bluuebunny/crossref_metadata_2025_split huggingface dataset:
Polymorphisms of GSTP1, GSTT1, GSTM1, MTHFR, TS, ERCC1, and ERCC2 in metastatic colorectal cancer treated by first-line mFOLFOX6 chemotherapy
Authors: Ishibashi K. , Okada N. , Ishiguro T. , Yokoyama M. , Miyazaki T. , Sano M. , Yamada H. , Ishida H. 
Published: August 2020

e14628 Background: It was reported that determining functional polymorphisms of genes involved in drug-metabolising pathways and DNA repair may be useful for predicting the response to 5-FU/oxaliplatin chemotherapy in Caucasian patients with metastatic colorectal cancer. This study was performed to examine whether determining these polymorphisms had any clinical value in Asian patients with colorectal cancer receiving 5-FU/oxaliplatin therapy. Methods: Genomic DNA was extracted from peripheral blood lymphocytes (n=25) or colonic mucosa (n=47) in Japanese patients with metastatic colorectal cancer who were receiving first-line therapy with the modified FOLFOX6 regimen followed by FOLFIRI (n=42). Polymorphisms of 5 genes involved in drug metabolism (glutathione S-transferase (GST) P1 (IIe 105 Val), GSTT1 deletion, and GSTM1 deletion, methylenetetrahydrofolate reductase (MTHFR) (Ala 677 Val), and a 6-base pair (bp) deletion in the 3’-untranslated region (UTR) of thymidylate synthase (TS)), and polymorphisms of two DNA repair genes (excision repair cross complementing group 1 (ERCC1): Asp 118 Asn and ERCC2: Lys 751 Gln) were assessed in these patients by PCR-RFLP or the invader technique. Correlations between polymorphisms of these genes and the response to therapy were evaluated. Results: The distribution of the genotypes of GSTP1, GSTT1, TS, ERCC1, and ERCC2 in the present Japanese patients (but not that of GSTM1 or MTHFR), differed significantly from the distribution of these genotypes in a Caucasian population. The response rate and progression-free survival were not correlated with any of the functional polymorphisms investigated. However, patients who had both alleles containing the 6-bp nucleotide fragment in the 3’UTR of TS showed significantly shorter overall survival than those who had at least one allele without the 6-bp nucleotide fragment (p=0.03). Conclusions: These results suggest that 3’UTR polymorphism of TS may be an important predictor of overall survival for Japanese patients with metastatic colorectal cancer receiving first-line 5-FU/oxaliplatin therapy. No significant financial relationships to disclose.
Mouse prostate tumor inhibition via disruption of murine telomerase: In vivo and in vitro data for a new preclinical cancer model
Authors: Xu T. , Xu Y. , Lao R. , He K. , Xue L. , Goldkorn A. 
Published: August 2020

e14631 Background: Telomerase-interference (TI), a novel therapeutic strategy, exploits the high telomerase activity in prostate cancer by introducing a mutated telomerase RNA (MT-Ter) that encodes toxic telomeres. Until now, TI has been tested by targeting human telomerase in tumor cells xenografted into immuno-deficient mice, an inadequate model for predicting efficacy and toxicity. We designed and validated 2 new TI gene constructs that specifically target murine telomerase RNA (mTER), enabling the study of TI in preclinical mouse models that are immuno-competent and that develop endogenous prostate tumors. Methods: We designed 2 constructs and cloned them into a lentiviral delivery system: MT-mTER and siRNA against wild type mTer (α-mTer-siRNA). Using a mouse prostate cancer cell line, E4, we tested the 2 constructs for expression (RT-PCR), telomerase activity (TRAP), and biologic activity (53bp1 DNA damage staining, MTS growth assay, TUNEL and caspase apoptosis assays), as well as in vivo efficacy (NOD-SCID allografts). Results: We confirmed MT-mTER expression (∼50-fold) and showed that α-mTer-siRNA specifically depleted WT-mTER (80% reduction) but not MT-mTER when the 2 constructs are co-expressed; thus, the 2 constructs in combination effectively substituted MT-mTer for WT-mTer in the mouse prostate cancer cells. MT-mTER caused mutant telomeric repeats (TTTGGG instead of TTAGGG) to be added to the ends of telomeres, resulting in rapid telomeric uncapping marked by 53bp1 DNA damage foci (an average 7.5 foci/cell vs. 1.4 foci/cell in vector control). This, in turn, led to rapid and significant apoptosis (>90% TUNEL and caspase +) and growth inhibition in vitro (90% reduction by MTS) and in vivo (75% reduction in tumor allograft size). Conclusions: We successfully designed and validated MT-mTer and α-mTer-siRNA, 2 novel gene constructs that specifically target and co-opt murine telomerase activity within mouse prostate cancer cells. These constructs offer a significant advantage, as they can be used to investigate TI in immuno-competent mice that develop prostate cancer, thereby modeling actual human disease and testing TI-based therapies in a much more informative and authentic manner. No significant financial relationships to disclose.
Effect of lenalidomide on the antiproliferative effect of gemcitabine against pancreatic tumor cells and on immune-mediated pancreatic cancer cell death
Authors: Wu L. , Liu W. , Galustian C. , Schafer P. , Dalgleish A. G. , Bartlett J. B. 
Published: August 2020

e14635 Background: Lenalidomide is an immunomodulatory and anti-angiogenic agent that has demonstrated activity against a range of hematological malignancies. Despite evidence of direct anti-proliferative activity against hematological cells in vitro, there is no evidence of single agent direct activity against solid tumor cells in vitro. To take advantage of its known immune-enhancing properties alongside direct anti-tumor agents, lenalidomide is being advanced in solid tumor indications in combination with other agents. There are few data regarding the combination of lenalidomide and standard of care chemotherapeutic agents, such as gemcitabine. Methods: Here, we assess the effects of lenalidomide alone, and in combination with gemcitabine, on pancreatic cancer cell growth and survival, and the ability of lenalidomide to enhance the ability of human PBMC to kill allogeneic pancreatic tumor cells (BxPC3, PANC-1 and MiaPaCa) in a PBMC:tumor cell co-culture model. Results: Lenalidomide alone had no effect on the proliferation of pancreatic cancer cells (BxPC-3 and Panc-1) whereas gemcitabine had moderate anti-proliferative activity. With combination therapy there was clear synergistic enhancement of anti-proliferative activity in both cell lines and additive effects were observed in a BxPC-3 xenograft mouse model of pancreatic cancer. About 20% of tumor cells were sensitive to immune-mediated cell death and, for BxPC3, this was increased significantly in the presence of lenalidomide. Lenalidomide significantly and dose-dependently enhanced immune-mediated killing (both T and NK cells are required for tumor cell killing in this model). For PANC-1 and MiaPaCa, immune-mediated killing was also increased by lenalidomide, albeit non-significantly. Conclusions: These results suggest that, in addition to anti-angiogenic and other effects within the tumor microenvironment, lenalidomide may act as an immune adjuvant to enhance the recognition and apoptosis of tumor cells by host T and NK cells. These studies support the potential utility of lenalidomide in combination with chemotherapeutic agents, gemcitabine in particular, in the treatment of patients with solid tumors including pancreatic cancer. [Table: see text]
Methylnaltrexone potentiates the antiangiogenic effects of the mTOR inhibitor, temsirolimus
Authors: Singleton P. A. , Moss J. , Garcia J. G. 
Published: August 2020

e14636 Background: Angiogenesis is important in cancer growth and metastasis. Recent therapeutic interventions cancer, include drugs such as temsirolimus that target both tumor growth and angiogenesis including the mammalian target of rapamycin (mTOR) inhibitor. Previously, we have demonstrated that the peripheral opioid antagonist methylnaltrexone (MNTX) potentiates the effects of bevacizumab and 5FU on endothelial cell (EC) migration and proliferation. Since mTOR inhibitor therapy is also associated with significant side effects, we examined potential adjunctive agents that could reduce the therapeutic dose and improve the therapeutic window for this approach. Methods: We determined the effect(s) of MNTX, naloxone and/or temsirolimus on VEGF-induced human pulmonary microvascular EC proliferation and migration assays. We used siRNA and specific inhibitors to analyze Src, Akt and mTOR regulation. Results: MNTX inhibited EC VEGF-induced proliferation and migration with an IC50 of ∼100 nM. Adding 10 nM MNTX to EC shifted the IC50 of temsirolimus inhibition of VEGF-induced proliferation from ∼10 nM to ∼1 nM. Further, adding 10 nM MNTX shifted the IC50 of temsirolimus on inhibition of EC migration from ∼50 nM to ∼10 nM. These synergistic effects were not observed with naltrexone, a tertiary mu opioid receptor antagonist. On a mechanistic level, we observed that treatment of human EC with MNTX, but not naltrexone, increased protein tyrosine phosphatase (PTP) activity which was independent of mu opioid receptor expression. Treatment of human EC with the PTP inhibitor, 3,4-Dephostatin, inhibited both the synergy between MNTX and temsirolimus and increased VEGF-induced tyrosine phosphorylation of Src with enhanced PI3 kinase and mTOR complex 2-dependent phosphorylation of Akt and subsequent activation of mTOR complex 1 (temsirolimus target), while silencing Src, Akt or mTOR complex 2 components blocked VEGF-induced angiogenic events. Conclusions: Synergy of MNTX with mTOR inhibitors may have therapeutic implications which await clinical proof-of-concept. [Table: see text]
Investigation on the mechanism of dichloroacetate (DCA) induced apoptosis in breast cancer
Authors: Ko L. , Allalunis-Turner J. 
Published: August 2020

e14637 Background: DCA is a generic, orally available, small molecule metabolic modulator that has an established history in the treatment of mitochondrial diseases and lactic acidosis. DCA inhibits pyruvate dehydrogenase kinase (PDK), an inhibitor of pyruvate dehydrogenase, a key enzyme in glucose metabolism. DCA preferentially shifts glucose metabolism in cancer cells from glycolysis to glucose oxidation, reversing the unique aerobic glycolysis found in solid tumors. DCA induced apoptosis in cancer cells as evidenced by the efflux of cytochrome c and apoptosis-inducing factor from the mitochondria. Recent studies suggested apoptosis induction by DCA in glioblastoma, endometrial, prostate, and non-small cell lung cancers. In this study we attempt to establish a link between DCA and apoptosis in breast cancer cell lines. Methods: Six breast cancer cell lines were investigated (BT474, MCF-7, MDA-MB231, MDA- MB468, SKBR3, T47D). Quantitative real-time polymerase chain reaction (RT-PCR) was performed using Taqman probes to identify increased DNA templates of PDK isoforms 1–4, using DCA concentrations from 0 to 20mM. Western blots were performed to identify increased expression of PDK isoforms and correlate findings with Taqman. MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)- 2-(4-sulfophenyl)-2H-tetrazolium) assays were performed to measure decreased mitochondrial activity and cytotoxicity. Flow cytometry using annexin V and propidium iodide was performed to measure apoptosis and cell death. Results: RT-PCR showed increased DNA expression of all PDK isoforms in MDA-MB231 cells after DCA treatment. The effect was most pronounced at 10mM concentration. 10mM of DCA also increased DNA expression of all PDK isoforms in MCF-7 cells, and PDK1 in T47D cells. MTS assays showed increased cell kill and decreased mitochondrial activity in all six cell lines, with IC50 ranging between 20mM and 30 mM. Flow cytometry showed increased apoptosis induced by DCA at IC50 for BT474 and MCF-7 cell lines. Conclusions: Apoptosis appears to play a role in the mechanism of DCA in breast cancer, with increased PDK isoform expressions, cytotoxicity and decreased mitochondrial activity. Data from flow cytometry suggested DCA-induced apoptosis in two cell lines. No significant financial relationships to disclose.
MicroRNAs 15a and 16–1 regulates tumor proliferation in multiple myeloma
Authors: Roccaro A. M. , Sacco A. , Leleu X. , Thompson B. , Azab A. , Runnels J. , Lin C. , Witzig T. , Anderson K. , Ghobrial I. 
Published: August 2020

e14640 Background: MicroRNAs (miRNAs) are non-coding RNAs that act as negative regulators of gene expression. They have been described to play roles in solid tumors and hematologic malignancies. However the role of miRNAs in multiple myeloma (MM) has not been yet fully described. Methods: We performed miRNA-profiling of primary bone marrow-derived CD138+ MM cells, compared to their normal cellular counterparts and validated data by qRT-PCR. In vitro and in vivo functional studies were performed on miRNA-15a- and -16–1-precursors-transfected MM cells. Effect of miRNA-15a and -16–1 on signaling cascades have been evaluated by western blot and immunofluorescence. NF-kB activity has been studied using the Active Motif TransAM kit. In vivo MM cell growth has been evaluated by either using an in vivo imaging model or bioluminescence. Angiogenesis has been studied both in vitro and in vivo using the chorioallantoic membrane model. Results: We identified a MM-specific miRNA signature characterized by down-expression of miRNA-15a, -16–1 and over-expression of miRNA-222/-221/-382/-181a/-181b (P<0.01). Data were confirmed by qRT-PCR on matched samples. Predicted targets for the decreased miRNAs in MM patients included pro-angiogenic cytokines, oncogenes, cell cycle regulators, NFkB activators. Conversely, predicted targets for the increased miRNAs included cell cycle inhibitors, suppressors of cytokine signaling, and pro-apoptotic factors. We demonstrated that miRNA-15a and - 16–1 regulate proliferation and growth of MM cells. Indeed, transfected cells showed decreased DNA synthesis; decreased cyclinD1/cyclinD3/CDC25a/pRb protein expression; phase G1 cell cycle arrest. Moreover, transfected cells showed inhibition of NFkB pathway as shown by reduced p65-/p50-/p52-NFkB activities; downregulation of p-p65/p50/p52 nuclear protein level; upregulation of cttosolic phospho-IkB; and inhibited translocation of p-p65 from the cytolplasm to the nucleus. Similarly, inhibition of MM cell growth was confirmed in vivo; and anti-angiogenic properties of miRNA-15a and -16–1 were demonstrated both in vitro and in vivo. Conclusions: These data indicate that miRNAs play a pivotal role in the biology of MM; providing the basis for the development of new miRNA-based targeted therapies in this disease. No significant financial relationships to disclose.
Preclinical study of lithium sensitivity in human breast PTEN knock out cell lines
Authors: Higgins M. J. , Abukhdeir A. M. , Gustin J. , Vitolo M. I. , Bachman K. E. , Park B. H. 
Published: August 2020

e14641 Background: Mutations of PIK3CA, the catalytic subunit of phosphatidylinositol 3-kinase (PI3K), are found in ∼25% of breast cancers. The tumor suppressor PTEN reverses the effects of PI3K and genetic alterations leading to loss or reduced function of PTEN also occur at high frequency in breast cancers. We have used somatic cell gene targeting to create paired isogenic cell lines with critical genetic alterations as their only differentiating factor. Our lab has introduced two “hotspot” oncogenic PIK3CA mutations into the MCF-10A human breast epithelial cell line. This study led to the discovery that lithium, an FDA approved therapy for bipolar disorders, has selective anti-neoplastic properties against human breast cancer cell lines containing oncogenic PIK3CA mutations. Due to the opposing interactions between PI3K and PTEN, we are now studying whether loss of PTEN also confers sensitivity to lithium using human somatic cell knock outs. Methods: Targeting vectors were created and used to knock out the PTEN gene in MCF-10A. Biochemical and phenotypical characterizations were performed. Paired isogenic cell lines were exposed to lithium to evaluate heterozygous and homozygous loss of PTEN as a predictive biomarker of response. Proteomic and phenotypic changes were measured and assessed. Results: Homozygous loss of PTEN (PTEN-/-) resulted in a transformed phenotype enabling growth in the absence of exogenous growth factors, a hallmark of malignant potential. Preliminary results suggest that treatment of PTEN-/- cells with lithium leads to a decrease in cell proliferation with selectivity for mutant PTEN knock out cell lines, although the effects are not as pronounced as that seen with mutant PIK3CA knock in cells. Conclusions: Our preliminary results suggest that loss of PTEN predicts for sensitivity to lithium in breast cell lines in a manner similar to that seen with oncogenic PIK3CA. We hypothesize that differences in the magnitude of response to lithium may be the result of varied levels of non-PI3K-AKT-mTOR activation between the PIK3CA knock in cell lines and the PTEN knock out cells. No significant financial relationships to disclose.
A pilot study to compare FXR expression in normal and malignant tissue in receptor-positive early-stage breast cancer
Authors: Kounalakis N. , Lau S. , Darling D. , Palomares M. , Senthil M. , Lai L. 
Published: August 2020

e14645 Background: Farnesoid X receptor (FXR), a nuclear receptor, is a ligand dependent transcriptional factor regulating cholesterol and carbohydrate metabolism. Recently, FXR was shown to have a contributing role in colorectal cancer. We hypothesize that FXR expression changes from normal to premalignant to malignant tissue in patients with breast cancer. Methods: We identified 16 paired formaldehyde fixed, paraffin embedded tissue (normal, premalignant, and malignant) from patients with receptor positive, early stage breast cancer. Clinical information was extracted from a prospective database initiated in 2006 under institutional approval. Immunohistochemical staining of FXR using a validated polyclonal antibody was completed with appropriate positive and negative controls. The slides were graded independently by two investigators using an agreed upon scale to detect the percentage of positively stained cells to the nearest 10th percentile. Statistical analysis was performed by ANOVA and Student's t-test. A p-value of 0.05 was considered significant in all analyses. Results: Normal tissue and invasive cancer was identified in all 16 patient specimens. Of the 16 invasive cancers, 12 were ductal and 4 were lobular. 8/16 (50%) of the specimens also contained non-invasive cancer. 5/16 patients (31%) had N1 disease. FXR expression did not correlate with grade, histology, stage, or lymph node status. However, FXR expression increases with malignant transformation of the breast cancer cell. The mean percentage of cells staining positive for FXR in normal breast tissue was 58%, non-invasive 72% and invasive 79%. FXR staining in normal breast tissue was significantly less when compared to both invasive and noninvasive cancer (p< 0.007). Conclusions: FXR expression is upregulated in breast cancer when compared with expression in normal tissue and appears to progressively increase along the continuum of malignancy. Our pilot study results warrant further evaluation into FXR as a predictive biomarker for breast cancer, given the ability to target FXR via development of non-toxic oral ligands. No significant financial relationships to disclose.
An open-label phase II trial to assess to efficacy of sunitinib as an antieffusion agent in patients with malignant peritoneal and/or pleural effusions
Authors: Trinh X. , van Dam P. , Tjalma W. , Wojtasik A. , Lybaert W. , Rasschaert M. , Rutten A. , Prové A. , Vermeulen P. , Dirix L. 
Published: August 2020

e14647 Background: Pleural and peritoneal effusions are common in advanced malignant disease and relative resistant to traditional treatments. VEGF-A is considered to be a crucial factor in the biology of accumulation of malignant effusions (ME). Sunitinib is a multi-targeted TKI with a high potency of inhibiting VEGFR2. A treatment that could symptomatically reduce ME would be useful, regardless of its anti-tumoural or anti-angiogenic effect. Methods: Refractory patients with symptomatic ME were enrolled to receive 2x6 weeks of daily 37.5mg sunitinib regardless of their primary histology or site. Outcome was measured by time to re-puncture or by measurement of volume ME by CT-scan every 6 weeks or at the end of treatment. Puncture fluids and blood samples were collected for extensive biomarker analysis (VEGFA&C, VEGFR1–3, b-FGF, PDGFA-B, PDGFRα-β, PlGF1–2) on consecutive samples. Results: At present 6 patients have been enrolled. Patients had ovarian cancer (N=3), breast cancer (N=2) and squamous cervical cancer (N=1) as primary tumours. All patients had been extensively pre-treated with 4 to 8 lines of systemic therapy. Efficacy: Three (3/6) patients showed regression of effusion fluid by consecutive CT-scans. Two (2/6) patients had unchanged volume and one (1/6) patient had an increase of fluid volume. Follow-up: Two (2/6) patients stopped the trial early due to toxicity. Two patients (2/6) died during treatment due to disease progression elsewhere. Two patients (2/6) completed the 2 cycles of 6 weeks treatment. Of which, one had sustained reduction in volume fluid accumulation, while the other one had initial stabilisation after one cycle but eventually had increasing volume of ascites. Conclusions: These findings support the hypothesis that sunitinib could be useful in the management of ME. Although the numbers are low, in a majority of patients (5/6) there was reduction or stabilisation of fluid accumulation. Toxicity led to study withdrawal in a third of our patients. An update of patient numbers as well as results of translational/biomarker analysis will be presented at the meeting. No significant financial relationships to disclose.