Based on the observation that removal of tumors from metastatic organs reversed their chemoresistance, we hypothesized that chemoresistance is induced by extracellular factors in tumor-bearing organs. By comparing chemosensitivity and proteins in different tumors (primary vs. metastases) and different culture systems (tumor fragment histocultures vs. monolayer cultures derived from the same tumor), we found elevated levels of acidic (aFGF) and basic (bFGF) fibroblast growth factors in the conditioned medium (CM) of solid and metastatic tumors. These CM induced broad spectrum resistance to drugs with diverse structures and action mechanisms (paclitaxel, doxorubicin, 5-fluorouracil). Inhibition of bFGF by mAb and its removal by immunoprecipitation resulted in complete reversal of the CM-induced chemoresistance, whereas inhibition͞removal of aFGF resulted in partial reversal. Using CM that had been depleted of aFGF and͞or bFGF and subsequently reconstituted with respective human recombinant proteins, we found that bFGF but not aFGF induced chemoresistance whereas aFGF amplified the bFGF effect. aFGF and bFGF fully accounted for the CM effect, indicating these proteins as the underlying mechanism of the chemoresistance. The FGF-induced resistance was not due to reduced intracellular drug accumulation or altered cell proliferation. We further showed that an inhibitor of aFGF͞bFGF (suramin) enhanced the in vitro and in vivo activity of chemotherapy, resulting in shrinkage and eradication of well established human lung metastases in mice without enhancing toxicity. These results indicate elevated levels of extracellular aFGF͞bFGF as an epigenetic mechanism by which cancer cells elude cytotoxic insult by chemotherapy, and provide a basis for designing new treatment strategies.R esistance of tumor cells to chemotherapy and the limited efficacy of chemotherapy in metastatic disease are two major challenges in patient management. A common resistance mechanism observed in preclinical studies is the overexpression of drug efflux proteins (1-3). However, clinical studies show that inhibition of the drug efflux proteins does not significantly improve the effectiveness of chemotherapy in patients (4, 5), suggesting the existence of other chemoresistance mechanisms.Using the transplantable, metastatic rat prostate MAT-LyLu tumor, we have shown that the antitumor activity of paclitaxel in lymph node metastases was 20-fold lower than in s.c. implanted primary tumors. When the metastatic tumor was reimplanted at the s.c. site, the resistance was lost in the second generation primary tumor but regained in the second generation metastases. We further found that the chemoresistance in metastatic tumors is not due to reduced intracellular drug accumulation or retention (6). These results led to the hypothesis of an epigenetic chemoresistance mechanism that is mediated by extracellular factors present in tumor-bearing organs. The present study tested the hypothesis, identified the factors that induce resistance, and determined the resto...

IntroductionOlaparib, a poly (ADP-ribose) polymerase (PARP) inhibitor, has been found to have therapeutic potential for treating cancers associated with impaired DNA repair capabilities, particularly those with deficiencies in the homologous recombination repair (HRR) pathway. Histone deacetylases (HDACs) are important for enabling functional HRR of DNA by regulating the expression of HRR-related genes and promoting the accurate assembly of HRR-directed sub-nuclear foci. Thus, HDAC inhibitors have recently emerged as a therapeutic agent for treating cancer by inhibiting DNA repair. Based on this, HDAC inhibition could be predicted to enhance the anti-tumor effect of PARP inhibitors in cancer cells by blocking the HRR pathway.MethodsWe determined whether suberoylanilide hydroxamic acid (SAHA), a HDAC inhibitor, could enhance the anti-tumor effects of olaparib on breast cancer cell lines using a cytotoxic assay, cell cycle analysis, and Western blotting. We evaluated how exposure to SAHA affects the expression of HRR-associated genes. The accumulation of DNA double strand breaks (DSBs) induced by combination treatment was assessed. Induction of autophagy was monitored by imaging green fluorescent protein-tagged microtubule-associated protein 1A/1B-light chain 3 (LC3) expression following co-treatment with olaparib and SAHA. These in vitro data were validated in vivo using a human breast cancer xenograft model.ResultsTriple-negative breast cancer cell (TNBC) lines showed heterogeneous responses to the PARP and HDAC inhibitors. Co-administration of olaparib and SAHA synergistically inhibited the growth of TNBC cells that expressed functional Phosphatase and tensin homolog (PTEN). This effect was associated with down-regulation of the proliferative signaling pathway, increased apoptotic and autophagic cell death, and accumulation of DNA damage. The combined anti-tumor effect of olaparib and SAHA was also observed in a xenograft model. These data suggest that PTEN expression in TNBC cells can sensitize the cell response to simultaneous inhibition of PARP and HDAC both in vitro and in vivo.ConclusionOur findings suggest that expression of functional PTEN may serve as a biomarker for selecting TNBC patients that would favorably respond to a combination of olaparib with SAHA. This provides a strong rationale for treating TNBC patients with PTEN expression with a combination therapy consisting of olaparib and SAHA.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-015-0534-y) contains supplementary material, which is available to authorized users.

AKAP12/Gravin, one of the A-kinase anchoring proteins (AKAPs), functions as a kinase scaffold protein and as a dynamic regulator of the b 2 -adrenergic receptor complex. However, the biological role of AKAP12 in cancer development is not well understood. The AKAP12 gene encodes two major isoforms of 305 and 287 kDa (designated AKAP12A and AKAP12B, respectively, in this report). We found that these two isoforms are independently expressed and that they are probably under the control of two different promoters. Moreover, both isoforms were absent from the majority of human gastric cancer cells. The results from methylation-specific PCR (MSP) and bisulfite sequencing revealed that the 5 0 CpG islands of both AKAP12A and AKAP12B are frequently hypermethylated in gastric cancer cells. Treatment with DNA methyltransferase inhibitor and/or histone deacetylase inhibitor efficiently restored the expression of AKAP12 isoforms, confirming that DNA methylation is directly involved in the transcriptional silencing of AKAP12 in gastric cancer cells. Hypermethylation of AKAP12A CpG island was also detected in 56% (10 of 18) of primary gastric tumors. The restoration of AKAP12A in AKAP12-nonexpressing cells reduced colony formation and induced apoptotic cell death. In conclusion, our results suggest that AKAP12A may function as an important negative regulator of the survival pathway in human gastric cancer.

Ldb1 and erythroid partners SCL, GATA-1, and LMO2 form a complex that is required to establish spatial proximity between the ␤-globin locus control region and gene and for transcription activation during erythroid differentiation. Here we show that Ldb1 controls gene expression at multiple levels. Ldb1 stabilizes its erythroid complex partners on ␤-globin chromatin, even though it is not one of the DNA-binding components. In addition, Ldb1 is necessary for enrichment of key transcriptional components in the locus, including P-TEFb, which phosphorylates Ser2 of the RNA polymerase C-terminal domain for efficient elongation. Furthermore, reduction of Ldb1 results in the inability of the locus to migrate away from the nuclear periphery, which is necessary to achieve robust transcription of ␤-globin in nuclear transcription factories. Ldb1 contributes these critical functions at both embryonic and adult stages of globin gene expression. These results implicate Ldb1 as a factor that facilitates nuclear relocation for transcription activation. (Blood. 2010;116(13):2356-2364) IntroductionThe ␤-globin locus control region (LCR) activates the adult globin genes (major and minor) over a distance of approximately 20 kb. Activation is accompanied by the establishment of proximity between these elements 1,2 through any of several mechanisms, including looping, linking, or tethering to a common nuclear substructure. 3 Intrachromosomal associations between genes and enhancers have also been observed in the ␣-globin locus in erythroid cells, 4 at the T H 2, interferon-␥, and major histocompatibility complex loci in T cells, the IgH and Ig loci in B cells, and the growth hormone locus in pituitary cells, among others. 5,6 Furthermore, close interactions can exist between chromosomes to regulate T H 1 and T H 2 cytokine and odorant receptor choice during differentiation. 7,8 How these interactions between regulatory elements and genes form and how they function to increase transcription remain unclear.The erythroid activators EKLF, GATA-1, and FOG-1, as well as the widely expressed nuclear factor lim domain binding protein 1 (Ldb1) are required to establish spatial proximity between the LCR and ␤-globin gene. [9][10][11] Ldb1, in a complex with LMO2 and DNA-binding partners GATA-1 and SCL, occupies the LCR and ␤-globin promoter in induced murine erythroleukemia (MEL) cells and fetal liver erythroid cells of mice, and shRNA-mediated reduction of Ldb1 prevents interaction between these elements. Ldb1 is the murine homolog of Drosophila melanogaster Chip and is proposed to be a general "facilitator" of long-range chromatin interactions. 12 Proximity between distant elements that are Ldb1-binding sites may depend on Ldb1 self-interaction, although this has not been firmly established. 13,14 It is not known whether all functions of Ldb1 involve long-range associations or whether all Ldb1 functions in erythroid cells are carried out through its interaction with DNA-binding partner GATA-1. However, there is a high correlation between pos...