

The transcription factor and tumor suppressor protein p53 is frequently inactivated in human cancers. In many cases, p53 gene mutations result in high levels of inactive, full-length p53 protein with one amino acid change in the core domain that recognizes p53 DNA-binding sites. The ability to endow function to mutated p53 proteins would dramatically improve cancer therapy, because it would reactivate a central apoptotic pathway. By using genetic strategies and p53 assays in yeast and mammalian cells, we identified a global suppressor motif involving codons 235, 239, and 240. These intragenic suppressor mutations, either alone or in combination, restored function to 16 of 30 of the most common p53 cancer mutants tested. The 235-239-240 suppressor motif establishes that manipulation of a small region of the core domain is sufficient to activate a large number of p53 cancer mutants. Understanding the structural basis of the rescue mechanism will allow the pursuit of small compounds able to achieve a similar stabilization of p53 cancer mutants.