bluuebunny/crossref_metadata_2025 huggingface dataset:
Translational relevance of baseline NLR and LMR ratio in predicting response of EGFR-TKI.
Authors: Varayathu Hrishi , Tiwari Rashi Yogendranath , Somayaji Shalaka , Mathew Smruthy Ann , Thomas Beulah Elsa , Sangi Lalram , Thomas Renjan , Balaram Gautam , Ghosh Mithua , Thungappa Satheesh Chiradoni , Mufti Suhail Sayeed , Bj Sriniivas , Munirathnam Vinayak , Naik Radheshyam , Ajaikumar Basavalinga Sadasivaiah
Published: June 2021

e15020

Background: Neutrophil-Lymphocyte Ratio (NLR) and Lymphocyte-Monocyte Ratio (LMR) have been under scrutiny for their potential as a prognostic tool in various cancers. While there have been conflicting opinions on their reliability, NLR has shown rather near accurate predictions in previous studies. Targeted tyrosine kinase inhibitors have shown good results in improving OS and PFS of patients with EGFR-mutated NSCLC but the prognostic indicators are limited. This study aims to decipher if baseline NLR and LMR ratios are valid tools to predict prognosis of patients with EGFR-mutated NSCLC. Methods: We analyzed 78 advanced NSCLC patients harboring Exon 19 Deletion and Exon 21 L858R mutation undergoing EGFR-TKI therapy. Baseline NLR and LMR were measured in peripheral blood within two weeks prior to the initiation of TKIs. Overall survival was used as an outcome measure for TKI therapy and it is defined as from the initiation of TKIs to death by any cause. The cut-off for NLR and LMR was calculated using an X-tile plot (version 3.6.1, Yale University, New Haven, CT, USA) by dividing marker data into three populations: low, middle, and high (i.e., two divisions), with randomized 1:1 “training” and “validation” cohorts. Results: Overall median survival of the study group was 31.5 months and median age was found to be 60 years. Gefitinib (35.9%) and afatinib (34.6%) were the most commonly prescribed followed by erlotinib (21.8%) and osimertinib (7.7%). With cut-points of 1.8 and 4.9 we analyzed three populations for baseline NLR as ≤1.8, 1.9-4.9 and > 4.9. Similarly, for LMR cut-points of 3.49 and 5 were found and we analyzed patients with LMR of ≤3.49, 3.5-5 and >5. There was a statistically significant overall median survival observed as per the obtained NLR as well as LMR cut-off as given in the table. Conclusions: A statistically significant co-relation was observed with baseline NLR and LMR ratio in predicting response to EGFR TKIs in NSCLC. Our study further strengthens the concept of using NLR and LMR as peripheral blood biomarkers for prognostic assessment in cancer patients undergoing treatment.[Table: see text]
Consistent performance of highly multiplexed RNA fusion detection with low RNA input and limit of detection.
Authors: Debruyne David , Li Chenyu , Spencer Julia , Kapoor Vidushi , Liu Lily Y. , Lee Lucie , Feigelman Rounak , Burdon Grayson , Liu Jeffrey , Liu Guoying , Liu Zhitong
Published: June 2021

e15029

Background: Gene fusions are usually caused by chromosomal rearrangements and frequently associated with various cancers. Detection of fusion mutations is an important part of cancer management, and are usually detected in RNA purified from biopsies of fresh tissue or FFPE sections. However, the amount of RNA from these samples is usually limited. We developed a new, PCR-based fusion detection method, which allows detection of unknown and novel fusion mutations. We demonstrate its consistent performance across various RNA inputs, and sequencing depths, in terms of technical performance and limit of detection. Methods: A multiplex PCR RNA fusion panel was designed to cover known RNA fusion targets and unknown fusion targets for target enrichment via single primer multiplex PCR. SeraSeq fusion Reference RNA was used to evaluate the limit of detection of the method. The amplicons were about 110 bp on average to allow for target amplification from challenging fragmented samples such as FFPE RNA. After reverse transcription, the panel was used in a multiplex PCR to amplify the targets, and a final PCR was used to add sequencing adapters and sample indexes. The resulting libraries were sequenced on Illumina sequencers. The data was analyzed for the detection of known fusion variants across varying initial inputs of targets, as well as the performance in terms of mapping and on-target rates, as well as the presence of human ribosomal RNA. Results: We found that the fragmented RNA length had little effect on the performance of the libraries. The required numbers of PCR cycles were optimized based on testing with various amounts of starting RNA input material. We observe a linear relationship between the library yields and RNA input amounts from 10 to 50ng, which had R1 mapping rates above 95% and 98%, respectively. We report the detection of all twelve fusion variants, which are targeted by our panel and present in the Seraseq Fusion RNA Mix v4 reference standard, and which are covered with an average sequencing depth of 0.2M reads, or 3600 reads/amplicon, across all assays. Even at inputs as low as 3 ng, all 12 fusions were typically detected. In general, the results for individual fusions among the different replicates were concordant, with limited observed variance in reads across some fusion junctions between assays and replicates. Over 90% of the reads supported the fusion calls from all inputs (ranging from 1.6 to 50 ng). Over 97% of the reads supported the fusion calls between 12 and 50 ng. Conclusions: A multi-lab validation confirmed the above results, and were also comparable when using an additional distinct CleanPlex fusion panel. To conclude, our CleanPlex fusion technology allows for reliable and simultaneous detection of 12 designed clinically relevant RNA fusions even at low input amounts.
Analysis of the relative copy numbers of TP63, YAP1 and KMT2D genetic loci and EGFR expression in orthotopic patient-derived xenografts of cardioesophageal cancer in immunodeficient mice.
Authors: Goncharova Anna S. , Kolesnikov Evgeniy N. , Zaikina Ekaterina V. , Volkova A. V. , Mindar M. V. , Khodakova D. V. , Lukbanova Ekaterina A. , Kurbanova L. Z. , Kozhushko Mikhail A. , Anisimov Aleksandr E. , Myagkov Roman E. , Sanamyants Sergey V. , Katsieva Tanzila B.
Published: June 2021

e15033

Background: Cardioesophageal cancer is a common tumor affecting the cardiac stomach and lower esophagus. Tumor heterogeneity is important, since it can cause the ineffectiveness of chemotherapy and radiotherapy due to individual characteristics of the neoplasm in different patients, as well as the intensification of invasion and metastasis. Our purpose was to analyze the relative copy numbers of TP63, YAP1 and KMT2D genetic loci and EGFR expression in orthotopic patient-derived xenografts of cardioesophageal cancer. Methods: The model of cardioesophageal cancer was created in Balb/c Nude mice with surgical bioptates of adenocarcinoma obtained from donors. The relative copy numbers of TP63, YAP1 and KMT2D genetic loci and EGFR expression were determined by Real-Time qPCR. Results: A PDX model of cardioesophageal cancer was successfully created by transplanting a moderately differentiated adenocarcinoma from a patient diagnosed with infiltrative ulcerative cancer of the lower third of the esophagus with a transition to the cardiac stomach into the distal esophagus of Balb/c Nude mice. The EGFR expression was elevated in patient tumor samples, compared to healthy tissues (p = 0.021) and gastric cancer xenografts (p = 0.07). Molecular genetic analysis demonstrated an association between an increased EGFR expression and changes in the relative copy numbers of TP63, YAP1, and KMT2D in donor samples compared to gastric cancer xenografts (p = 0.02, p = 0.026 and p = 0.042). Conclusions: The relative copy numbers of TP63, YAP1, and KMT2D were associated with intensified EGFR expression and changed with each new generation of orthotopic xenografts.
Development of a DNA damage response (DDR) therapeutics platform for oncology.
Authors: Das Debanu , Duncton Matthew , Pellicena Patricia , Deacon Ashley , Wilson David , Georgiadis Millie
Published: June 2021

e15036

Background: Cancer cells respond to increases in DNA damage by deploying their DNA damage response (DDR) pathways. We are building a platform for the discovery and development of target-specific DDR therapeutics, including small molecule inhibitors and targeted protein degradation warheads, founded on fragment- and structure-based drug discovery. Methods: Our DDR platform, which includes hit-to-lead, lead optimization and candidate selection, starts with hit generation from a new technology that uses high-throughput protein X-ray crystallography to directly screen compound libraries. Our hit generation process produces empirical evidence of direct target engagement. The elucidation of high-quality ligand-bound 3D structures reveals the location and pose of the ligand and details of the protein-ligand interactions. Thus we can predict the structure-function consequences of the hit molecule engagement, which sets the stage for rapid assessment of synthetic tractability and intellectual property. After hit identification, we apply a multi-pronged approach in hit-to-lead conversion and lead optimization using iterative biophysical and biochemical assays, coupled with crystallography. We are applying our approach to several new targets in DDR and will present some early progress in this space. Results: Apurinic/apyrimidinic endonuclease 1 (APE1) is the major repair enzyme for abasic sites in DNA and contributes to DNA strand break processing. Many studies have associated increased APE1 levels with enhanced growth, migration, and drug resistance in human tumor cells, and with decreased patient survival. APE1 has been implicated in over 20 human cancers, including glioblastoma, making the protein an attractive target for the development of anticancer therapeutics. Despite intensive effort, there are no clinical endonuclease inhibitors of APE1. We have identified 25 diverse small molecule fragments that bind to APE1 at two distinct sites, including the endonuclease site. Pol eta (or PolH) is a DNA polymerase implicated, among other things, in the development of cisplatin resistance in a subset of ovarian cancers. In our quest to develop PolH inhibitors, we have identified 5 diverse fragments that bind to two distinct sites in the polymerase including a new potential allosteric site. Our results on APE1 and PolH represent the first known cases of crystal structures of small molecules bound to these proteins. Flap endonuclease (FEN1) is implicated in several cancers including for example ER/tamoxifen-resistant breast cancer. We are developing a targeted protein degradation approach using PROTACs (Proteolysis-Targeting Chimeras) toward the development of novel therapeutics against FEN1. Conclusions: Our results will help us develop small molecule inhibitors and targeted protein degradation against DDR targets that may be effective as single therapies or be used to make existing therapies more effective.
Towards the clinical validity of tumor organoid drug screens: Establishing a framework for organoid disease models.
Authors: Slostad Jessica , Masood Ashiq , Swoboda April T. , Levy Mia Alyce
Published: June 2021

e15037

Background: Despite advances in biomarker-directed cancer therapies to predict tumor response to treatment, precision oncology remains an imprecise science. Tumor organoid drug screens present an opportunity to test multiple potentially effective therapies simultaneously before exposing a patient to treatment toxicities. Preliminary studies have established the analytic validity; however, the clinical validity of the tumor organoid response to a drug to predict the tumor response in patients is unclear. Methods: In order to establish the clinical validity of an organoid drug screen, a clinical disease model should have the following features to enable comparison of the tumor’s clinical response to treatment and the organoid’s response to the same treatment. First, a fresh tissue biopsy (non-bone) needs to be obtained for organoid development prior to the start of systemic treatment. Priority should be given to tumors known to successfully grow organoids. Disease models where the standard of care systemic treatment is a single cytotoxic or targeted agent would best assess correlation between the drug screen and patient response, such as metastatic breast, cervical, or prostate cancer. This would often lead to enrolling patients on later line systemic therapies. Studies should avoid drugs whose mechanism of action leverages the patient metabolism or tumor microenvironment (e.g. immunotherapy, aromatase inhibitors, VEGF inhibitors). Patient should have measurable disease that can be measured clinically, radiographically, or pathologically and using standardized response evaluation criteria (e.g. RECIST). Next generation sequencing would assess genomic concordance between the tumor in the patient and organoid. These studies would determine the feasibility and timeliness of prospectively developing tumor organoids that is sufficient to perform a drug screen. Results: We propose two research models to evaluate the clinical validity of tumor organoid drug screens: a metastatic solid tumor and neoadjuvant solid tumor disease model. Metastatic disease models provide an opportunity to assess response across multiple cancer types at the time of progression and initiation of next line of therapy. The efficacy of chemotherapy can be determined using objective data from radiologic response (RECIST). In neoadjuvant models, pathologic (pCR) and radiologic response can provide objective data for organoid response. These study design features will lay the framework for determining the clinical utility of organoid drug screens. Conclusions: We call for clinical studies assessing the clinical validity of tumor organoid drug screens and determining their concordance with patient response to systemic therapy. Advancements in clinically validated tumor organoids have the potential to fundamentally shift clinical paradigms and improve patient outcomes in cancer treatment.
Cytoprotective effects of spirulina extract against doxorubicin‐induced cardiotoxicity: Biochemical evidences and translational perspectives in cardio-oncology.
Authors: Maurea Nicola , Quagliariello Vincenzo , Paccone Andrea , De Laurentiis Michelino , Buccolo Simona , Iovine Martina , Botti Gerardo
Published: June 2021

e15042

Background: Spirulina, a blue-green algae used in the daily diet of natives of Africa and America, has antioxidant, anticancer, antiviral, hepatoprotective, immune enhancing and lipid-lowering effects. Considering that doxorubicin-induced cardiotoxicity involves myocardial inflammation, ferroptosis and overexpression of several cytokines involved in cell dead and apoptosis, the use of spirulina during exposure to doxorubicin could be an intriguing method to reduces myocardial injuries in cancer patients undergoing treatment with anthracyclines. Methods: Human fetal cardiomyocytes (HFC cell line) were exposed to doxorubicin (from 10 to 500 nM), alone or in combination with spirulina extract (from 10 to 100 µg/ml) for 24 and 48h. After the incubation period, we performed the following tests: determination of cell viability, through analysis of mitochondrial dehydrogenase activity, study of lipid peroxidation (quantifying cellular Malondialdehyde and 4-hydroxynonenal), intracellular Ca2+ homeostasis. Moreover, pro-inflammatory studied were also performed (activation of NLRP3 inflammasome; expression of TLR4/MyD88; mTORC1 Fox01/3a; transcriptional activation of p65/NF-κB and secretion of cytokines involved in cardiotoxicity (Interleukins 1β, 8, 6). Results: Spirulina extract, co-incubated with doxorubicin exerts cardioprotective effects, enhancing cell viability of 20-36.7 % compared to untreated cells (p<0,01 for all); spirulina extract reduced significantly the cardiotoxicity through MyD88/NF-KB/cytokines axis and mTORC1 Fox01/3α mediated mechanisms and reduces oxidative damages of doxorubicin resulting in lower intracellular calcium content. Conclusions: For the first time, it was demonstrated that spirulina extract exert anti-inflammatory and cytoprotective properties in myocardial cells exposed to growing concentration of doxorubicin leading to preclinical studies in mice analyzing any effects on cardiac strain and left ventricular ejection function.
New tropolone derivative to inhibit cell motility in colorectal cancer cells.
Authors: Mezhevova Irina V. , Filippova Svetlana Yu. , Timofeeva Sofia V. , Sitkovskaya Anastasia O. , Shamova Tatiana V. , Lukbanova Ekaterina A. , Goncharova Anna S. , Gusakov Evgeniy A. , Sayapin Yuriy A. , Kolesnikov Vladimir E. , Kolesnikov Valeriy V. , Sidorenko Maria , Panchenko Pavel , Kit Oleg I.
Published: June 2021

e15043

Background: Tropolones are small, naturally occurring organic compounds that have anti-cancer effects in a variety of cancer cell lines. Among proposed mechanisms of tropolone derivatives anti-cancer activity are caspase activation, matrix metalloproteinases and histone deacetylases inhibition and other actions. Since matrix metalloproteinases are the major proteins that contribute to tumor cell tissue invasion and migration their blockers are extensively studied as prospective anti metastatic agents. We assumed that tropolones may also have potential as metastases prevention drugs. The aim of the study was to investigate the influence of new tropolone derivative 2-(6,8-dimethyl-5-nitro-4-chloroquinolin-2-yl)-5,6,7-trichloro-1,3-tropolone on the cell migration capacity in colorectal cancer cells. Methods: Cell migration in permanent colorectal cancer cell lines HT29, Caco2 and HCT116 was studied using the standard wound healing assay with automated image acquisition and analysis on Lionheart FX (BioTek) cell imager. Cell cultures were maintained in DMEM medium with L-glutamine (1 μM) (Gibco) and 10% FBS (Gibco) at 37C0 and 5.0% CO2. The day before assay establishment cells were seeded at 250 000 cells per well on 24-well plates. After attachment of cells, a vertical wound in cell layer was made manually in each well, and then medium with studied tropolone derivative (5 μM) was added. Plates with cells were continuously incubated and photographed in cell imager at 37C0 and 5.0% CO2. The extent of cells migration was measured as the percent of wound area decrease after 48 hours of incubation in relation to starting time point. Data are given as: Mean ± 95% confidence interval. Results: The new tropolone derivative under study exhibited pronounced anti migratory activity in all studied colorectal cancer cell cultures. The most prominent effect was observed in HCT116 cell line: after 48 hours of incubation relative wound area decrease was 48.2%±5.7% in control and 7.2%±3.15% in experimental groups. In Caco2 cell line cell migration inhibition was the least with 65%± 6.9% relative wound area decrease in control and 51.5%± 7.7% in experimental groups. The moderate effect was observed in HT29 cell line (measured parameter was 63.6%±7.6% and 36.9%±11.6% in control and experimental groups accordingly). The obtained difference between control and experimental groups was significant in all tested cell cultures at 0.05 level (df = 30). Conclusions: Results of the present study prove tropolone derivative to be prospective candidates for anti-metastatic drug development. Further investigations are needed to reveal the exact mechanisms of tropolones anti-migratory activity.
Antimigratory effect of berberine in T98G, U87MG and primary glioma cell culture.
Authors: Mezhevova Irina V. , Filippova Svetlana Yu. , Timofeeva Sofia V. , Sitkovskaya Anastasia O. , Shamova Tatiana V. , Enin Yaroslav S. , Burov Oleg N. , Pozdnyakova Viktoria V. , Khokhlova Olga , Pushkareva Tatiana F. , Mezentsev Stanislav S. , Novikova Inna A. , Kit Oleg I.
Published: June 2021

e15045

Background: Berberine is an alkaloid compound with a structure that is highly similar to that of intercalating agents. It affects numerous cell signaling pathways and is widely studied as potential anticancer drug. It is known that berberine affects cancer cells migration through metalloproteinase-2 inhibition, but this effect was never studied on glioma cells. Anti-migratory drugs are of special interest in brain cancer therapy since glioma's highly invasive nature makes total surgical removal of tumor practically impossible. The aim of the study was to evaluate berberine anti-migratory activity on glioma cells. Methods: Cell migration capacity of T98G and U87MG cell lines, as well as primary glioma cell culture established in our laboratory, was assessed via standard wound healing assay with automated image acquisition and analysis on Lionheart FX (BioTek) cell imager. Prior to assay setting up cell cultures were maintained in DMEM medium with L-glutamine (1 μM) (Gibco) and 10% FBS (Gibco) at 37C0 and 5.0% CO2. Cells were seeded at 250 000 cells per well on 24-well plates and incubated overnight in order to attach to plate bottom. After that a vertical wound was made manually in each well, and berberine was added to experimental wells to final concentration 50 mg/L. Plates with cells were continuously incubated and photographed in cell imager at 37C0 and 5.0% CO2. The extent of cells migration was measured as the percent of wound area decrease after 24 hours of incubation in relation to starting time point. Data are given as: Mean ± 95% confidence interval. Results: In our study we berberine exhibited anti-migratory activity in all cell cultures under study. In rather fast growing primary cell culture wound area decrease was 99.23%±0.62% in control sample and 91.75%±0.28% in experimental sample. The difference was small but significant at p < 0.001 level (df = 30). Popular permanent glioma cell lines T98G and U87MG showed more prominent decrease in studied parameter with higher degree of variance at the same time. In T98G wound area decrease was 71.6%±12.3% in control and 48.8%± 7.6% in experimental samples after 24 hours of cultivation in presence of 50 mg/L berberine. While U87MG demonstrated 60.28%±5.13% and 37.5%± 8.34% wound area decrease accordingly. The obtained difference between control and experimental groups in permanent cell cultures was statistically significant at the 0.05 level (df = 30). Conclusions: Our preliminary research proved berberine to be potent anti-migratory agent in glioma treatment. Further investigations are needed to evaluate its ability to inhibit glioma cell expansion in vivo.
The functional significance of ATM phosphorylation of Bub3 on Serine 135 in sensitivity to DNA damaging agents.
Authors: Zhang Siyue , Xiao Mingming , Liu Zhuang , Mo Yaqi , Liu Hong , Xu Bo
Published: June 2021

e15048

Background: Identification of biomarkers to assess and modify the sensitivity of cancer cells to radiotherapy and chemotherapy is critical to improve cancer treatment outcome. Budding uninhibited by benzimidazoles 3 (Bub3) is a mitotic checkpoint protein, and it is frequently overexpressed in many cancers and associated with low survival rates. Bub3 is involved in the repair of DNA damage induced by radiotherapy and chemotherapy. We recently identified that the ATM kinase phosphorylated Bub3 on Serine 135 (S135) via the stable isotope labeled amino acid in cell culture -mass spectrometry analysis. This study aims to explore the mechanism of ATM-phosphorylated Bub3 in the DNA damage response (DDR) and its effect on tumor sensitivity to DNA damaging agents. Methods: The radiosensitivity of the cells was detected by clonal formation assay, the proliferation ability of the cells was detected by the MTS assay. The expression of DDR protein γ-H2AX in the nucleus was detected by immunofluorescence assay. Genomic instability was observed by multinuclear formation. Co-immunoprecipitation and Western Blot were used to explore the internal mechanism of ATM phosphorylation of Bub3 in DDR. Results: We showed that ionizing radiation (IR) could induce Bub3 S/TQ (the specific ATM consensus motif) phosphorylation in an ATM dependent manner. Mutation of Bub3 Serine 135 to alanine (S135A) led to a phosphorylation defect. Phenotypic experiments showed hypersensitivity to IR in cells expressing Bub3 S135A. Bub3 S135A prolonged existence of γ-H2AX foci and increased the proportion of cells containing micronuclei. Further, we found that Ku70 and Ku80 showed a significant increase after IR in their interactions with Bub3, while Bub3 S135A mutation significantly reduced the interaction, leading to impaired DNA repair. Conclusions: We demonstrate that Bub3 S135 phosphorylation mediated by ATM is essential for an optimal DDR and disruption of this pathway increases tumor sensitivity to DNA damaging agents.
Bispecific anti-PD-1/PD-L1 antibody CTX-8371 to promote cellular clustering and PD-1 shedding, antitumor activity and tolerability.
Authors: Albu Diana I. , Qin Yan , Wang Xianzhe , Li Vivian , Kim Taeg , Gong Bing , Schuetz Thomas , Kuklin Nelly
Published: June 2021

e15056

Background: Checkpoint blockade therapies targeting PD-1 and PD-L1 have shown great success for the treatment of various malignancies. However, a substantial fraction of patients with PD-L1-positive tumors remain unresponsive to these therapies. Novel therapy with significantly greater activity than the leading PD-1/PD-L1 inhibitors is expected to bring additional clinical benefit to patients. Here we describe the preclinical evaluation of CTX-8371, which combines anti-PD-1 and anti-PD-L1 monoclonal antibodies in one bispecific tetravalent molecule. Methods: The immune-enhancing activity of CTX-8371 was tested in vitro in T cell activation assays and tumor cell killing assay. CTX-8371 anti-tumor efficacy in vivo was assessed using mouse tumor cells expressing human PD-L1 implanted in transgenic mice humanized at the PD-1 and PD-L1 loci. CTX-8371 anti-tumor activity was also tested in xenograft tumor models. The mechanism of action of CTX-8371 was investigated in vitro using Jurkat cells expressing PD-1 or PD-L1, human PBMCs, and in vivo in tumor-bearing, chimeric PD-1/PD-L1 transgenic mice. CTX-8371 PK was determined in mice using an MSD ELISA-based assay and in cynomolgus monkeys using a qualified ELISA method. Dose range finding and toxicokinetic studies were performed in cynomolgus monkeys. Results: CTX-8371 potently enhanced T cell activation and function in vitro and showed curative efficacy as monotherapy in multiple solid tumor models, isografts or xenografts. Furthermore, CTX-8371 demonstrated superior anti-tumor efficacy compared to Keytruda or atezolizumab in checkpoint inhibitors-sensitive and resistant syngeneic mouse tumor models. Mechanistically, in addition to blocking PD-1 interaction with PD-L1, CTX-8371 bispecific antibody facilitated cell to cell bridging between cells expressing PD-1 and cells expressing PD-L1. Furthermore, we show that simultaneous binding of CTX-8371 to both PD-1 and PD-L1 resulted in long term PD-1 shedding. This suggests that CTX-8371 may prevent or overcome T cell exhaustion within the tumor microenvironment, thus providing additional advantage over existing therapies. Lastly, excellent tolerability was observed in non-human primates given 2 weekly drug infusions at up to 50 mg/kg dose. Conclusions: CTX-8371 displays multiple mechanisms of action over monoclonal PD1/PD-L1 blockade. These unique pharmacological properties of CTX-8371 could explain the enhanced T cell responses to tumor antigens and superior efficacy over current monoclonal antibody therapies. With favorable PK/PD and toxicology profiles in mice and cynomolgus monkeys, CTX-8371 warrants further advancement to clinical testing.
An open-label, fixed-sequence study to evaluate the effect of encequidar (HM30181) an oral P-gp inhibitor, on the pharmacokinetics of dabigatran etexilate (a P-gp substrate) in healthy male volunteers.
Authors: Jackson Christopher G. C. A. , Hung Noelyn Anne , Cutler David , Kramer Douglas , Zhi Jay , Chan Wing Kai , Kwan Min-Fun Rudolf , Hung Tak
Published: June 2021

e15060

Background: Oral co-administration of encequidar (a selective, minimally absorbed oral P-gp inhibitor) 12.9 mg with paclitaxel (a P-gp substrate) 205 mg/m2 for 3 consecutive days per week can achieve comparable AUC exposure to that of IV paclitaxel 80mg/m2 with a significantly lowered Cmax and has been demonstrated its improved tumor response with reduced neuropathy compared to IV paclitaxel 175 mg/m2 Q3W for the treatment of patients with metastatic breast cancer. Because of its pharmacology as an inhibitor of P-gp, encequidar may increase the bioavailability of orally administered drugs that are substrates of P-gp, such as dabigatran etexilate. Methods: To determine the effect of a therapeutic dose and regimen (3 once-daily 12.9 mg doses) of encequidar on the single dose PK of dabigatran etexilate, an open-label, fixed-sequence study was performed in 20 healthy male subjects. Participants received a single oral dose of dabigatran etexilate 75 mg on Day 1 of Treatment Period 1 (reference) and, after a washout period of at least 7 days, on Days 3, 17 and 31 of Treatment Period 2, after receiving once-daily oral doses 12.9 mg encequidar on Days 1 to 3 of Period 2. The PK sampling for determination of plasma concentrations of total and unconjugated dabigatran lasted up to 48 hours postdose of each dabigatran etexilate dosr. Results: Mean AUC and Cmax values for dabigatran were both increased ̃ 95% without changing t½ when dabigatran etexilate was administered 1 hour post the 3rd dose of 12.9 mg encequidar compared to when dabigatran etexilate was administered alone. When dabigatran etexilate was administered 2 weeks after encequidar administration, no apparent differences in dabigatran AUC or Cmax were detected compared to those of dabigatran etexilate alone. When administered 4 weeks after discontinuation of encequidar, dabigatran AUC and Cmax were both slightly lower than Reference dabigatran etexilate (̃ 25% lower for AUC and 34% lower for Cmax). Both unconjugated and total dabigatran PK data were analyzed and shown to be similar. Encequidar and dabigatran etexilate were well tolerated and had acceptable safety findings in this healthy subject population. Conclusions: Concomitant dosing of encequidar with dabigatran etexilate resulted in < 2-fold increase in exposure to dabigatran, which had abated by the time of the first re-test, 14 days after the last dose of encequidar. The observed changes do not warrant dose adjustment of dabigatran etexilate when administered with encequidar. Clinical trial information: ACTRN12618000791235.
Synthetic lethality of LP-184, a next generation acylfulvene, in ex vivo PDX models with homologous recombination defects.
Authors: Kulkarni Aditya , Restifo Diana , Astsaturov Igor A. , Kathad Umesh , McDermott Joseph , Bhatia Kishor , Sharma Panna
Published: June 2021

e15064

Background: The clinical success of PARP inhibitors (PARPi) in homologous recombination (HR) deficient (HRD+) solid tumors has broadened the scope of identifying additional agents and vulnerabilities in cancers with DNA repair deficiencies. However, with more than 40% of BRCA1/2-deficient patients failing to respond to PARPi or acquiring resistance with prolonged PARPi administration, newer agents are also needed. LP-184, an acylfulvene, is a prodrug activated by PTGR1. Threshold expression levels of PTGR1 are higher in several tumors, providing a window of specificity for its cytotoxic action. DNA damage inflicted by acylfulvene (AF) agents is reliant upon HR pathway genes including BRCA1 for correction and removal. We hypothesized that tumors with high PTGR1 expression and HR deficiency will therefore be uniquely targeted and demonstrate synthetic lethality when exposed to LP-184. Methods: We evaluated ex vivo antitumor activity of LP-184 in selected PDX models representing lung, pancreatic and prostate cancers with high PTGR1 and known HR defects. Dissociated tumor fragments were treated with LP-184 across a concentration range of 5 nM to 36 uM for 5 days. Cell viability was quantified by CellTiter Glo. LP-184 IC50s were compared with PARPi efficacy. We further confirmed the dependency of PTGR1 in HR deficient tumor cells by comparing LP-184 sensitivity in the BRCA2 deficient cell line CAPAN-1 and the ATM mutant cell line PANC03.27, with and without PTGR1 suppression following an engineered CRISPR knockout of PTGR1. We also analyzed TCGA data to estimate the percentage of tumors with elevated PTGR1 and co-occurring damaging mutations in a panel of 60 HR genes. Results: The mean LP-184 IC50 across 15 HRD+ PDX models tested was 288 nM (range 31 - 2900 nM). LP-184 turned out to be 6 - 340X more potent ex vivo than the PARPi Olaparib in these models. 9 of 15 models were associated with no clinical response to or initial response followed by progression on approved standard of care (SOC) agents. 6 of 15 models showed < 10% tumor growth inhibition in vivo with SOC treatment. Regardless of cancer type, models with high-impact, loss-of-function mutations in ATM, ATR and BRCA1 showed exquisite sensitivity to LP-184 (mean IC50 ̃ 60 nM). CRISPRi-mediated stable suppression of PTGR1 in the pancreatic cancer cell lines CAPAN-1 and PANC03.27 entirely abrogated LP-184 sensitivity relative to isogenic parental cell lines. 17.6% of lung adenocarcinomas (n = 517), 4.5% of pancreatic adenocarcinomas (n = 179) and 9.6% of prostate adenocarcinomas (n = 498) displayed elevated PTGR1 along with damaging HR related mutations, and are likely to be responsive to LP-184 based on analysis of TCGA data. Conclusions: LP-184 is broadly effective in HRD+ tumors that may be less responsive to SOC including PARPi and could be useful clinically in a subset of tumors with high PTGR1 and HR defects.
A small molecule LN435a targeting TGFβR1 exerts promising antitumor effects on breast cancer.
Authors: Zhang Yuzhu , Li Huachao , Zhang Hongyan , Liu Xiaoyuan , Luo Tianyu , Lin Huan , Xu Rui , Ren Liping , Li Dongmei , Chen Yihua , Chen Qianjun , Huang Luqi
Published: June 2021

e15069

Background: Breast cancer has overtaken lung cancer as the most diagnosed cancer. Despite conventional treatment, metastases occur in 20-30% of patients, resulting in death. This study aims to screen of effective drugs by metastatic patient-derived organoid and the potential molecular mechanism. Methods: Breast Cancer patient-derived organoid (PDO) model was established from the patient who have multiple drug resistance, multiple visceral and contralateral breast metastases. The organoid morphologies was tested by hematoxylin-eosin (HE) staining and immunohistochemistry (IHC). Then, pharmacological activity assay of 2370 natural product monomer (from Selleck) was performed with organoids. we modified the structure of harmine(HM) and screened the best active drugs. Cell proliferation assay and wound healing assay were used to detect LN435a anticancer activity in vitro. Orthotopic, Metastatic Xenograft and Patient-Derived tumor Xenograft(PDX) model of Breast Cancer were used to detect LN435a anticancer activity in vivo. In order to explore the anti-cancer target of LN435a, we used RNA transcriptome and proteomics sequencing. To further validate anti-cancer targets,TGFβ receptor 1 (TGFβR1), we used real-time quantitative qPCR, western blot, lentiviral packing and biolayer interferometry assay. To investigate whether LN435a inhabition of EMT and stem cell markers, we performed flow cytometry, immunohistochemistry and fluorescence. Results: We observe that organoid morphologies typically matched the histopathology, hormone receptor status, and HER2 status of the original tumor. In the first anti-cancer drug screening, HM showes the best effect on PDO. Because HM contains β-carbine alkaloids as the structural units, we designe a series of active drugs based on this and did anticancer screening. We find LN435a as one of the lead compounds exerting anti-metastatic activity in the nanomolar range in PDO and breast cancer cells. Proteomic and biochemical studies identify TGFβR1 as the direct target of LN435a. And then it inhibits EMT and stem cell markers. In parallel, loss of TGFβR1 or pharmacological inhibition of TGFβR1 by LN435a reduces breast cancer extravasation into the lung in an experimental metastasis mouse model, which reveals an essential role of TGFβR1 in breast cancer progression. Conclusions: Altogether, LN435a is a novel inhibitor of promising anti-tumor effects on breast cancer that works by blocking TGFβ signaling pathways.
AKT1 in Asia: A landscape analysis of 11,813 Chinese tumor samples.
Authors: Gan PingPing , Lu Yi , Li Xiaomin , Deng Wanglong , Han Tiantian , Xi Wenjing , Bu Fanfeng
Published: June 2021

e15076

Background: AKT1 is a key gene of the PI3K/AKT/mTOR signalling pathway. The recent development of ATK1 targeting drug has shown great promise. It is thus important to understand the genomic landscape of AKT1 in a specific population. Methods: Sequencing data of 11,813 tumor samples collected in China were analyzed for AKT1 mutation status. Concomitant genomic aberrations were further analyzed in tumors with AKT1 mutations. Results: AKT1 mutations were observed in 173 of 11,813 tumor samples (1.46%). The most common cancer types in our study included cervical (5.17%), breast (5.15%), osteosarcoma (3.51%), brain (3.14%), head and neck (2.22%) and colorectal (2.02%). We included only the cancer types with more than 50 cases for analysis. Higher frequencies of AKT1 mutation were observed in several common cancers, including brain glioma (1.38% vs 0.67%), breast (5.15% vs 2.92%), cervical (5.17% vs 2.93%), head and neck (2.22% vs 0.95%) cancer in our study than reported in TCGA, but showed no statistically significant difference. The main common type of variation was SNV/Indel (67.63%), amplification (20.81%), deletion (11.56%), fusion (1.16%). Among SNV/Indel, E17K accounted for 32.9% of AKT1 mutations, Q79K only accounted for 0.79%. AKT1 E17K was more commonly found in breast (3.09%), brain (1.96%) and cervical (1.72%) cancer. There is no significant difference between wild type and AKT1 mutated groups in age and gender. III/IV patients were identified in 85.71% of AKT1 mutated group, which were significantly higher than that of wild type group (73.76%; p<0.05).MSI-H patients were identified in 12.16% of AKT1 mutated group, which were significantly higher than that of wild type group (1.8%; p<0.001). Frequencies of specific concomitant aberrations varied between AKT1 mutated and wild type groups. In AKT1 mutated groups, 57 genes are more frequently altered, and mainly enriched in PI3K-Akt signaling pathway (26, q= 2.97E-20, MAPK signaling pathway (count:20, q= 9.74E-15), Central carbon metabolism in cancer (19, q= 5.01E-25), EGFR tyrosine kinase inhibitor resistance (19, q= 3.36E-24). Conclusions: Higher frequencies of AKT1 mutation were observed in several common cancers, including cervical, breast, osteosarcoma, brain, head and neck and colorectal in our study. AKT1 E17K was more commonly found in breast, brain and cervical cancer. Given the fact that III/IV patients are more common in AKT1 mutated group, it coincides with AKT inhibitors therapeutic prospects in a variety of advanced malignant tumors. MSI-H patients are more common in AKT1 mutated group, and it provides possibility for immune checkpoint inhibitors combined with targeted signaling pathway therapy. The enrichment results of the common co-occurring genes with AKT1 in this study, further investigations are still needed.
NTRK genomic alterations in Latin-American cancer patients.
Authors: Carvajal Daniel , Salas Claudio , Marcelain Katherine , Perez Francisco , Rivas Solange , Feliu Evelin , Schalper Kurt , Armisén Ricardo
Published: June 2021

e15088

Background: The neurotrophic receptor tyrosine kinase genes NTRK1-3 encode the tropomyosin receptor kinase proteins TRKA, TRKB, and TRKC, respectively. TRK fusions lead to overexpression of the chimeric protein, resulting in constitutively active, ligand-independent downstream signaling. NTRKs act as oncogenes, they can be targeted, and it is estimated that they are present in up to 0.3% of tumors. The incidence of actionable alterations in these genes is currently unknown in Latin-American patients. We investigated the presence of NTRK1-3 mutations/rearrangements in 3 independent patient series from several tertiary hospitals from Chile, Brazil and Peru. Methods: 1795 FFPE tumor samples from multiple institutions divided in 3 series were analyzed using 2 NGS panels: Oncomine Focus Assay (OFA; 52 genes) and Oncomine Comprehensive Assay (OCA; 161 genes. Data on NTRK1-3 SNVs, indels, CNVs and fusions was obtained. Bioinformatic workflows were used to study the biologic significance of these alterations. Results: Using OCA (series 1-2), 31 somatic variants were found in gastric, CRC, gallbladder, lung and pancreatic cases out of 300 patients. From these, 13 were located in the tyrosine kinase domain. Using OFA, no NTRK alteration were detected (series 3, 1495 NSCLC cases). Conclusions: NTRK alterations are rare events in Latin-American cancer patients. In 3 series including 1,795 patients, 31 somatic variants were detected. No NTRK fusions or CNVs were identified. The presence of NTRK mutations in the tyrosine kinase domain warrants further research into their potential to benefit from FDA/EMA-approved targeted therapies. [Table: see text]
Significance of scores generated by a cancer therapy matching engine for patient outcomes.
Authors: Romm Eden , Davtyan Emiliya , Bush Kevin , Sutton Ramsay , Patek Itamar , Imanirad Parisa , Sicklick Jason K. , Kato Shumei , Kurzrock Razelle , Perlina Ally
Published: June 2021

e15099

Background: Oncologists need effective precision medicine tools to navigate the myriad of therapeutic options to best address each patient’s unique tumor profile. Described here is a digital solution, which analyzes tumor data (NGS, other), scores and ranks known and novel matching combinations of cancer drugs with molecular-level precision for decision support. Methods: All data was acquired through the published supplement of the I-PREDICT study (NCT 02534675). Patients with available progression-free survival (PFS) and sequencing data who received <3 matched non-experimental cancer drugs in combination were evaluated (n=77). To determine each patient’s score (represented as %), the system used expert-curated content integrated within an artificial intelligence (AI) reasoning framework that computed how well the therapy each patient received matched their tumor’s genomic profile. The dataset was binarized at 36 possible thresholds, which split the data into higher vs. lower score bins. Relationship with PFS and the degree of separation between bins were evaluated with Kaplan-Meier plots and assessed in terms of p-value, hazard ratios and related confidence intervals (Table). To prevent sampling bias at high and low thresholding extremes of this dataset, a limit was imposed of >25 patients per bin. Results: Significant separation between high and low scoring bins was observed in >73% of evaluable thresholds. The mean p-value was 0.044 (range, 0.011-0.13). The hazard ratio was consistently ̃2 (mean, 1.81, range, 1.50-2.08). A similar level of statistical significance was observed for all thresholds up to and including bracketing of 60% (Table). Despite the small size of this dataset and the disparity between sample counts per bin seen at the end of the range, significant p-values of <0.05 were achieved. Ex., a 68-year-old female with an ampullary adenocarcinoma harboring ERBB2, CDK6 and other alterations, received a combination of pertuzumab and trastuzumab scoring 56%. The PFS was 745+ days. However, had palbociclib, ribociclib, or abemaciclib been added to this combination, the resulting 3-drug options would have scored even higher due to also targeting CDK6. Conclusions: Our study indicates that therapies with higher scores were predictive of better PFS, while lower scores were predictive of worse PFS. The results will be validated on larger datasets but are already demonstrative of the utility of this system in providing digital guidance to oncologists in selecting the most suitable treatment. [Table: see text]
Comparing of pan-cancer tumor immune microenvironment.
Authors: Ye Jiaan , Cui Longgang , Zhao Xiaochen , Lan Guanghui
Published: June 2021

e15100

Background: Cancer treatment has entered the era of immune checkpoint inhibitors (ICI), but different tumors have different responses to ICI drugs. For example, non-small cell lung cancer and melanoma have higher response rates to ICIs than colorectal cancer and liver cancer patients. Previous studies have shown that tumor immune microenvironment have a great impact on the efficacy of ICI. Methods: This study retrospectively included pan-cancer patient specimens, using multiple fluorescent labeling immunohistochemistry to explore the differences in the immune microenvironment of different tumors. Shapiro-Wilk was used for normality test, and ANOVA or Kruskal Wallis test was used according to the results. Two-sided P < 0.05 was considered a significant difference. Results: The study included 308 patients, including 119 (38.6%) NSCLC patients, 72 (23.4%) Colorectal cancer patients, 51 (16.6%) Hepatobiliary cancer patients and 66 (21.4%) Others types of cancer patients. Among them, there was 192 (62.3%) Male, and 116 (37.7%) Female, and the median age was 57 (50-66). The proportion of CD8+ T cells and natural killer cell in tumor was statistically different. The proportion of CD8+ T cells in NSCLC, Colorectal cancer, Hepatobiliary cancer and others was 2.16%, 1%, 1.77% and 2.63%, p < 0.01; the proportion of natural killer cell was 16.44 %, 4.91%, 5.58% and 3.29%, p < 0.01. Conclusions: Different tumor types have different immune microenvironments. These results may provide valuable clues for future ICI trail design.