Poly(ADP-ribose) polymerases (PARPs) are a family of enzymes that use NAD+ as a substrate to synthesize polymers of ADP-ribose (PAR) as post-translational modifications of proteins. PARPs have important cellular roles that include preserving genomic integrity, telomere maintenance, transcriptional regulation, and cell fate determination. The diverse biological roles of PARPs have made them attractive therapeutic targets, which have fueled the pursuit of small molecule PARP inhibitors. The design of PARP inhibitors has matured over the past several years resulting in several lead candidates in clinical trials. PARP inhibitors are mainly used in clinical trials to treat cancer, particularly as sensitizing agents in combination with traditional chemotherapy to reduce side effects. An exciting aspect of PARP inhibitors is that they are also used to selectivity kill tumors with deficiencies in DNA repair proteins (e.g. BRCA1/2) through an approach termed “synthetic lethality.” In the midst of the tremendous efforts that have brought PARP inhibitors to the forefront of modern chemotherapy, most clinically used PARP inhibitors bind to conserved regions that permits cross-selectivity with other PARPs containing homologous catalytic domains. Thus, the differences between therapeutic effects and adverse effects stemming from pan-PARP inhibition compared to selective inhibition are not well understood. In this review, we discuss current literature that has found ways to gain selectivity for one PARP over another. We furthermore provide insights into targeting other domains that make up PARPs, and how new classes of drugs that target these domains could provide a high degree of selectivity by affecting specific cellular functions. A clear understanding of the inhibition profiles of PARP inhibitors will not only enhance our understanding of the biology of individual PARPs, but may provide improved therapeutic options for patients.

RNA-binding protein HuR binds U- or AU-rich sequences in the 3′-untranslated regions of target mRNAs, stabilizing them and/or modulating their translation. Given the links of HuR with cancer, we studied the consequences of modulating HuR levels in pancreatic cancer cells. HuR-overexpressing cancer cells, in some instances, are roughly up to 30-fold more sensitive to treatment with gemcitabine, the main chemotherapeutic component of treatment regimens for pancreatic ductal adenocarcinoma (PDA), compared with control cells. In pancreatic cancer cells, HuR associates with deoxycytidine kinase (dCK) mRNA, which encodes the enzyme that metabolizes and thereby activates gemcitabine. Gemcitabine exposure to pancreatic cancer cells enriches the association between HuR and dCK mRNA and increases cytoplasmic HuR levels. Accordingly, HuR overexpression elevates, whereas HuR silencing reduces, dCK protein expression in pancreatic cancer cells. In a clinical correlate study of gemcitabine treatment, we found a 7-fold increase in risk of mortality in PDA patients with low cytoplasmic HuR levels compared with patients with high HuR levels, after adjusting for other treatments and demographic variables. These data support the notion that HuR is a key mediator of gemcitabine efficacy in cancer cells, at least in part through its ability to regulate dCK levels posttranscriptionally. We propose that HuR levels in PDA modulate the therapeutic efficacy of gemcitabine, thus serving as a marker of the clinical utility of this common chemotherapeutic agent and a potential target for intervention in pancreatic cancer. [Cancer Res 2009;69(11):4567–72]

The mechanism by which malignant melanoma (MM) cells survive in lymph nodes is poorly understood. One possible mechanism by which MM cells can escape immune surveillance is through upregulation of immunomodulatory enzymes such as indoleamine 2,3-dioxygenase (IDO). In this study, 25 cases of MM lymph node metastases from patients with long and short survival were evaluated for expression of IDO and the number of Forkhead box p3 (FOXP3)-expressing regulatory T cells. Moderate to strong cytoplasmic IDO expression was present in all (15/15) MM lymph node metastases in patients with poor survival. Eight of 10 patients with metastatic MM and long survival were negative or only weakly positive for IDO. Upregulation of IDO in metastatic MM cells was associated with an increased number of regulatory T cells (Tregs). There was a statistically significant association between shorter survival and both a stronger IDO expression (p=0.0019) and a higher number of FOXP3 expressing Tregs (p

The recently discovered indoleamine 2,3-dioxygenase-2 (IDO2) gene has 2 functional polymorphisms that abolish its enzymatic activity. We hypothesize that expression of the IDO2 enzyme in primary pancreatic ductal adenocarcinomas (PDA) can help cancer cells evade immune detection. Because the IDO2 enzyme might be the preferential target of d-1-methyl-tryptophan, a clinical lead inhibitor of IDO currently being evaluated in phase I trials, we sequenced IDO2 in 36 pancreatic specimens and evaluated its expression. We found that 58% (21 of 36) of cases were heterozygous for the R248W polymorphism; 28% (10 of 36) were homozygous wild-type; and only 14% (5 of 36) were homozygous for the functionally inactive polymorphism. As for the Y359STOP polymorphism, we found that 27% (10 of 36) of cases were heterozygous, 62% (22 of 36) were homozygous wild-type, and only 11% (4 of 36) were homozygous for this functionally inactive allele. Ruling out the possibility of compound polymorphic variants, we estimated 75% of our resected patient cohort had an active IDO2 enzyme, with a conservative estimate that 58% of the patients had at least 1 functional allele. IDO2 was expressed in PDA tissue from each genetically polymorphic subgroup. We also detected IDO2 protein expression in the genetically distinct pancreatic cancer cell lines after exposure with interferon-gamma. This is the first study to report IDO2 expression in PDA and related cancers indicating that IDO2 genetic polymorphisms do not negate interferon-gamma-inducible protein expression. Taken together, our data strongly suggest that the clinical lead compound d-1-methyl-tryptophan might be useful in treatment of PDA.

Pancreatic ductal adenocarcinoma (PDA) is the fourth leading cause of cancer-related death in the United States, with a 95% five-year mortality rate. For over a decade, gemcitabine (GEM) has been the established first-line treatment for this disease despite suboptimal response rates. The development of PARP inhibitors that target the DNA damage repair (DDR) system in PDA cells has generated encouraging results. Ubiquitin-specific peptidase 11 (USP11), an enzyme that interacts with the DDR protein BRCA2, was recently discovered to play a key role in DNA double-strand break repair and may be a novel therapeutic target. A systematic high-throughput approach was used to biochemically screen 2,000 U.S. Food and Drug Administration (FDA)-approved compounds for inhibition of USP11 enzymatic activity. Six pharmacologically active small molecules that inhibit USP11 enzymatic activity were identified. An in vitro drug sensitivity assay demonstrated that one of these USP11 inhibitors, mitoxantrone, impacted PDA cell survival with an IC₅₀ of less than 10 nM. Importantly, across six different PDA cell lines, two with defects in the Fanconi anemia/BRCA2 pathway (Hs766T and Capan-1), mitoxantrone is 40- to 20,000-fold more potent than GEM, with increased endogenous USP11 mRNA levels associated with increased sensitivity to mitoxantrone. Interestingly, USP11 silencing in PDA cells also enhanced sensitivity to GEM. These findings establish a preclinical model for the rapid discovery of FDA-approved compounds and identify USP11 as a target of mitoxantrone in PDA. Implications: This high-throughput approach provides a strong rationale to study mitoxantrone in an early-phase clinical setting for the treatment of PDA. Mol Cancer Res; 11(8); 901–11. ©2013 AACR.