Targeting DNA repair for cancer treatment: Lessons from PARP inhibitor trials

Ionizing radiation is frequently used to treat solid tumors,as it causes DNA damage and kill cancer cells.However,damaged DNA is repaired involving poly-(ADP-ribose)polymerase-1(PARP-1)causing resistance to radiation therapy.Thus,PARP-1 represents an important target in multiple cancer types,including prostate cancer.PARP is a nuclear enzyme essential for single-strand DNA breaks repair.Inhibiting PARP-1 is lethal in a wide range of cancer cells that lack the homologous recombination repair(HR)pathway.This article provides a concise and simplified overview of the development of PARP inhibitors in the laboratory and their clinical applications.We focused on the use of PARP inhibitors in various cancers,including prostate cancer.We also discussed some of the underlying principles and challenges that may affect the clinical efficacy of PARP inhibitors.

Bioactive lipids in cancer stem cells

Tumours are known to be a heterogeneous group of cells,which is why they are difficult to eradicate.One possible cause for this is the existence of slow-cycling cancer stem cells(CSCs)endowed with stem cell-like properties of self-renewal,which are responsible for resistance to chemotherapy and radiotherapy.In recent years,the role of lipid metabolism has garnered increasing attention in cancer.Specifically,the key roles of enzymes such as stearoyl-CoA desaturase-1 and 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase in CSCs,have gained particular interest.However,despite accumulating evidence on the role of proteins in controlling lipid metabolism,very little is known about the specific role played by lipid products in CSCs.This review highlights recent findings on the role of lipid metabolism in CSCs,focusing on the specific mechanism by which bioactive lipids regulate the fate of CSCs and their involvement in signal transduction pathways.

Establishment of organoid models for pancreatic ductal adenocarcinoma and screening of individualized therapy strategy

Background:Patients with pancreatic ductal adenocarcinoma(PDAC)who undergo surgical resection and receive effective chemotherapy have the best chance for longterm survival.Unfortunately,because of the heterogeneity of pancreatic cancer,it is difficult to find a personalized treatment strategy for patients.Organoids are ideal preclinical models for personalized medicine.Therefore,we explore the cultivation conditions and construction methods of PDAC organoid models to screen the individualized therapy strategy.Methods:Fresh PDAC tissues from surgical resection were collected and digested with digestive enzymes;then the tumor cells were embedded in Matrigel with a suitable medium to establish the PDAC organoid models.The genetic stability of the organoids was analyzed using whole exon sequencing;hematoxylin and eosin staining and immunohistochemistry of organoids were performed to analyze their consistency with the pathological morphology of the patient's tumor tissue;After 2 days of organoid culture,we selected four commonly used clinical chemotherapy drugs for single or combined treatment to analyze drug sensitivity.Results:Two cases of PDAC organoid models were successfully established,and the results of their pathological characteristics and exome sequencing were consistent with those of the patient's tumor tissue.Both PDAC organoids showed more sensitivity to gemcitabine and cisplatin,and the combined treatment was more effective than monotherapy.Conclusion:Both organoids better retained the pathological characteristics,genomic stability,and heterogeneity with the original tumor.Individual PDAC organoids exhibited different sensitivities to the same drugs.Thus,this study provided ideal experimental models for screening individualized therapy strategy for patients with PDAC.

Comparisons of biophysical properties and bioactivities of mono-PEGylated endostatin and an endostatin analog

Background:Endostatin(ES) is a well-established potent endogenous antiangiogenic factor.An ES variant,called zinc-binding protein-ES(ZBP-ES),is clinically available;however,its use is limited by rapid renal clearance and short residence time.PEGylation has been exploited to overcome these shortcomings,and mono-PEGylated ES(called M_2ES) as well as mono-PEGylated ZBP-ES(MZBP-ES) are developed in our study.This study aimed to compare the biophysical properties and biological effects of M_2ES and MZBP-ES to evaluate their druggability.Methods:Circular dichroism and tryptophan emission fluorescence were used to monitor the conformational changes of M_2ES and MZBP-ES.Their resistance to trypsin digestion and guanidinium chloride(GdmCl)-induced unfolding was examined by Coomassie staining and tryptophan emission fluorescence,respectively.The biological effects of M_2ES and MZBP-ES on endothelial cell migration were evaluated using Transwell migration and wound healing assays,and the uptake of M_2ES and MZBP-ES in endothelial cells was also compared by Western blotting and immunofluorescence.Results:Structural analyses revealed that M_2ES has a more compact tertiary structure than MZBP-ES.Moreover,M_2ES was more resistant to trypsin digestion and GdmCI-induced unfolding compared with MZBP-ES.In addition,although M_2ES and MZBP-ES showed comparable levels of inhibiting transwell migration and wound healing of endothelial cells,M_2ES displayed an increased ability to enter cells compared with MZBP-ES,possibly caused by the enhanced interaction with nucleolin.Conclusions:M_2ES has a more compact tertiary structure,is more stable for trypsin digestion and GdmCI-induced unfolding,exhibits increased cellular uptake and shows equivalent inhibitory effects on cell migration relative to MZBP-ES,indicating that M_2ES is a more promising candidate for anticancer drug development compared with MZBP-ES.

Chimeric oncogenic interferon regulatory factor-2 (IRF-2): Degradation products are biologically active

Interferon Regulatory Factor-2 (IRF-2) belongs to IRF family, was identified as a mammalian transcription factor involved in Interferon beta (IFNβ) gene regulation. Besides that IRF-2 is involved in immunomodulation, hematopoietic differentiation, cell cycle regulation and oncogenesis. We have done molecular sub-cloning and expression of recombinant murine IRF-2 as GST (Glutathione-S-Transferase)- IRF-2 fusion protein in E. coli/XL-1blue cells. Recombinant IRF-2 with GST moiety at N-terminus expressed as GST-IRF-2 (~66 kd) in E. coli along with different low molecular mass degradation products revealed approximately 30, 42, 60 and 62 kd by SDS-PAGE and Western blot, respectively. We further confirm that degradation takes place at C-terminus of the fusion protein not at N-terminus as anti-GST antibody was detecting all bands in the immunoblot. The recombinant IRF-2 was biologically active along with their degradation products in terms of their DNA binding activity as assessed by Electrophoretically Mobility Shift Assay (EMSA). We observed three different molecular mass DNA/protein complexes (1 - 3) with Virus Response Element (VRE) derived from human Interferon IFNβ gene and five different molecular mass complexes (1 - 5) with IRF-E motif (GAAAGT)4 in EMSA gel. GST only expressed from empty vector did not bind to these DNA elements. To confirm that the binding is specific, all complexes were competed out completely when challenged with 100-X fold molar excess of IRF-E oligo under cold competition. It means degradation products along with full-length protein are able to interact with VREβ as well as IRF-E motif. This means degradation products may regulate the target gene (s) activation/repression via interacting with VRE/IRF-E.

Pharmacometrics of nutraceutical sulforaphane and its implications in prostate cancer prevention

Sulforaphane(SFN), found in broccoli and other cruciferous vegetables, has a beneficial effect in chemoprevention of prostate cancer, whose incidence and associated mortality have gradually increased worldwide. There is great enthusiasm for bench-to-bedside development of SFN as a potent chemopreventive agent, possibly alone or as an adjunct to existing chemotherapy regimens, in the oncology care setting to reduce toxicity of chemotherapeutics and potentially enhance their cancer cell-kill efficacy. In this review, we appreciate existing knowledge on SFN using a pharmacometrics approach, which is fast becoming a gold standard in discovery research and validation of New Chemical Entities and New Biological Entities in pharmaceutical industry. We discuss the epistemology of SFN target engagement and quantitative systems pharmacology with due emphasis on mechanistic pharmacology, pharmacodynamics, pharmacogenomics and metabolism of SFN. In addition, we explore the quantitative freeway to SFN translational medicine by assessing the preclinical and clinical PK/metabolism aspects of SFN that form the cornerstone of SFN pharmacometric evaluation, as well as the promise of SFN in prostate cancer. Taken together, we share perspectives on the exciting developments in translational cancer chemoprevention, with emphasis on the pharmacometric aspects, of the nutraceutical SFN which is currently in clinical trials, and suggest that the pharmacometric approach holds great promise in the SFN translational pharmacology paradigm for prostate cancer.

Differential signaling regulatory networks governing hormone refractory prostate cancers

To understand the organization of the biological networks that might potentially govern the pathogenesis of hormone refractory prostate cancer(HRPC), we investigated the transcriptional circuitry and signaling in androgen-dependent 22Rv1 and MDA PCa 2b cells, androgen- and estrogen-dependent LNCaP cells, and androgen-independent DU 145 and PC-3 prostate cancer(PCa) cell lines. We used microarray analyses, quantitative real-time PCR, pathway prediction analyses, and determination of Transcription Factor Binding Site(TFBS) signatures to dissect HRPC regulatory networks. We generated graphical representations of global topology and local network motifs that might be important in prostate carcinogenesis. Many important putative biomarker ‘target hubs' were identified in the current study including AP-1, NF-?B, EGFR, ERK1/2, JNK, p38 MAPK, TGF beta, VEGF, PDGF, CD44, Akt, PI3 K, NOTCH1, CASP1, MMP2 and AR. Our results suggest that complex cellular events including autoregulation, feedback loops and cross-talk might govern progression from early lesion to clinically diagnosed PCa, as well as metastatic potential of pre-existent high-grade prostate intraepithelial neoplasia(HG-PIN) and/or advancement to HRPC. The identification of TFBS signatures for TCF/LEF, SOX9 and ELK1 in the regulatory elements suggests additional biomarkers for the potential development of chemopreventive/therapeutic strategies against PCa. Taken together, in this study, we have identified putative biomarker ‘target hubs' in the architecture of PCa signaling networks, and investigated TFBS signatures that might enhance our understanding of key regulatory nodes in the progression and pathogenesis of HRPC.

BCL9 regulates CD226 and CD96 checkpoints in CD8^(+)T cells to improve PD-1 response in cancer

To date,the overall response rate of PD-1 blockade remains unsatisfactory,partially due to limited understanding of tumor immune microenvironment(TIME).B-cell lymphoma 9(BCL9),a key transcription co-activator of the Wnt pathway,is highly expressed in cancers.By genetic depletion and pharmacological inhibition of BCL9 in tumors,we found that BCL9 suppression reduced tumor growth.

Endostatin specifically targets both tumor blood vessels and lymphatic vessels

Endostatin,a 20 kDa C-terminal fragment of collagen XVIII,was first identified as a potent angiogenic inhibitor.The anti-angiogenic function of endostatin has been well documented during the past decade.Recently,several studies demonstrated that endostatin also inhibits tumor lymphangiogenesis and lymphatic metastasis.However,the exact mechanism that endostatin executes its anti-angiogenic and anti-lymphangiogenic functions remains elusive.In the current mini-review,we briefly summarize recent novel findings,including the functions of endostatin targeting not only angiogenesis but also lymphangiogenesis,and the underlying mechanism by which endostatin internalization regulates its biological functions.

The phytochemical, biological, and medicinal attributes of phytoecdysteroids: An updated review

The phytoecdysteroids(PEs)comprise a large group of biologically-active plant steroids,which have structures similar to those of insect-molting hormones.PEs are distributed in plants as secondary metabolites that offer protection against phytophagus(plant-eating)insects.When insects consume the plants containing these chemicals,they promptly molt and undergo metabolic destruction;the insects eventually die.Chemically,ecdysteroids are a group of polyhydroxylated ketosteroids that are structurally similar to androgens.The carbon skeleton of ecdysteroids is termed as cyclopentanoperhydro-phenanthrene with aβ-side chain at carbon-17.The essential characteristics of ecdysteroids are a cis-(5β-H)junction of rings A and B,a 7-en-6-one chromophore,and a trans(14α-OH)junction of rings C and D.Plants only synthesize PEs from mevalonic acid in the mevalonate pathway of the plant cell using acetyl-CoA as a precursor;the most common PE is 20-hydroxyecdysone.So far,over 400 PEs have been identified and reported,and a compilation of 166 PEs originating from 1998 has been previously reviewed.In the present review,we have summarized 212 new PEs reported between 1999 and 2019.We have also critically analyzed the biological,pharmacological,and medicinal properties of PEs to understand the full impact of these phytoconstituents in health and disease.

Xanthophyll β-cryptoxanthin treatment inhibits hepatic steatosis without altering vitamin A status inβ-carotene 9',10'-oxygenase knockout mice

Background:β-cryptoxanthin(BCX),one of the major carotenoids detected in human circulation,can protect against the development of fatty liver disease.BCX can be metabolized throughβ-carotene-15,15'-oxygenase(BCO1)andβ-carotene-9',10'-oxygenase(BCO2)cleavage pathways to produce both vitamin A and apo-carotenoids,respectively,which are considered important signaling molecules in a variety of biological processes.Recently,we have demonstrated that BCX treatment reduced hepatic steatosis severity and hepatic total cholesterol levels in both wide type and BCO1^(-/-)/BCO2^(-/-)double knock out(KO)mice.Whether the protective effect of BCX is seen in single BCO2^(-/-)KO mice is unclear.Methods:In the present study,male BCO2^(-/-)KO mice at 1 and 5 months of age were assigned to two groups by age and weight-matching as follows:(I)-BCX control diet alone(AIN-93 purified diets);(II)+BCX 10 mg(supplemented with 10 mg of BCX/kg of diet)for 3 months.At 4 and 8 months of age,hepatic steatosis and inflammatory foci were evaluated by histopathology.Retinoids and BCX concentrations in liver tissue were analyzed by high-performance liquid chromatography(HPLC).Hepatic protein expressions of SIRT1,acetylated and total FoxO1,PGC1α,and PPARαwere determined by the Western blot analysis.Real-time PCR for gene expressions(MCAD,SCD1,FAS,TNFα,and IL-1βgene expression relative toβ-actin)was conducted in the liver.Results:Steatosis was detected at 8 months but not at 4 months of age.Moreover,BCX supplementation significantly reduced the severity of steatosis in the livers of BCO2KO mice,which was associated with changes in hepatic SIRT1 acetylation of FOXO1,PGC1αprotein expression and PPARαprotein expression in BCO2^(-/-)KO mice.HPLC analysis showed that hepatic BCX was detected in BCX supplemented groups,but there were no differences in the hepatic levels of retinol and retinyl palmitate(RP)among all groups.Conclusions:The present study provided experimental evidence that BCX intervention can reduce liver steatosis independent of BCO2.

Elucidation of regulatory interaction networks underlying human prostate adenocarcinoma

The incidence of prostate cancer is rising in the Asia-Pacific region as well as other countries. Androgen-ablation therapy is clinically useful in the androgen-dependent phenotype however, many patients progress to hormone refractory prostate cancer that is difficult to treat and needs newer interventions that are more effective. The objective of this study was to determine functionally-relevant biological networks, to appreciate the potential crosstalk between signaling members, and to identify biomarker signatures in prostate cancer. We used microarray analyses to identify key genes that were upregulated or down regulated at least five-fold in human prostate cancer and constructed canonical interaction networks that are important in prostate cancer through metabolomics analyses. Our prostate cancer network architecture revealed several key biomarkers including ERK1/2, JNK, p38, MEK, PI3 K, NFκB, AP-1, 14-3-3, VEGF, PDGF, Rb, WNT8 A, WNT10 A, CD44, ESR2, FSH and LH. Furthermore, the top ten transcription factors identified by TFBS-association signature analysis in the regulatory elements of co-regulated biomarkers were delineated, which may crosstalk with upstream or downstream genes elicited in our network architecture. Taken together, our results demonstrate that the regulatory interaction networks in prostate cancer provide a universal view of crosstalk between important biomarkers, i.e., key players in the pathogenesis of this disease. This will facilitate more rapid screening of functional biomarkers in early/intermediate drug discovery.

Nrf2-mediated antioxidant and detoxifying enzyme induction by a combination of curcumin and sulforaphaneNrf2-mediated antioxidant and detoxifying enzyme induction by a combination of curcumin and sulforaphane

The dietary phytochemicals curcumin(CUR) and sulforaphane(SFN) have shown remarkable cancer chemopreventive effects in many model systems. This study was designed to investigate the induction of Nrf2-mediated antioxidant enzymes by combining doses of CUR and SFN and the effect of their combination on the Nrf2-ARE(antioxidant response element) response in Hep G2-C8 cells. We hypothesized that the combination of the polyphenol CUR and the isothiocyanate SFN could enhance the induction of AREs and Nrf2-target enzymes. Hep G2-C8 cells were treated with a combination of low doses of CUR, SFN or both. The induction of Nrf2-mediated antioxidant and phase II detoxifying enzymes–heme oxygenase-1(HO-1) and UDP-glucuronosyltransferase-1A(UGT1A)–was measured by real-time RT-PCR and western blotting. ARE-luciferase activity was also quantified. Low doses of CUR(10 μM) and SFN(12.5 μM) significantly induced the expression of HO-1 and UGT1A1 proteins. Through the use of chemical inhibitors of mR NA and protein synthesis, the combination of CUR and SFN was shown to affect the transcriptional regulation of both HO-1 and UGT1A1. Additionally, the combination of CUR and SFN synergistically induced the expression of Nrf2- and ARE-luciferase activity in Hep G2-C8 cells. Thus, CUR and SFN at low concentrations augment therapeutic effects in Hep G2-C8 cells. The enhanced ARE-luciferase activity of combined CUR and SFN treatment could partly explain the significant induction of the Nrf2-target enzymes HO-1 and UGT1A1. Taken together, our results suggest that combining low doses of CUR and SNF could be a promising strategy for cancer chemoprevention in humans.