bluuebunny/crossref_metadata_2025_split huggingface dataset:
Ultra-sensitive detection of biomarkers and applications in pharmaceutical discovery and development. Example application to rat cardiac troponin I assay via nanoparticle probes
Authors: Bao P. , Lubben T. , Holzman T. 
Published: August 2020

e14594 Although specific to heart, rat cardiac troponin I (cTnI) is an example of an important biomarker for assessing drug-induced cardiotoxicity in animal models used in various phases of drug discovery and development. Current commercially available assays can only detect 10 ∼ 100 pg/mL in serum at the lowest limits. To improve the sensitivity of rat cTnI assay, we have developed a generically applicable, microarray based nano-probe test. Our rat cTnI assay algorithm uses a multi-step robotic process, which relies on non- isotropically oriented antibodies on functionalized glass as multiplexed microarrays to capture cTnI from serum. Functionalized, 130 angstrom diameter gold nano-probes (measured by static light scattering, 5 nm S.D.) also bind to the troponin through a molecular-scale complex containing antibodies. The troponin-bound molecular complex is then quantified through silver enhancement of the functionalized gold. Assays in this format can be rapidly configured and implemented for a wide array of potential biomarkers. For cTnI we have demonstrated a robust and ultra-sensitive assay with an LOD of less than 500 femtograms of rat cTnI per mL serum, and an overall CV of less than 20%. The assay also shows very low background, a broad dynamic range and over 3 logs of linear dose response. As an example of the potential of high sensitivity, the nanoparticle-based rat cTnI assay could significantly increase the effectiveness of measuring drug-induced heart damage at very low drug dosages and early times. Such sensitive and early measurements can improve examination of the safety of drug candidates while correspondingly reducing drug development time and cost. [Table: see text]
Epidermal growth factor receptor intron 1 (CA)n dinucleotide repeat polymorphism and mutations associated with the responsiveness of molecular targeted therapy in lung cancer
Authors: Wang J. , Zhang S. , Lai R. 
Published: August 2020

e14595 Background: To explore EGFR gene intron 1 (CA)n repeat polymorphism and mutations associated with the responsiveness of molecular targeted therapy in lung cancer. Methods: Both observed groups consisted of 116 somatic specimens of lung cancer and controls consisted of 20 peripheral blood samples were analyzed by direct DNA sequencing of EGFR mutations at exons 18,19,21. Also (CA)n repeat polymorphisms in intron 1 of EGFR from 48 specimens were analyzed. 45 lung cancer patients were followed up. Results: EGFR mutations were found in 24 of 116 somatic specimens (20.69%). Response rate and disease control rate to EGFR TKIs was significantly higher in the patients with EGFR mutations (62.5% vs. 0)(P<0.0l),(100% vs.44.4%) (P<0.05)respectively than those without it in observed groups.In patients harboring EGFR mutations, disease control rate to patients treated with Iressa (100%) was significantly higher than those who never treated with it (40%)(P<0.05). The frequency distribution of EGFR intron 1 (CA)n repeat in 48 specimens was 23(47.9%)low (CA)n repeat(CA≤16)and 25 (52.1%) high repeat(CA>16). There was no significant difference in response rate and disease control rate of EGFR TKIs treatment between low and high (CA)n repeat numbers both in 16 patients with mutations and 18 patients without mutations in observed groups(P>0.05). Conclusions: Somatic mutations of EGFR is a major determinant of EGFR TKIs response in lung cancer. There was no significant difference in response rate and disease control rate of EGFR TKIs treatment between low (CA) n repeat patients and high repeat ones under the consideration of the mutant factor. No significant financial relationships to disclose.
Telomerase interference in combination with docetaxel therapy for maximal prostate cancer inhibition: In vitro data for a novel therapeutic approach
Authors: Sahu D. , Xu T. , Lau R. , Xue L. , Goldkorn A. 
Published: August 2020

e14596 Background: Metastatic castration-resistant prostate cancer (mCRPC) carries a median survival of 18 months with standard docetaxel based therapies. Telomerase interference (TI) is a promising novel therapeutic strategy that exploits the high telomerase activity in cancer cells by introducing a mutated telomerase RNA (MT-Ter) that encodes toxic telomeres and rapidly induces apoptosis. We investigated whether TI can be combined with docetaxel therapy to achieve greater growth inhibition in mCRPC. Methods: PC3 and DU145 mCRPC cell lines were treated with docetaxel in the presence of TI or vector control. TI was accomplished by concurrent lentiviral expression of 2 constructs: telomerase RNA with an altered template region (MT-Ter) and siRNA targeting wild-type telomerase RNA (anti-Ter siRNA). Telomere length and telomerase activity were assessed using RT-PCR and TRAP, respectively. Proliferation, apoptosis, and DNA damage were quantified using MTS, TUNEL, and 53bp1 staining, respectively. Statistical significance was calculated using a 2-sided t-test. Results: Docetaxel (10nM) induced 22% inhibition (p=0.01) of PC3 proliferation in the presence of vector control and 41% inhibition (p=0.001) in the presence of TI (results were similar in DU145 cells). This near-doubling of efficacy was attributable to an independent inhibitory effect (17% inhibition, p=0.04) from TI treatment alone, which occurred without change in bulk telomere length or telomerase activity. TI alone generated increased numbers of DNA damage foci (7/cell vs. 2/cell with vector control) while docetaxel alone did not generate significant increases in DNA damage. Both TI and docetaxel induced a marked increase in the rate of apoptosis. Conclusions: Docetaxel and TI each exerted a pro-apoptotic effect which, when combined, produced an additive inhibition of mCRPC proliferation. TI-mediated apoptosis ensued from DNA damage, consistent with its known telomeric-uncapping effect, while docetaxel-induced apoptosis was not associated with direct DNA damage, also consistent with known docetaxel mechanisms of action. These findings underscore the therapeutic promise of combining standard agents with TI to improve efficacy and reduce toxicity. No significant financial relationships to disclose.
Pilot study of fixed-dose bevacizumab (200 mg) for solid malignancies
Authors: Reddy S. K. , Curti M. , Janis M. , Minow R. 
Published: August 2020

e14600 Background: We report our initial experience with fixed dose bevacizumab at 200mg (approximately 3mg/kg). Phase I studies suggested that an optimal dose for phase II studies with bevacizumab is 3mg/kg and that circulating VEGF was undetectable at 0.3mg/kg. (Gordon et al. JCO 2001) We proposed a fixed-dose regimen of bevacizumab which we hypothesized would yield equivalent response rates with reduced toxicities and cost versus higher-dose regimens. Patients with advanced malignancies for whom bevacizumab would be indicated were analyzed. Methods: 15 patients were treated with 200mg bevacizumab in combination with antineoplastic therapy. 6 patients had NSCLCa, 8 patients had Colorectal cancer, and 1 patient had BRCA. Results: 15 patients are evaulable for response and have completed a total of 234 doses of bevacizumab (median number of doses =13) with no grade III/IV toxicity, or bevacizumab associated toxicities seen. No grade III or greater hypertension was observed. Proteinuria was not formally assessed, but no grade 3 or greater proteinuria was reported. All patients are evaluable for response with overall response rate of 33% (5/15). With a median follow-up (from the start of bevacizumab) of 452 days (222–1,699 days), median survival has not been reached with only 2 deaths. Conclusions: Fixed dose bevacizumab appears to be effective, less toxic, significantly less expensive and supported by biologic rationale and prior phase I studies and warrants further investigation. Additional patients will be accrued in a prospective phase II trial. [Table: see text]
Activity of mTOR-Inhibitor Rad001 (everolimus) in differentiated and anaplastic thyroid cancer cell lines
Authors: Behlendorf T. , Voigt W. , Mueller T. , Jordan K. , Arnold D. , Schmoll H. 
Published: August 2020

e14608 Background: Recent data suggest that aberrant activation of PIK3/AKT-pathway and mTOR are involved in the development of thyroid cancer, particularly of anaplastic (ATC) and follicular (FTC) subtype. Therefore, mTOR could be a potential treatment target in thyroid cancer. Methods: To asses the potential role of mTOR as target for the treatment of thyroid cancer, two human ATC cell lines SW1736 and 8505C, the papillary thyroid cancer (PTC) cell line BCPAP and the FTC cell line FTC133 were exposed for 96h to increasing concentrations of the mTOR inhibitor RAD001 (Everolimus, kindly provided by Novartis, Switzerland). For combination experiments 10 nM of RAD001 were combined with increasing concentrations of either doxorubicin (DOX) or cisplatin (CDDP) continuously. Cytotoxicity was measured using the sulforhodamine B assay. IC50-values were calculated with Sigma Plot (Jandel Scientific) and drug interaction was determined by the model of Drewinko. Results: The observed IC50-values of RAD001 were 1nM (FTC133), 10 nM (BCPAP), 1000 nM (SW1736) and 9400 nM (8505C). In contrast to the pronounced differences in sensitivity as assessed on the basis of IC50, a growth inhibitory effect ≥ 25 % was seen in all cell lines at a concentration of 1 nM of RAD001. IC50 for CDDP ranged from 1,3–4,8 μM and for DOX from 8–40 nM. Combination of 10 nM RAD001 with either DOX or CDDP resulted in additive drug interaction with the exception in cell line 8505C where significant synergy was found for the combination with CDDP. Conclusion: RAD001 exerted interesting preclinical activity in two differentiated thyroid cancer cell lines. Mainly additive drug interaction in thyroid cancer cell lines was observed for combinations with CDDP and DOX. Mechanistic investigations are underway and will be presented at the meeting. At least for differentiated thyroid cancer mTOR-inhibition appeared promising, further evaluation in thyroid cancer seems warranted. No significant financial relationships to disclose.
Dual inhibition of αV integrins and Src kinase activity in colon cancer cells
Authors: Starodub A. , Jia J. , Cushman S. , Marshall D. , Hurwitz H. I. , Nixon A. B. 
Published: August 2020

e14609 Background: Integrins are commonly upregulated in tumor cells and are important regulators of invasion and metastasis. Integrin signaling is initiated upon engagement of ECM and requires Src as well as various other proteins including ILK, FAK, and paxillin. Methods: Presence of αv integrins on CRC cell lines was established by flow cytometry. CNTO 95 (10μg/ml, a monoclonal anti-αv integrin antibody; Ortho Biotech), and dasatinib (<200nM; BMS, src small molecule inhibitor) were used to study the effects of combined integrin and src inhibition on proliferation and serum-induced migration of colon cancer cell lines in vitro. Downstream signaling changes were assessed by immunoblotting for phosphorylated forms of src, FAK (Y397, Y576/577 and Y925), paxillin, GSK3β and AKT (S473) in HT29, HCT116 and RKO. Results: Proliferation was inhibited by dasatinib in HT29, HCT116, DLD1 and HCT15 in a dose-dependent manner, while RKO and SW48 were resistant. Combining CNTO 95 with dasatinib further inhibited proliferation in all dasatinib-sensitive cells, yet resistant cells were unaffected. Migration was blocked by both CNTO 95 and dasatinib in all cell lines, and combining the two drugs produced augmented effect. Src activation and src- dependent FAK phosphorylation at sites 576 and 925 were blocked by dasatinib; CNTO 95 had little effect alone, but potentiated the effect of dasatinib. Paxillin phosphorylation was modestly blocked by both compounds, but the combination produced significantly augmented inhibition in the three cell lines tested. The phosphorylation status of AKT and GSK-3β, which are downstream of ILK, was inhibited by both drugs as single agents. The combination of dasatinib and CNTO 95 produced further inhibition. Conclusions: Dual inhibition of Src by dasatinib and αv integrins by CNTO 95 produced additive to synergistic inhibition of proliferation in dasatinib sensitive CRC cell lines, and inhibition of migration in all cell lines tested. Decreased phospho-paxillin levels may be responsible for the pronounced inhibition of migration observed after dual treatment with dasatinib and CNTO 95 in these CRC cell lines. These data support the rationale for combined Src/integrin inhibition in colon cancer, and further suggest approaches to patient selection strategies. [Table: see text]
APX3330 inhibition of the redox function of ape-1/ref-1 (Ref-1) in promyelocytic leukemia cells enhances retinoic acid (ATRA) induced myeloid differentiation and limits cell proliferation as an approach to the prevention of the retinoic acid syndrome (RAS)
Authors: Robertson K. A. , Colvin E. S. , Kelley M. R. , Fishel M. L. 
Published: August 2020

e14613 Background: ATRA + chemotherapy has improved the treatment of promyelocytic leukemia(APL). However, 25% of ATRA treated APL patients experience toxicities that comprise the RAS (life-threatening respiratory distress, edema, renal failure, hypotension, coagulopathy and rising blast count). One approach to prevent RAS is to limit blast proliferation and enhance myeloid differentiation. Ref-1 is a DNA repair protein that functions in redox regulation of cellular proteins, such as Fos, Jun, p53, and NFkB. HL60 myeloid leukemia cells are promyeloblasts that respond to ATRA with granulocytic differentiation/growth arrest. Prior studies suggest Ref-1 redox control is integral to ATRA-induced differentiation. To define the role of the redox function of Ref-1, we used the Ref-1 specific drug, APX3330, to block Ref-1 redox function and examined the response of HL60 cells to ATRA. Methods: Cell growth assessed using trypan blue. Differentiation was evaluated by morphology and expression of CD11b by flow cytometry. Apoptosis was assayed by annexin-PI staining on flow cytometry and cell cycle analysis assayed with propidium iodide flow cytometry. To assess activation of the MAPK pathway, BLR-1 expression was determined by real time PCR. Results: 1) APX3330 blockade of Ref-1 redox function resulted in limited cell growth yet a profound increase in differentiation and a moderate increase in apoptosis. 2) dose dependent studies with ATRA showed a similar degree of differentiation in cells treated with 10 μM ATRA to cells treated with APX3330 + 0.01 μM ATRA; allowing HL60 cells + APX3330 to give a similar response to a 1000 fold lower dose of ATRA. APX3330 alone did not induce differentiation and induced only minimal apoptosis but in combination with ATRA, increased the number of cells in G1/G0 phase significantly. 3) APX3330 + ATRA increased BLR-1 expression significantly by real time PCR suggesting enhanced activation of the MAPK pathway. Conclusions: APX3330 + ATRA limits HL60 growth and dramatically enhances terminal granulocytic differentiation. These finding may provide a therapeutic approach for prevention of the RAS. No significant financial relationships to disclose.
Preclinical development of orally available sphingosine kinase inhibitors
Authors: Smith C. D. , Maines L. W. , Zhuang Y. , Green C. L. , Keller S. N. , Beljanski V. , Knaak C. , Wang W. 
Published: August 2020

e14615 Background: Sphingolipid-metabolizing enzymes control the dynamic balance of important bioactive lipids, including apoptotic ceramide and proliferative sphingosine 1-phosphate (S1P). Several growth factors and inflammatory cytokines promote the cleavage of sphingomyelin and ceramide leading to rapid elevation of S1P levels through the action of sphingosine kinases (SKs). SK1 is oncogenic and is markedly overexpressed in a variety of human cancers, making this enzyme a potential molecular target for cancer therapy. Methods: SK inhibitors were synthesized and evaluated in a variety of cellular and in vivo models. Antitumor activity was assessed in an allogeneic model utilizing murine JC mammary adenocarcinoma cells growing in Balb/c mice, and a xenograft model of human Bxpc pancreatic adenocarcinoma cells growing in scid-mice. Results: The lead SK inhibitors, called ABC294640 and ABC294735, inhibit SK1 and SK2 at low micromolar concentrations, and are nontoxic to rodents at acute doses up to at least 1000 mg/kg (po). ABC294640 is orally-available, and has excellent pharmacokinetics, with serum levels exceeding the effective SK inhibitory concentration for at least 8 hours. Acute- and chronic- toxicology studies indicate that ABC294640 induces a transient minor decrease in the hemotocrit of rats and mice receiving 100 and 250 mg/kg/day; however, this normalizes by 28 days of treatment. No other changes in hematology parameters, or gross or microscopic tissue pathology result from treatment with ABC294640. Oral administration of ABC294640 to mice bearing JC adenocarcinomas results in dose-dependent antitumor activity associated with depletion of S1P levels in the tumors and progressive tumor cell apoptosis. ABC294735 is also orally-active in both JC and Bxpc tumor models, and demonstrates synergistic antitumor activity when combined with cisplatin or gemcitabine. Conclusions: These newly developed SK inhibitors provide orally-available drug candidates for the treatment of pancreatic cancer and other diseases. IND-enabling CMC and toxicology work is currently underway, and an SK inhibitor is expected to enter clinical trials in early 2010. [Table: see text]
Evaluation of anticancer drugs in NSCLC
Authors: Guirgis H. M. 
Published: August 2020

e14616 Background: Methodology to evaluate anti-cancer drugs in GI malgnancies was previoulsy reported. It has been estimated that the society would be willing to pay $50,000-$100,000 per life-year S gain (S Negrier et, 2007). Objectives: Design scoring model methodology to evaluate and grade anti-cancer drugs in NSCLC with emphasis on the association between between S, AEs, A and C. Methodology: 0S and AEs data were abstracted from J Douillard, R Herbert, Le Chevalier, C Mangeold, A Sandler, F Shepard, R Rossell and C Thienelt (2005–2008). A was graded in points (P) from 3 oral to zero daily injections. AEs: 4 (placebo) to 0 (fatalities) using CTCAE v 3.0. S gain (days,d) was calculated in a corrected ratio of S/360 (SR); Cost R: as C/d/s/$1500, divided by (A+ AEs), corrected and expressed as C scores. C in US $ was based on treating one patient for entire course,70 kg or 1.7/ m2. Using a grading scale from A to D, Survival > 360d, SR>10; C/d/s up to $63 and CR up to 4.2 were rated as (A+) while S< 15d, SR <0.42, C/d/s> $1500 and CR>100 as (D). Results: Examples shown in Table . Adj Cis + Vin after resection in stage II and III scored the highest SR and lowest CR with A+ rating. However, there was a lesser SR using the same combination in mNSCLC. OS markedly improved with Er and BV in 2nd-line. Addition of Cetuximab (Cet) to Cis and Vin demonstrated modest S gain at a much higher CR. Bevacizumab (BV) combined with Pac + Car improved OS over Pac + Car in 1st and 2nd lines at higher CR. Combination of BV, Cis and Gemcitabine demonstrated the lowest S, highest CR and D rating. Discussion: S and C were calculated as Ratios. C scores were based on the association of S with arbitrary but relative weights assigned to AEs and A. BV and Er improved OS in 2nd line, in contrast to phase III Beta study. Adjuvant Cis+Vin seems to be cost-effective. Increased CR was not necessarily associated with improved S. Grading of drugs varied depending on S or C. Higher C/d/s, increased AEs and cumbersome A resulted in higher C scores. Conclusions: SR, CR and possibly C scores might be advantageous in evaluation, grading and comparison of anti-cancer drugs. [Table: see text] No significant financial relationships to disclose.
MET in small cell lung carcinoma (SCLC): Effects of a MET inhibitor in SCLC cell lines and prognostic role of MET status in patients
Authors: Cañadas I. , Arumi M. , Lema L. , Martinez A. , Grande E. , Bellosillo B. , Rojo F. , Rovira A. , Albanell J. , Arriola E. 
Published: August 2020

e14617 Purpose: HGF/MET pathway is aberrantly activated by receptor overexpression and mutations in SCLC preclinical models, enhancing their oncogenicity. The significance of MET expression in SCLC remains unclear. Our aim was to analyze the effects of MET inhibition in chemosensitive/refractory SCLC models and to study the expression pattern and prognostic impact of total and phosphorylated (p) MET in SCLC patients. Methods: Total and p-MET expression (Western Blot), gene copy number (FISH), and exon 14 activating mutations (sequencing) were evaluated in H69 and H69AR SCLC cell lines. PHA-665752 (PHA), alone or combined with doxorubicin, was used to study the effects of pathway inhibition on viability, colony formation and invasion assays in basal/stimulated conditions (HGF). Fifty-eight SCLC cases were evaluated for MET and p-MET expression by immunohistochemistry. Survival analyses were performed. Results: H69 and H69AR (both R988C mutated) expressed MET at basal conditions, but not p-MET. HGF induced MET phosphorylation, increased proliferation (20%) and protected cells from doxorubicin cytotoxicity. PHA 0.5μM blocked MET phosphorylation, decreased colony formation by 50% in H69, counteracted the cytoprotective effect of HGF and inhibited invasion in H69AR. MET expression was found in 98% normal bronchial epithelia, and 78% tumor samples (overexpression 38%). Activated MET was focally detected in normal and metaplastic mucosa and expressed in 22% tumors. MET expression was associated with improved overall and disease free survival (p: 0.06 and 0.051, respectively). All p-MET positive cases within MET expressing tumors, showed relapsed disease (83% in negative p-MET samples, p=0.065), suggesting MET activation may revert the good prognosis linked to total MET expression. Conclusions: MET activation, results in a more aggressive phenotype in SCLC cells. PHA at MET inhibiting concentrations reverses this phenotype. In SCLC specimens, MET expression was more prevalent than p-MET and associated with raised prognosis. All these data suggest that studies with MET inhibitors should focus on p-MET positive SCLC. [Table: see text]