bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2183: Protective autophagy elicited by RAF®MEK®ERK inhibition suggests a treatment strategy for pancreatic cancer
Authors: Kinsey Conan G. , Camolotto Soledad A. , Boespflug Amelie M. , Guillen Katrin P. , Foth Mona , Truong Amanda , Schuman Sophia S. , Shea Jill E. , Seipp Michael T. , Yap Jeffrey T. , Burrell Lance D. , Lum David H. , Whisenant Jonathan R. , Gilcrease G. Weldon , Cavalieri Courtney C. , Rehbein Kaitrin M. , Cutler Stephanie L. , Affolter Kajsa E. , Welm Alana L. , Welm Bryan E. , Scaife Courtney L. , Snyder Eric L. , McMahon Martin
Published: August 2019

Pancreatic ductal adenocarcinoma was responsible for ~43,000 deaths in the USA in 2017 and is the epitome of a recalcitrant cancer driven by an, as yet, pharmacologically intractable oncoprotein, KRAS. Although the clinical picture remains grim, the mechanisms by which key alterations in tumor suppressors and proto-oncogenes contribute to PDA have been dissected. Downstream of KRAS, the RAF→MEK→ERK signaling pathway plays a central role in pancreatic carcinogenesis. However, to date, pharmacological inhibition of this pathway has provided no clinical benefit to PDA patients. Here we show that inhibition of KRAS→RAF→MEK→ERK signaling in PDA cell lines elicits autophagy, a process of cellular recycling that protects pancreatic cancer cells from the potentially cytotoxic effects of KRAS pathway inhibition. Furthermore, combined inhibition of MEK1/2 plus autophagy displays synergistic anti-proliferative effects against PDA cell lines in vitro, and promotes regression of xenografted patient-derived PDA tumors in mice. Finally, treatment of a PDA patient with the combination of trametinib plus hydroxychloroquine resulted in a partial, but nonetheless striking disease response. These data suggest that this combination therapy may represent a novel strategy to target RAS-driven cancers such as PDA.

Citation Format: Conan G. Kinsey, Soledad A. Camolotto, Amelie M. Boespflug, Katrin P. Guillen, Mona Foth, Amanda Truong, Sophia S. Schuman, Jill E. Shea, Michael T. Seipp, Jeffrey T. Yap, Lance D. Burrell, David H. Lum, Jonathan R. Whisenant, G. Weldon Gilcrease, Courtney C. Cavalieri, Kaitrin M. Rehbein, Stephanie L. Cutler, Kajsa E. Affolter, Alana L. Welm, Bryan E. Welm, Courtney L. Scaife, Eric L. Snyder, Martin McMahon. Protective autophagy elicited by RAF®MEK®ERK inhibition suggests a treatment strategy for pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2183.
Abstract 2185: Efficacy of Rac/Cdc42 inhibitors in multiple cancer models
Authors: Calderon Jean F. Ruiz , Castillo-Pichardo Linette , Lopez-Lopez Irmaris , Dharmawardhane Suranganie
Published: August 2019

The RhoGTPases Rac and Cdc42 are excellent therapeutic targets for inhibiting metastasis because these homologous proteins regulate numerous cellular functions that promote tumorigenesis, angiogenesis, invasion and therefore, metastasis. Also, Rac1, Cdc42, and their effector p21-activated kinase (PAK) are overexpressed in multiple cancer types, including breast, gastric, and pancreatic, and are associated with higher mortality rates. Furthermore, Rac and Cdc42 are activated through the signaling of oncogenic cell surface receptors such as EGFR and HER2, therefore hyperactivating their signaling pathways. Our laboratory has developed two small molecules targeting Rac and Cdc42, Ehop-016 and MBQ-167, that inhibit Rac1 and Cdc42 with IC50s in the low micro- and nano-molar concentrations, respectively (Montalvo-Ortiz et al. 2012, J Biol Chem; Humphries-Bickley et al. 2017, Mol Cancer Therap), making them the most potent Rac/Cdc42 inhibitors developed to date. We hypothesize that targeting Rac1, 3, and Cdc42 is a rational strategy to inhibit aggressive TNBC, pancreatic and gastric cancer. To test this hypothesis, we treated the TNBC cell lines MDA-MB-231, MDA-MB-468 and 4T1 (murine breast cancer), gastric cancer cells NCI-N87, and pancreatic cancer cells MIA-PaCa-2 with 4uM and 500nM of Ehop-016 and MBQ-167 respectively, and measured cell viability. Both TNBC cell lines and the pancreatic cancer cell line exhibited a drastic reduction in cell viability (>60%) after 48hrs of treatment Ehop-016 or MBQ-167. Additionally, experimental and spontaneous metastasis assays were performed with TNBC and gastric cancer xenografts in immunocompromised and immunocompetent mice. Treatment with 1 mg/kg BW MBQ-167 3X week resulted in reduced tumor growth by 60-90% in TNBC and gastric cancer models, and metastasis inhibition by ~90% in the MDA-MB-231 model. In the syngeneic experimental metastasis study with 4T1, both Ehop-016 and MBQ-167 reduced lung metastasis in comparison to the vehicle treated group. Moreover, these compounds were not toxic to both immunocompromised and immunocompetent mice, as observed by no significant changes in weight and liver enzymes. Therefore, both Ehop-016 and MBQ-167 are promising targeted therapeutics against cancers that overexpress Rac, Cdc42, and PAK.

Citation Format: Jean F. Ruiz Calderon, Linette Castillo-Pichardo, Irmaris Lopez-Lopez, Suranganie Dharmawardhane. Efficacy of Rac/Cdc42 inhibitors in multiple cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2185.
Abstract 2187: Tryptophan Metabolism Plays a Central Role in Immunosuppression
Authors: Geeganage Sandaruwan , Sams Lillian , Henry James , Dorsey Frank , Roth Kenneth , Nikolayev Alexander , Benhadji Karim , Gilmour Raymond , Cerezo Ana , Peregrina Sandra , Hoyo Gloria Martínez-del , Campos-Olivas Ramón , Funes Juan Manuel , Diezma Laura , Velasco-Miguel Susana
Published: August 2019

Tryptophan metabolism plays a central role in immunosuppression through the local depletion of tryptophan with the concomitant production and accumulation of kynurenine, both of which are immunosuppressive. Indoleamine 2,3-dioxygenase 1 (IDO1), a heme-dependent enzyme, catalyzes the initial and rate-limiting step of the kynurenine pathway. Tumor cells selectively upregulate IDO1 as an immune-evasion mechanism either through intrinsic expression of IDO1, or in response to IFN-γ, a cytokine secreted by immune cells during an active immune response.

In addition to the immuno-suppressive role of IDO1 and kynurenine, we investigated their additional roles in tumor cells. Here we describe an NMR-based readout assay for IDO1 activity, in which we are able to trace the extra- and intracellular destination of the single tryptophan carbon groups. This assay allowed us to detect contribution of the tryptophan catabolism to purine synthesis in tumor cells, and suggested further roles of tryptophan catabolism in tumors, apart from kynurenine production, that contribute to the modulation of tumor growth. We detected the incorporation of labeled tryptophan carbon units in intracellular purine pools. Our highly potent, orally available, and CNS penetrant small molecule inhibitor of IDO1 (LY3381916) inhibits this incorporation of tryptophan-derived one carbon units into purines in vitro and in vivo. This activity leads to pre-clinical efficacy and may be an added advantage of inhibiting IDO1 in cancer.

Citation Format: Sandaruwan Geeganage, Lillian Sams, James Henry, Frank Dorsey, Kenneth Roth, Alexander Nikolayev, Karim Benhadji, Raymond Gilmour, Ana Cerezo, Sandra Peregrina, Gloria Martínez-del Hoyo, Ramón Campos-Olivas, Juan Manuel Funes, Laura Diezma, Susana Velasco-Miguel. Tryptophan Metabolism Plays a Central Role in Immunosuppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2187.
Abstract 2189: Evaluating efficacy of repurposed drugs in treatment of glioblastoma
Authors: Neal Tristan , Tang Nanyun , Reid George , Byron Sara , Dhruv Harshil , Berens Michael
Published: August 2019

Glioblastoma (GB) is the most aggressive adult brain tumor with a devastating median survival time of about fourteen months post-surgery and standard of care therapy with radiation and temozolomide. The low incidence of GB is a dis-incentive to develop novel therapies. To overcome that obstacle, we investigated the efficacy of repurposing four FDA approved drugs known to cross the BBB, minocycline, propranolol, chlorpromazine, and metformin, to inhibit signaling and metabolism in GB cells. Minocycline is a tetracycline class broad spectrum antibiotic commonly used to treat severe acne and other skin infections. Propranolol is a beta blocker type heart medication primarily used to treat high blood pressure and irregular heartbeat. Chlorpromazine is a phenothiazine antipsychotic usually used for schizophrenia. Metformin is the most widely used first-line oral treatment for type-2 diabetes. Based on a literature survey, minocycline is expected to prevent the phosphorylation of STAT3, a transcription factor downstream of EGFR; propranolol is expected to disrupt EGFR trafficking; chlorpromazine is expected to target the mTOR/AKT pathway; metformin is believed to exploit vulnerabilities in cancer cell metabolism. Efficacy of minocycline in inhibiting EGFR-driven STAT3 activation was investigated using western blot analysis. Our results demonstrate that Minocycline effectively inhibits activation of EGFR-driven STAT3 in U373 glioma cells at 100μM. The ability of chlorpromazine to inhibit the mTOR/AKT pathway was similarly tested via western blot, which showed inhibition of phosphorylated Akt and S6 at 10μM. Efficacy of propranolol in perturbing EGFR trafficking was evaluated using flow cytometry and immunofluorescence, which depicted altered membrane-associated EGFR abundance. Finally, concentration-dependent inhibition of colony formation was tested for all four drugs. Propranolol and minocycline showed potential stimulatory effects at 10μM, but all drugs inhibited cell growth at 50μM and higher. Efficacy of these drugs in treatment of GB is being further evaluated using in vitro neurosphere cultures from patients identified as having the cellular vulnerabilities potentially targeted by these drugs. Successful completion of this project will lead to in vivo efficacy testing of these four drugs in orthotopic GB PDX models.

Citation Format: Tristan Neal, Nanyun Tang, George Reid, Sara Byron, Harshil Dhruv, Michael Berens. Evaluating efficacy of repurposed drugs in treatment of glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2189.
Abstract 2195: Real-world experience with afatinib as first-line treatment in advanced non-small cell lung cancer in a university hospital
Authors: Jang Taewon , Kim Jaehoon
Published: August 2019

Purpose: In patients with non-small cell lung cancer (NSCLC), mutations in the epidermal growth factor receptor (EGFR) have been associated with sensitivity to EFGR-Tyrosin Kinase Inhibitors (TKIs). The aim of this study was to analyze clinical efficacies of first line afatinib treatment.

Experimental Design: We collected patients with EGFR mutation positive NSCLC who started first-line afatinib treatment between Jan 2015 and Sep 2017. We reviewed clinical characteristics of the patients, laboratory finding, tumor response rate, side effects, progression free survival (PFS), 2nd biopsy result, and overall survival (OS), retrospectively.

Results: Forty six patients were available for analysis among all 62 patients (1st to 8th line chemotherapy). At the time of data analysis in October 2018, nine patients were still on afatinib. Median age was 67.4 years-old. 20 (44%) patient had EGFR 19 deletion, 25 (54%) EGFR L858R mutation, 1 (2%) S768I. 10 patients (21.7%) had brain metastasis at baseline study. Tumor response was 58.7% with partial response, no change 19.6%, progressive disease 2%, and non-evaluable 19.6%. Mean follow-up time was 21.6 months (13.5 to 42.5 month) from start of afatinib treatment. Twenty-eight progression or death events were observed. Median PFS was 13.5 months (95% CI: 10.5 to 16.5 months). There were no differences in smoking, gender, EGFR types. 37 patients (80.4%) had dose reduction from initial dose. 6 patients (13.1%) discontinued due to afatinib toxicity. 6 patients (13%) had G3 toxicity, but usually manageable by dose reduction or supportive care. 1 case of doxycycline induced pneumonitis occurred. The PFS of dose reduction group due to adverse events was longer than standard group (14.9 months vs 10.1 months, p=0.023). In dose reduction group, body weight was low (53.1 kg vs 68.4 kg, p=0.002), the ratio of never smoker (87.9% vs 37.5%, p=0.001) and female (87.4% vs 47.4%, p=0.048) were higher than standard group. After progression, 2nd biopsy was done in 17 patients. T790M mutation was detected in 8 patients, so detection rate was 47.1%. 6 patients received oximertinib treatment. Median OS was 38.6 months.

Conclusions: Afatinib is one of appropriate first-line treatment option for patients with advanced EGFR positive NSCLC. The dose reduction due to adverse events was common in low-body weight female never smoking patients, but the PFS was increased. With use of new 3rd-gen EGFR TKIs in the first-line setting, optimal sequencing of EGFR TKIs needs to be validated in maximizes treatment outcomes.

Citation Format: Taewon Jang, Jaehoon Kim. Real-world experience with afatinib as first-line treatment in advanced non-small cell lung cancer in a university hospital [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2195.
Abstract 2199: BOS172738: A novel highly potent and selective RET kinase inhibitor in Phase 1 clinical development
Authors: Keegan Mitchell , Wilcoxen Keith , Ho Peter T.
Published: August 2019

Background: RET gene alterations including point mutations and gene fusions have been detected in multiple cancers, including colorectal cancer, NSCLC and various thyroid cancers. RET fusion events involve the kinesin family member 5B (KIF5B-RET), as well as a number of other fusion partners such as CCDC6, TRIM33, and NCOA, and are considered to be the driver mutations in NSCLC and other tumors harboring RET fusions. Selective inhibitors of RET kinase recently has been associated with durable objective responses in patients with RET gene-altered NSCLC and papillary thyroid carcinoma. BOS172738 (formerly DS-5010) is an orally available small-molecule RET kinase inhibitor that has previously been shown to have potent in vitro RET inhibitor activity and in vivo potency against transfected allograft and xenograft models. In this study, we discuss additional in vitro characterization and in vivo efficacy in patient derived xenograft (PDX) models driven by RET fusions and emergent resistant mutations.

Results: An expanded kinase profile was conducted with BOS172738 against over 450 kinases, and demonstrated exquisite potency for RET and RET mutations with Kd values ≤ 1 nM for RETwt, RET(M918T), RET(V804L) and RET(V804M). BOS172738 also demonstrated high selectivity over KDR (>300 fold). To confirm this selectivity in vitro, BOS172738 was compared to ponatinib, a multikinase inhibitor with potent activity against both KDR and RET. While BOS172738 demonstrated similar potency to ponatinib (IC50 0.5 µM vs. 0.3 µM) in inhibiting the in vitro proliferation of a NCO4-RET human CRC cell line (CR1520), it resulted in significantly less anti-proliferative activity in HUVEC normal endothelial cells (IC50 2.9 µM vs 0.2 µM). BOS172738 was also evaluated in a number of PDX models of RET fusion cancers, including three CRC PDX models (CR2518:CCDC6-RET, CR1520: NCOA4-RET and CR2545: CCDC6-RET with a V804M mutation) and one NSCLC PDX model (CTG-0838:KIF5B-RET). In all cases tested, BOS172738 demonstrated potent and durable tumor regression at doses of 30 mg/kg, with tumor growth inhibition at lower doses. BOS172738 also demonstrated potent tumor regression on larger, established tumors (>1000 mm3).

Conclusion: These results support that BOS172738 is a potent and selective RET kinase inhibitor, with strong antitumor activity against a range of RET fusion proteins, as well as resistant RET active site mutations. BOS172738 is currently being evaluated in a Phase 1 clinical study.

Citation Format: Mitchell Keegan, Keith Wilcoxen, Peter T. Ho. BOS172738: A novel highly potent and selective RET kinase inhibitor in Phase 1 clinical development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2199.
Abstract 2205: CBT-102, an oral small molecule multi-kinase inhibitor, demonstrates favorable CSF-1R activity, offering means of controlling tumor associated macrophages
Authors: Liu Elaine , Yang Lan , Choy Gavin , Zhang Xiaoling , Pearce Tillman , Reddy Mamatha , Redkar Sanjeev , Shi Qian
Published: August 2019

Background: The role of the tumor microenvironment (TME) in fostering the development of malignancies is prompting the pursuit of anticancer therapies that target its components as to opposed to the tumor itself. As part of their immune surveillance duties, immune cells form part of this microenvironment, and yet, cancer cells have devised means to downplay their tumoricidal capabilities. Colony stimulating factor 1 receptor (CSF-1R) may offer such a means of controlling tumor associated macrophages in the tumor microenvironment.

Methods: The IC50 of CBT-102 against CSF1R was evaluated in both cell free system and cell based assays, where enzyme activity was evaluated with radiometric assay format, and cell growth inhibition was evaluated in the Ba/F3 hCSF1R cell line by the CellTiter-Glo (CTG) method. Further evaluation of CBT-102 IC50 was undertaken in M-NFS-60 syngeneic cell lines and human monocytes. In vivo studies of CBT-102 in combination with anti-PD1 antibody were performed in H22 and MC38 syngeneic models.

Results: CBT-102 demonstrated a reproducible activity against CSF1R in a radiometric enzyme activity assay. Using engineered BaF3 hCSF1R cell lines, we demonstrated the IC50 value of CBT-102 was 0.588 µM compared to 1.333 µM of sulfatinib, a similar multi-kinase inhibitor, and 0.279 µM of GW2580, a more specific CSF1R kinase inhibitor. Similar studies conducted in M-NFS-60 cell lines and human monocyte also showed sub-micromolar activities of CBT-102. In vivo study combining CBT-102 with anti-PD1 antibody in syngeneic models demonstrated a favorable combination effect compared to each of the single agents.

Conclusions: We demonstrated previously that CBT-102 effectively inhibits VEGFR and angiogenesis. The newly revealed mechanism of CBT-102, inhibiting CSF1R and macrophages, offers a promising dual mechanism of action in addition to targeting VEGFR and angiogenesis. Rational combination with check-point inhibitors (CPIs) may improve the efficacy of CBT-102 and broaden the impact of CPIs. Further studies may be needed to delineate the interplay among CBT-102’s different mechanisms and its impact in combination with CPIs.

Citation Format: Elaine Liu, Lan Yang, Gavin Choy, Xiaoling Zhang, Tillman Pearce, Mamatha Reddy, Sanjeev Redkar, Qian Shi. CBT-102, an oral small molecule multi-kinase inhibitor, demonstrates favorable CSF-1R activity, offering means of controlling tumor associated macrophages [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2205.
Abstract 2210: Discovery of a novel third generation EGFR TKI and identification of its potential mechanism of resistance
Authors: Zhang Tao , Qu Rong , Chen Chengbin , Tong Linjiang , Feng Fang , Chen Yi , Geng Meiyu , Ding Ke , Xie Hua , Ding Jian
Published: August 2019

Non-small cell lung cancer (NSCLC) patients with activating EGFR mutations initially respond to first generation EGFR inhibitors (such as gefitinib and elortinib), however, clinical efficacy is limited by acquired resistance, frequently driven by the EGFR T790M mutation. In this work, we identified a pyrimidine-backbone compound D7 as a novel irreversible inhibitor of EGFR T790M mutation and sensitizing mutation, with selectivity over the wild type EGFR (WT EGFR). D7 exhibited potent in vitro anti-proliferation activity in tumor cells harboring EGFR T790M (NCI-H1975 cells) and sensitizing mutations (PC-9 and HCC827 cells), whereas it showed moderate or weak inhibition in cells expressing WT EGFR. Oral administration of D7 induced dramatic tumor regression in EGFR mutated NSCLC tumor xenograft models as well as in patient-derived xenograft (PDX) model harboring EGFR T790M mutation. To further explore potential mechanisms mediating drug resistance, we constructed D7-resistant cells by prolonged exposure of parental NCI-H1975 cells to gradually increasing concentration of this compound. Resistant cells exhibited signs of apoptosis-resistance to D7 treatment and demonstrated increased sensitivity when combined with Src kinase inhibitors both in vitro and in vivo. Mechanistic investigation revealed that hyperphosphorylation of Src and subsequent activation of anti-apoptotic signaling via Akt pathway contributed to D7 resistance. These results support D7 as a promising third generation EGFR inhibitor deserving further development, and identify potential combination strategies with potential biomarkers for NSCLC tumors that is resistant to this compound.

Keywords: EGFR inhibitor, lung cancer, drug resistance, combination therapy

Citation Format: Tao Zhang, Rong Qu, Chengbin Chen, Linjiang Tong, Fang Feng, Yi Chen, Meiyu Geng, Ke Ding, Hua Xie, Jian Ding. Discovery of a novel third generation EGFR TKI and identification of its potential mechanism of resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2210.
Abstract 2211: Discovery of orally available and potent MPS1(TTK) kinase inhibitors for anti-cancer drugs
Authors: Lee Younho , Kang Juhee , Kang Sein , Kim Hyunkyung , Seo Kyung-ah , Choi Jieun , Ahn Jaeyoung , Son Jung Beom , Kim Nam Doo
Published: August 2019

There are many cell cycle regulating kinases such as CDKs, Polo, Aurora, Bub, NEK/NimA, etc. Dual specificity protein kinase, MPS1(TTK) is one of them. The role of MPS1 is to regulate spindle assembly checkpoint(SAC) during cell mitosis. The inhibition of MPS1 activity causes cell death by premature mitotic exit. Some of MPS1 inhibitors (NMS-P153, BOS-172722, CFI-402257, BAY-1217389, and BAY-1161909) are in clinical trials. These compounds are treated for solid tumors with or without Paclitaxel. We discovered new orally available and potent inhibitors of MPS1 kinase (MPS1 enzyme IC50 < 10nM / MDA-MB-231 GI50 < 50nM, MDA-MB-468 GI50 < 50nM / HCT116 GI50 < 50nM, F% > 30%). The compounds selectively inhibit MPS1 based on kinase profiling and decrease phosphorylation of MPS1 and Phospho-HH3 signaling, effectively. Some of compounds were selected for further preclinical studies.

Citation Format: Younho Lee, Juhee Kang, Sein Kang, Hyunkyung Kim, Kyung-ah Seo, Jieun Choi, Jaeyoung Ahn, Jung Beom Son, Nam Doo Kim. Discovery of orally available and potent MPS1(TTK) kinase inhibitors for anti-cancer drugs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2211.
Abstract 2215: SLFN11 and EZH2 protein expression and localization in circulating tumor cells to predict response or resistance to DNA damaging therapies in small cell lung cancer
Authors: Byers Lauren Averett , Stewart Allison , Gay Carl , Heymach John , Fernandez Luisa , Lu David , Rich Robin , Chu Lincy , Wang Yipeng , Dittamore Ryan
Published: August 2019

Background: A recent phase II trial in recurrent small cell lung cancer (SCLC) demonstrated that patients with high SLFN11 expression in tissue biopsies had improved survival when treated with PARP inhibition (PARPi). EZH2 is also highly expressed in SCLC tissue and is associated with chemo-resistance through epigenetic silencing of SLFN11. Here we developed a liquid biopsy test to explore the clinical feasibility of utilizing SLFN11 and EZH2 expression on circulating tumor cells (CTCs) to aid in the prediction of drug response/resistance of DNA damaging chemotherapy or PARPi in SCLC.

Material and methods: 26 blood samples (19 baseline and 7 on-therapy) from SCLC patients treated at MDACC were sent to Epic Sciences for CTC enumeration (intact DAPI, CK+, cancer cell morphology) and characterization with SLFN11 and EZH2 immunoassays. Nuclear localization and expression of SLFN11 and EZH2 were quantified.

Results: 17/19 (89%) of baseline and 5/7 (71%) post-therapy samples had detectable CTCs. 19/26 (73%) samples had SLFN11+ CTCs and 8/26 (31%) samples had nuclear localized SLFN11. For samples with SLFN11+ CTCs detected, the percentage of SLFN11+ cells ranged from 2.4 to 100%. 10/26 (38%) samples had EZH2+ CTCs and 9/26 (35%) samples had nuclear localized EZH2. For samples with EZH2+ CTCs, the percentage of EZH2+ cells ranged from 19 to 83%. 6 samples had both nuclear localized SLFN11 and EZH2 CTCs detected.

Conclusions: Inter- and intra-sample heterogenous SLFN11 and EZH2 protein expression was observed in SCLC patients. Use of these assays is underway in SCLC patient samples to assess correlation with PARPi and platinum agent response.

Citation Format: Lauren Averett Byers, Allison Stewart, Carl Gay, John Heymach, Luisa Fernandez, David Lu, Robin Rich, Lincy Chu, Yipeng Wang, Ryan Dittamore. SLFN11 and EZH2 protein expression and localization in circulating tumor cells to predict response or resistance to DNA damaging therapies in small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2215.
Abstract 2224: A panel of autoantibodies against multiple tumor-associated antigens in the early immunodiagnosis of lung cancer
Authors: Jiang Di , Wang Tingting , Pei Lu , Wang Peng , Ye Hua , Song Chunhua , Wang Kaijuan , Zhang Jianying , Dai Liping
Published: August 2019

Lung cancer is the most common cancers worldwide and with very low 5-year survival rate due to lack of effective early diagnostic methods. In our previous study, nine autoantibodies against tumor-associated antigens (TAAs) were identified to be potential biomarkers in detection of lung cancer and this study aimed to find the optimal panel of TAAs to detect lung cancer in the early stage.

Methods A total of 1,130 serum samples were included in this study, including patients with lung cancer, normal control and benign lung diseases patients. Autoantibodies against PSIP1,TOP2A,ACTR3,RPS6KA5,HMGB3,MMP12,GREM1,ZWINT and NUSAP1 were detected by enzyme-linked immunosorbent assay. Logistic regression models were generated from the training set and further validated in another independent set. We also evaluated the ability of the model to detecting benign lung diseases and early-stage lung cancer.

Results Two greater models were selected. One model including 5 autoantibodies panel (GREM1,HMGB3,ZWINT,TOP2A and PSIP1) and another model including 3 autoantibodies panel (GREM1,HMGB3 and PSIP1). Finally, the panel of 3 autoantibodies showed high diagnostic accuracy with areas under the curve (AUC) of 0.711(95 % CI 0.674-0.746) in training cohort and 0.891(95% CI 0.845-0.927) in validation cohort, respectively. The AUC of the model of 3 autoantibodies panel was 0.948(95% CI 0.912-0.972) in discriminate lung cancer and benign pulmonary disease. This model could discriminate early-stage lung cancer patients from normal controls, with AUC of 0.686(95 % CI 0.633-0.735) in training cohort and AUC of 0.911(95 % CI 0.849-0.954) in validation cohort, and the overall AUC for early-stage lung cancer was 0.779(95 % CI 0.738-0.816) when the two cohorts were combined.

Conclusions In summary, it would be more cost-effective and more efficient to use the model with 3 autoantibodies panel which has a high diagnostic performance for lung cancer detection, especially for early-stage lung cancer. And the model may apply a new help for the discrimination for benign pulmonary disease and lung cancer.

Note: This abstract was not presented at the meeting.

Citation Format: Di Jiang, Tingting Wang, Lu Pei, Peng Wang, Hua Ye, Chunhua Song, Kaijuan Wang, Jianying Zhang, Liping Dai. A panel of autoantibodies against multiple tumor-associated antigens in the early immunodiagnosis of lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2224.
Abstract 2228: Comparison of the Epic CTC HRD assay vs. tumor DDR mutations in metastatic castration-resistant prostate cancer (mCRPC) patients (pts)
Authors: Schonhoft Joseph , Jendrisak Adam , Lee Jerry , Rodriguez Angel , Sutton Ramsay , Wang Yipeng , Dittamore Ryan , Landers Mark
Published: August 2019

Background: PARPi have recently demonstrated clinical activity in a subset of mCRPC pts with defects in homologous recombination (HRD). Predicting PARPi response with HRD gene mutations have struggled to show clinical specificity. Previously, we described a novel circulating tumor cell (CTC) based method (CTC-HRD) that identifies a specific cell morphology associated with genomic scarring [e.g. large scale transitions (LST)] and demonstrated CTC-HRD’s clinical utility in identifying pts likely to respond to PARPi and likely to resist to standard of care (SOC) therapies (Tx), including AR signaling inhibitors. We also observed some discordance between CTC-HRD and DNA damage response (DDR) mutational status from tissue sequencing. Here we compare the prevalence of CTCs, # LSTs/CTC and the % of CTCs with BRCA2 loss in mCRPC patient samples with matched tumor sequencing DDR status.

Materials and methods: 302 CTCs from 26 mCRPC pts collected prior to Tx were collected from pts with matched tissue biopsy sequencing. CTCs were identified, assayed for CTC-HRD status, and low pass whole genome sequenced for LSTs and gene copy number alterations (CNA). We compared the prevalence of CTCs, HRD properties of each identified cell including BRCA2 copy loss, # of LSTs/CTC and their prevalence in cells derived from pts vs. CTC-HRD score and tissue DDR mutational status.

Results:

CTC-HRDDDR Tumor# PtsMedian CTC/mLMedian LST/CTCMedian % CTC BRCA2 Loss per Pt++913.83259.00%+-1010.42347.20%-+72.430.00%

CTC-HRD positive pts, independent of tissue DDR status had higher CTC/mL, higher median # LST/CTC and higher percentages of BRCA2 loss in their CTCs vs. those pts who were CTC-HRD negative. Notably, we identified 10 pts that were negative for tissue DDR mutation but were CTC-HRD+ with a high frequency of CTC BRCA2 loss.

Conclusions: These results highlight the concordance of the Epic CTC-HRD assay with known markers of HRD and demonstrate differences in CTC-HRD status from tissue DDR mutations which have struggled to achieve the required clinical specificity required for PARPi patient selection.

Citation Format: Joseph Schonhoft, Adam Jendrisak, Jerry Lee, Angel Rodriguez, Ramsay Sutton, Yipeng Wang, Ryan Dittamore, Mark Landers. Comparison of the Epic CTC HRD assay vs. tumor DDR mutations in metastatic castration-resistant prostate cancer (mCRPC) patients (pts) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2228.
Abstract 2238: Characterization of T cell clonality and tumor mutation burden utilizing the TCRSeq NGS and the comprehensive NGS assays
Authors: Gong Hua C. , Gong Hua , Papoutsis Andreas , Wu Wen , Laing Christian
Published: August 2019

Background: Somatic mutations are the basis for the generation of potential neoantigens recognized by antitumor T lymphocytes, which may contribute to increase T cell diversity [1]. Next-generation sequencing of expressed T cell receptor (TCR) transcripts allows for quantification and assessing clonality of T cell diversity [2]. Tumors that exhibit a low TCR diversity and potentially having a high clonality have been shown to be lower responders to immunotherapy treatments.

Methods: Utilizing the Shannon entropy to arrive at a clonality readout, we sought to gain mechanistic insights and demonstrate a link between clonality and total mutation load. We conducted the analysis using a diverse set of solid tumors and hematological malignancies across three different specimen types (bone marrow (BM), peripheral blood (PB), formalin-fixed paraffin embedded (FFPE tissue)). Specifically, we used the beta TCRSeq NGS and the Comprehensive NGS assays (internally developed) to characterize sample clonality and mutation load respectively.

Results: The TCRSeq NGS assay sequences the V(D)J domains of the variable region of the TCR heavy chain (involved in epitope recognition), while the hybrid capture Comprehensive NGS assay detects variants across a 236 gene panel optimized for SNV and INDEL detection. Initial analysis across the different specimen types resulted in average clonalities of BM = 0.255, PB = 0.149, FFPE = 0.092, with average tumor mutation loads of 7.6, 7.6, and 21 respectively. Categorization of all samples by increasing clonality, independent of specimen type, established a clear differentiator with samples harboring a clonality of less than 0.10 to exhibit an average mutation load of 26.33. This was statistically significant from samples harboring clonalities between 0.10 and 0.19, which had an average mutation load of 7.86, and samples with clonalities equal to or greater than 0.20, having an average mutation load of 7.67.

Conclusions: Based on two different categorical analyses, lower T cell clonality readouts appear to be associated with solid tumors harboring greater genetic diversity/neoantigen burden than hematological malignancies with higher clonalities. As tumor mutation burden has been correlated with clinical outcomes of varying efficacy, this study provides further insight into the utility of T cell clonality as a “molecular tag” for immune response and resistance in oncological therapeutics.

Citation Format: Hua C. Gong, Hua Gong, Andreas Papoutsis, Wen Wu, Christian Laing. Characterization of T cell clonality and tumor mutation burden utilizing the TCRSeq NGS and the comprehensive NGS assays [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2238.
Abstract 2241: Molecular characterization of circulating tumor cells in head and neck squamous cell carcinoma: Direct comparison of a label-independent size-based microfluidic device with EpCAM-based CTC enrichment
Authors: Zavridou Martha , Mastoraki Sophia , Strati Areti D. , Koutsodontis George , Klinakis Apostolos , Psyrri Amanda , Lianidou Evi S.
Published: August 2019

Background: Circulating tumor cells (CTCs) heterogeneity is highly affecting the efficiency of their isolation and thus the reliability of downstream analysis. Especially in Head and Neck Squamous Cell Carcinoma (HNSCC) epithelial mesenchymal transition (EMT) is highly affecting CTC isolation and downstream analysis. We directly compared two different approaches used for CTC isolation, a label-independent size-based microfluidic-based system versus an EpCAM-based positive selection for downstream molecular characterization of CTC both at the gene expression and DNA methylation level in HNSCC.

Methods: Peripheral blood (PB) in EDTA (20mL) was collected from 50 HNSCC patients and 18 healthy donors (HD). A size-based microfluidic device (Parsortix, ANGLE) and an EpCAM-based positive immune-magnetic isolation procedure were applied in parallel, using 10mL PB in each case. Total RNA was isolated from enriched CTCs and RT-qPCR was used to study the expression levels of CK-19, PD-L1, EGFR, TWIST1, CDH2 and B2M. Real time methylation specific PCR (MSP) was used to study the methylation status of SOX17, RASSF1A and MLL3 genes in DNAs isolated from the same enriched CTCs.

Results: In identical blood draws, the label-free size-based CTC-isolation system was superior in terms of sensitivity when compared to the EpCAM-based CTC enrichment, since a significantly higher percentage of identical PB samples was found positive for all genes tested both at the gene expression and DNA methylation level, while the specificity was not affected.

Conclusions: In HNSCC CTC molecular characterization at the gene expression and DNA methylation level should be based on a label-free size-based isolation system.

Citation Format: Martha Zavridou, Sophia Mastoraki, Areti D. Strati, George Koutsodontis, Apostolos Klinakis, Amanda Psyrri, Evi S. Lianidou. Molecular characterization of circulating tumor cells in head and neck squamous cell carcinoma: Direct comparison of a label-independent size-based microfluidic device with EpCAM-based CTC enrichment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2241.
Abstract 2252: YIV-906 (PHY906) enhanced the anti-tumor activity of immune checkpoint blockade therapy (Anti-PD1) against liver cancer by changing the tumor micro-environment associated with M1 macrophages infiltration
Authors: Lam Wing , Yang Xiaochen , Jiang Zaoli , han Xue , Guan Fulan , Hu Rong , Xu Chang-Hua , Cai Wei , Cheng William , Liu Shwu-Huey , Cai Yuping , Rattray Nicholas , Johnson Caroline , Chen Lieping , Cheng Yung-chi
Published: August 2019

YIV-906 (PHY906) is inspired by a traditional 1800-year Chinese herbal formulation, “Huang Qin Tang”, which is commonly used for treating diarrhea. Following chemotherapy and radiation, preclinical and clinical results suggest that YIV-906 has the potential to improve the patient’s quality of life and prolonging survival. Consistent preparations of PHY906 could be manufactured apart 15 years. The effects of YIV-906 were studied on the anti-tumor activity of anti-PD1 using BDF1 mice bearing Hepa 1-6 tumors. Results indicated that anti-PD1 alone had moderate effects on tumor growth however YIV-906 plus anti-PD1 eradicated all tumors in all tumor bearing mice. Further re-implantation of Hepa 1-6 cells did not grow in the “cured” mice, but implanted CMT167 (non-small lung carcinoma) cells or Pan02 (Pancreatic Ductal Adenocarcinoma) cells did grow; suggesting that YIV-906 plus anti-PD1 created a tumor-specific vaccine-like effect. The combination treatment exhibited a highly inflamed tumor microenvironment with more M1-like macrophage expression over M2. In culture YIV-906 could potentiate the action of IFNg (interferon gamma) to polarize bone marrow-derived macrophages (BMDM) into M1 macrophages while inhibiting IL4 action for M2 macrophage polarization. YIV-906 potentiated IFNg action through: 1) stimulating IFNg secretion, 2) phosphorylation of JAK1/2 and STAT1 and 3) increasing IRF1 protein expression. Scutellaria baicalensis Georgi (S) and its flavonoids of YIV-906 were responsible for potentiate the IFNg to polarize macrophage into M1. In conclusion, YIV-906 enhanced the anti-tumor activity of anti-PD1 by enhancing inflammation in the tumor microenvironment and enriching M1-like macrophages. This suggests the potential use of combination YIV-906 and anti-PD1 in cancer treatment. This work was supported by grant (1PO1CA154295-01A1) from National Cancer Institute (NCI), NIH, USA. Dr. Yung-Chi Cheng is a fellow of National Foundation for Cancer Research (NFCR), USA.

Citation Format: Wing Lam, Xiaochen Yang, Zaoli Jiang, Xue han, Fulan Guan, Rong Hu, Chang-Hua Xu, Wei Cai, William Cheng, Shwu-Huey Liu, Yuping Cai, Nicholas Rattray, Caroline Johnson, Lieping Chen, Yung-chi Cheng. YIV-906 (PHY906) enhanced the anti-tumor activity of immune checkpoint blockade therapy (Anti-PD1) against liver cancer by changing the tumor micro-environment associated with M1 macrophages infiltration [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2252.
Abstract 2254: Solid tumor activity with a selective PI3Kdelta inhibitor as a single agent and in combination with anti-PD-1
Authors: Methot Joey L. , Zhou Hua , McGowan Meredith A. , Trotter Benjamin W. , Altman Michael , Fradera Xavier , Lesburg Charles , Li Chaomin , Alves Stephen , Chappell Craig P. , Jain Renu , Mangado Ruban , Pinheiro Elaine , Williams Sybil M. , Goldenblatt Peter , Hill Armetta D. , Shaffer Lynsey , Chen Dapeng , McLeod Robbie L. , Lee Hyun-Hee , Shah Sanjiv , Katz Jason D.
Published: August 2019

Inhibitors of PI3Kdelta are approved as monotherapy for the treatment of hematologic malignancies such as follicular B-cell non-Hodgkin lymphoma, chronic lymphocytic leukemia, and small lymphocytic lymphoma. Recent reports suggest that inhibition of PI3Kdelta in mice is effective against solid cancers as well. Mechanistic studies by several groups have suggested a possible adaptive immune-mediated anti-tumor response involving immunosuppressive Tregs and/or MDSCs in the tumor microenvironment. Our structure-guided discovery program led to the identification of a novel heterocycloalkyl purine inhibitor class with excellent isoform and kinome selectivity, as well as good solubility. Optimization included potency enhancements, addressing metabolism and increasing half-life in preclinical species to provide a PI3Kdelta inhibitor with a low once-daily predicted human dose. In FoxP3 reporter mice harboring MC38 tumors treated with inhibitor, we observed reduced peripheral Treg function and an increased ratio of CD8 to Treg in both spleen and tumor-infiltrating lymphocytes. In three-week efficacy studies, modest tumor growth inhibition was found in 4T1, MC38 and CT26 syngeneic tumor models when our inhibitor was dosed as a single agent and we demonstrated a trend towards better efficacy when our inhibitor was dosed in combination with an anti-mPD-1 antibody in CT26 tumor-bearing mice as compared to anti-mPD-1 alone. These preliminary studies confirm the impact of PI3Kdelta inhibition on T-cell subpopulations, and support continued exploration of PI3Kdelta inhibitors for the treatment of solid tumors, either as monotherapy or in combination with other agents.

Citation Format: Joey L. Methot, Hua Zhou, Meredith A. McGowan, Benjamin W. Trotter, Michael Altman, Xavier Fradera, Charles Lesburg, Chaomin Li, Stephen Alves, Craig P. Chappell, Renu Jain, Ruban Mangado, Elaine Pinheiro, Sybil M. Williams, Peter Goldenblatt, Armetta D. Hill, Lynsey Shaffer, Dapeng Chen, Robbie L. McLeod, Hyun-Hee Lee, Sanjiv Shah, Jason D. Katz. Solid tumor activity with a selective PI3Kdelta inhibitor as a single agent and in combination with anti-PD-1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2254.
Abstract 2291: Single cell capture and molecular analysis of live CTCs using integrated microwells and single cell aspirator
Authors: Stahlfeld Charlotte N. , Tokar Jake J. , Quigley David , Niles David , Sperger Jamie M. , Feng Felix , Lang Joshua M.
Published: August 2019

Resistance to targeted therapies can be caused by acquired genomic alterations in genes that code for the targeted protein. Circulating tumor cells (CTCs) provide an accessible source of tumor cells from diverse metastatic lesions from which to evaluate the frequency of genomic alterations that can be matched to distinct phenotypes, including therapeutic resistance. Successful analysis of the diversity of molecular mechanisms of therapeutic resistance requires isolation of live tumor cells for nucleic acid extraction. We have developed a semi-automated single cell aspirating platform to enrich and isolate live, rare cell populations for downstream molecular analysis. Using immortalized cell lines, we were able to enrich a target population of cells from a mixed population resulting a high purity sample (71.7-83.8%) to interrogate. Target cells were captured for nucleic acid extraction for downstream gene expression to validate enrichment. We demonstrate the ability to achieve high quality whole genome and transcriptome endpoints from low cell numbers in cell lines: the lower threshold for cellular input required to attain whole genome amplification (WGA) products reliable for downstream sequencing applications is now being validated. We also isolated CTCs from patient blood samples and showed phenotypic heterogeneity of EpCAM expression among individual CTCs. EpCAMhigh and EpCAMlow CTCs were detected, with a broad range of fluorescent intensities (141.2-5804.56 MFI). Using the phenotypic characterization of EpCAM expression, individual live CTCs were identified and captured for evaluation of their genomic heterogeneity. We have identified molecular heterogeneity in these cells that may reflect heterogeneity driving treatment resistance in CRPC. Early identification of treatment resistant clones may help develop intervention strategies that prolong the efficacy of molecular targeted therapies.

Citation Format: Charlotte N. Stahlfeld, Jake J. Tokar, David Quigley, David Niles, Jamie M. Sperger, Felix Feng, Joshua M. Lang. Single cell capture and molecular analysis of live CTCs using integrated microwells and single cell aspirator [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2291.