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Bone metastases are common complications of solid tumors. The outcome is poor despite major progress in cancer therapies. We describe a multicenter, open-label, phase 1, dose escalation and expansion trial of JMT103, a novel fully humanized receptor activator of nuclear factor kappa-B ligand (RANKL)-targeting monoclonal antibody, in adults with bone metastases from solid tumors. The study assessed the safety, tolerability, and pharmacokinetics/pharmacodynamics of JMT103. Patients received JMT103 at doses of 0.5, 1.0, 2.0, and 3.0 mg/kg every 4 weeks for 3 cycles. Among 59 patients enrolled, 20 and 39 patients participated in the dose-escalation and dose-expansion phases, respectively. One dose-limiting toxicity was observed at 2.0 mg/kg. The maximum tolerated dose was not determined. Treatment-related adverse events were reported in 29 (49.2%) patients, most commonly hypophosphatemia (30.5%), hypocalcemia (23.7%), and hypermagnesemia (10.2%). No treatment-related serious adverse events were reported. Two patients died due to disease progression, which were attributed to gastric cancer and lung neoplasm malignant respectively. Dose proportionality occurred between exposure levels and administered dose was within a dose range of 0.5 to 3.0 mg/kg. The suppression of urinary N-telopeptide corrected for creatinine was rapid, significant, and sustained across all doses of JMT103, with the median change from baseline ranging from –61.4% to –92.2% at day 141. JMT103 was well tolerated in patients with bone metastases from solid tumors, with a manageable safety profile. Bone antiresorptive activity shows the potential of JMT103 for treatment of bone metastases from solid tumors.Registration No.: NCT03550508; URL: https://www.clinicaltrials.gov/

Our aim was to identify plasma microRNA (miRNA)-based signatures to predict 3-year postoperative recurrence risk for patients with stage II and III gastric cancer (GC), so as to provide insights for individualized adjuvant therapy. Plasma miRNA expression was investigated in three phases, involving 407 patients recruited from three centers. ABI miRNA microarray and TaqMan Low Density Array were adopted in the discovery phase to identify potential miRNAs. Quantitative reverse-transcriptase polymerase chain reaction was used to assess the expression of selected miRNAs. Logistic regression models were constructed in the training set (n = 170) and validated in the validation set (n = 169). Receiver operating characteristic analyses, survival analyses and subgroup analyses were further used to assess the accuracy of the models. We identified a 7 miRNA classifier and 7miR + pathological factors index that provided high predictive accuracy of GC recurrence (area under the curve = 0.725 and 0.841 in the training set; and 0.627 and 0.771 in the validation set). High-risk patients defined by the signatures had significantly shorter disease-free survival and overall survival than low-risk patients. The 7 miRNA classifier is an independent prognostic factor, and could add predictive value to traditional prognostic factors. Subgroup analyses revealed the satisfactory performance persisted regardless of stage, and the two models both displayed high accuracy in stage IIA patients. In conclusion, identified microRNA signature may potentially provide some additional benefit for prediction of disease recurrence in patients with stage II and III GC.

3106 Background: WX390 is a high potent PI3K-mTOR dual inhibitor targeting pan-PI3K and mTOR. WX390 exhibited anti-tumor activity and high potency in xenograft models with inhibition of AKT phosphorylation. Methods: Patients (pts) with advanced lymphoma and solid tumors who have failed standard therapies were enrolled in dose escalation cohorts and received WX390 administered daily orally in 28-day cycles until confirmed progressive disease, intolerable toxicity or withdrawal of consent. Plasma samples were collected up to 7 days after the first dose and up to 24 hours at C1D28. Results: As of the cut-off date 2021 Feb 10, 25 patients (median age 52 years) were enrolled in 6 dose levels (0.1mg, 0.2mg, 0.4mg, 0.7mg, 1.1mg and 1.4mg) of the dose-escalation cohorts. Cohorts 0.1 mg to 1.1 mg were completed without dose-limiting toxicities (DLT), cohort 1.4 mg was completed with 1 DLT (Diabetic ketoacidosis combined with grade 3 hyperglycemia) out of 4 evaluable patients. Common treatment-related adverse events (TRAEs) (all grades, ≥20%) included hyperglycemia (20 pts, 80%), proteinuria (10 pts, 40%), creatinine increased (11 pts, 44%), hypophosphatemia (10 pts, 25%), anemia (8 pts, 33.3%), thrombocytopenia (7 pts, 29.2%), and aspartate aminotransferase increased (25%). Grade≥3 TRAEs were hyperglycemia (7 pts, 28%, 1 pt combined with diabetic ketoacidosis), hypophosphatemia (2 pts, 8%), neutropenia (2 pts, 8%), dermatitis (1 pt, 4%), and maculopapular rash (1 pt, 4%). PK showed fast absorption (Tmax 0.5-4 h) and dose proportionality for Cmax and AUC0-∞. The mean multiple dose T1/2 was approximately 12.4 hours across all cohorts with minimum accumulation. Among 14 tumor response evaluable patients, there were 3 patients with confirmed PIK3CA mutation. All 3 patients experienced tumor shrinkage and were still on treatment by the cut-off date. 2 (1 pt with head and neck cancer and 1 pt with cervical cancer) of them have achieved partial response (PR). The head and neck cancer patient (PR) was treated with WX390 0.7mg/day without any severe AE and the duration of response was beyond 15 months. Stable disease (SD) was observed in 6 patients (n = 5 with unknown PIK3CA mutation status and n = 1 with confirmed PIK3CA mutation). Of note, 1 patient (follicular lymphoma) treated with 0.1mg experienced SD for 14 months. Another patient (SD, head and neck cancer with PIK3CA mutation) experienced tumor shrinkage for more than 8 months and was still receiving WX390 treatment by the cut-off date. Conclusions: the PI3K-mTOR dual inhibitor WX390 was well tolerated with manageable safety profile, and showed encouraging antitumor activity. A RP2D of 1.1 mg qd is selected to be explored for different indications in solid tumor clinical studies. Clinical trial information: NCT03730142.

Hypoxia and dysregulation of microRNAs (miRNAs) have been identified as crucial factors in carcinogenesis. However, the potential mechanisms of HIF-1α and miR-421 in gastric cancer have not been well elucidated. In this study, we found that miR-421 was up-regulated by HIF-1α. Overexpression of miR-421 promoted metastasis, inhibited apoptosis, and induced cisplatin resistance in gastric cancer in vivo and in vitro. E-cadherin and caspase-3 were identified as targets of miR-421. Besides, relative mRNA expression of miR-421 was significantly increased in gastric cancer tumor tissues compared with non-tumor tissues in a cohort of gastric cancer specimens (n=107). The expression of miR-421 was higher in advanced gastric cancers compared with localized ones. Moreover, Kaplan-Meier analysis illustrated that those patients with low levels of miR-421 had a significant longer overall survival (p = 0.006) and time to relapse (p = 0.007). Therefore, miR-421 could serve as an important prognostic marker and a potential molecular target for therapy in gastric cancer.

Cytokine-induced killer (CIK) cells represent a realistic approach in cancer immunotherapy with confirmed survival benefits in the context of metastatic solid tumors. However, therapeutic effects are limited to a fraction of patients. In this study, immune-resistance elements and ideal combination therapies were explored. Initially, phenotypic analysis was performed to document CD3, CD56, NKG2D, DNAM-1, PD-L1, PD-1, CTLA-4, TIM-3, 2B4, and LAG-3 on CIK cells. Upon engagement of CIK cells with the tumor cells, expression of PD-1 on CIK cells and PD-L1 on both cells were up-regulated. Over-expression of PD-L1 levels on tumor cells via lentiviral transduction inhibited tumoricidal activity of CIK cells, and neutralizing of PD-L1/PD-1 signaling axis could enhance their tumor-killing effect. Conversely, blockade of NKG2D, a major activating receptor of CIK cells, largely caused dysfunction of CIK cells. Functional study showed an increase of NKG2D levels along with PD-L1/PD-1 blockade in the presence of other immune effector molecule secretion. Additionally, combined therapy of CIK infusion and PD-L1/PD-1 blockade caused a delay of in vivo tumor growth and exhibited a survival advantage over untreated mice. These results provide a preclinical proof-of-concept for simultaneous PD-L1/PD-1 pathways blockade along with CIK infusion as a novel immunotherapy for unresectable cancers.

TPS1101 Background: MRG002 is an antibody drug conjugate (ADC) composed of a humanized anti-HER2 IgG1 monoclonal antibody conjugated to a microtubule disrupting agent monomethyl auristatin E (MMAE). MRG002 is presently being investigated in an ongoing phase I study for safety, tolerability, pharmacokinetics (PK), and preliminary antitumor activity in patients (pts) with solid tumors. Methods: MRG002 is evaluated as monotherapy for the treatment of pts with confirmed HER2 positive locally advanced or metastatic cancers, including breast cancer (BC), gastric cancer (GC), salivary gland cancer (SGC) and others. The primary objective is to determine the maximum tolerated dose (MTD) and a recommended phase II dose (RP2D). Secondary objectives include evaluation of PK, tumor response, and immunogenicity. In the dose escalation phase with “3+3” design, approximately 24 pts will be enrolled to identify MTD. The starting dose of MRG002 is 0.3 mg/kg, followed by 0.6, 1.2, 1.8, 2.2, 2.6, and 3.0 mg/kg. In the dose expansion phase, about 50 pts with HER2 positive advanced cancers will be enrolled to further evaluate the safety, antitumor activity, and PK at an appropriate confirmed dose. In this phase I study, each pt receives single agent MRG002 once every 3 weeks (Q3W) for a maximum of 8 treatment cycles. Pts with BC and GC should have HER2 positive advanced solid tumors per College of American Pathologists (CAP) guidelines. For other cancers, pts must have IHC status of 2+ or 3+, regardless of FISH results. Pts should have either failed or are ineligible for standard treatments. All pts must have at least 1 measurable lesion per RECIST 1.1. ECOG should be 0-1, and bone marrow, hepatic, renal, cardiac functions should be adequate. Observations include adverse events (AEs), dose-limiting toxicity (DLT), and antitumor activity, which is assessed every two treatment cycles. Further clinical trial details can be found on chinadrugtrials.org.cn (CTR20181778). Enrollment is ongoing since November 2018. Clinical trial information: CTR20181778 .

3638 Background: Bone is one of the most common metastatic sites of malignancies. The metastatic tumor cells activate osteoclast activity and promote bone resorption via RANKL/RANK signaling pathway, and lead to osteolytic destruction. JMT103 is an innovative fully human IgG4 monoclonal antibody targeting RANKL. It can block RANKL/RANK signaling pathway, inhibit bone resorption, and protect bones from tumor metastasis. This study aimed to evaluate the safety and tolerability of JMT103 in patients with bone metastasis. Methods: This is a multi-center, open label, dose escalation, phase I clinical trial. The patients (ECOG score: 0-1) with bone metastasis from solid tumor who had not received bisphosphonates within 6 weeks before enrollment and were naïve to denosumab were enrolled. The initial dose was 0.5 mg/kg, sequentially escalated to 1.0, 2.0, and 3.0 mg/kg. JMT103 was injected subcutaneously by accelerated titration in 0.5 and 1.0 mg/kg dose groups, but via traditional "3+3" dose-escalation design in 2.0 and 3.0 mg/kg dose groups. Expansion study was conducted for subjects in 1.0, 2.0, and 3.0 mg/kg dose groups. Specifically, 3 additional doses (q4w) were injected after the end of 12-week single-dose study. The primary endpoints were maximum tolerated dose and safety. The secondary outcome measures included PK profile, preliminary efficacy biomarkers, immunogenicity, and bone mineral density (BMD). Results: From May 2018 to January 2020, 56 patients (13 males, 43 females, mean (SD) age: 55.57 (11.42) years) were enrolled, including bone metastasis from breast cancer (n = 36), gastric cancer (n = 5), lung cancer (n = 4), rectal cancer (n = 3), colorectal cancer (n = 2), or other solid tumors (n = 6). Nineteen patients participated in the dose-escalating study and 37 patients participated in the expansion study. JMT103 showed overall good safety. There were 74 drug-related AEs in all, including grade 3 (DLT hypocalcemia, n = 1; hypophosphatemia, n = 3), grade 2 (n = 15), and grade 1 (n = 55) AEs. The most common drug-related AEs were hypophosphatemia (n = 15), hypocalcemia (n = 12), and hypermagnesemia (n = 6). Median uNTx/Cr decrease from baseline was 76.6% (n = 20). Conclusions: JMT103 shows good safety and tolerability in patients with bone metastasis. Clinical trial information: NCT03550508 .

CTDP1 catalyzes serine phosphorylation and dephosphorylation of the mobile carboxy-terminal domain of the RNA polymerase II. It is conserved among eukarya and is essential for cell growth for its ability in regulation of transcription machinery. However, its function in the process of tumorigenesis is unclear. In the present study, we aim to explore the roles of CTDP1 in the progression of human lung cancer. To our knowledge, this is the first study that reports the functions of CTDP1 in human lung cancer. We first detected the expression level of CTDP1 in four human lung cancer cell lines: H-125, H1299, LTEP-A-2 and NCI-H446 by semiquantitative RT-PCR. We compared the expression level of CTDP1 in lung cancer tissues and paired adjacent normal tissues on 29 pathologically confirmed patients by real-time quantitative PCR. To further explore the effect of CTDP1 on cell proliferation, a lentiviral vector expressing CTDP1 short hairpin RNA (shRNA) was constructed and infected into human lung cell lines H1299. Interference efficiency was determined by western blot analysis and real-time quantitative PCR. The effects of knockdown of CTDP1 on cell growth, cell cycle and apoptosis and cell colony formation were explored by Cellomics, fluorescence-activated cells sorting and fluorescence microscopy, respectively. CTDP1 was expressed in all four human lung cancer cell lines. The expression of CTDP1 in tumor tissues was significantly higher than paired adjacent normal tissues in 29 patients with lung cancer. The expression of CTDP1 was markedly reduced in cells infected with lentivirus delivering shRNA against CTDP1. Inhibition of CTDP1 expression significantly suppressed cell growth, induced G0/G1 phase arrest and repressed cell colony formation. Our results demonstrated that CTDP1 was upregulated in human lung cancer tissues. In addition, it implied that CTDP1 played an important role in cell proliferation and may be a useful therapeutic target in human lung cancer.

// Jinjia Chang 1,* , Shanshan Wang 1,* , Zhe Zhang 1 , Xinyang Liu 1 , Zheng Wu 1 , Ruixuan Geng 1 , Xiaoxiao Ge 1 , Congqi Dai 1 , Rujiao Liu 1 , Qunling Zhang 1 , Wenhua Li 1 and Jin Li 1 1 Department of Medical Oncology, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China * These authors contributed equally to this work Correspondence: Jin Li, email: // Keywords : drug resistance, gastric cancer, AZD4547, FGFR2 Received : September 16, 2014 Accepted : December 09, 2014 Published : December 10, 2014 Abstract Fibroblast growth factor receptor 2 (FGFR2)-targeted therapy has attracted considerable attention as novel anticancer agents in gastric cancer (GC). However, intrinsic or acquired drug resistance has emerged as a major challenge to their clinical use. In this study, we demonstrated that several receptor tyrosine kinase (RTK), including EGFR, HER3 and MET, activations contributed to AZD4547 (a selective FGFR2 inhibitor) hyposensitivity in FGFR2 amplified GC cells. The rescue effect was abrogated by inhibiting these RTKs with their targeted tyrosine kinase inhibitors (TKIs). In addition, synergy in growth inhibition was observed when the GC cells were treated with a combination of AZD4547 and cetuximab (an EGFR monoclonal antibody) both in vitro and in vivo . More importantly, tissue microarray analysis revealed that these resistance-conferring RTKs were highly expressed in FGFR2 positive GC patients. Taken together, these observations demonstrated RTKs including EGFR, HER3 and MET activations as novel mechanisms of hyposensitivity to AZD4547. It will be clinically valuable to investigate the involvement of RTK-mediated signaling in intrinsicor acquired resistance to FGFR2 TKIs in GC. A combination targeted therapeutic strategy may be recommended for treating FGFR2 amplified GC patients with these RTK activations.

Crizotinib has been reported to be particularly effective and to have acceptable toxicity in advanced anaplastic lymphoma kinase (ALK)-positive, non-small cell lung cancer (NSCLC). In this study, we analyzed the efficacy and tolerability of crizotinib in the treatment of 72 Chinese patients with ALK-positive, advanced NSCLC. All patients received oral crizotinib 250 mg twice daily in 28-day cycles during the period June 1, 2013, to October 15, 2014. The tumor response was assessed after the first cycle of crizotinib and then after every two cycles using the Response Evaluation Criteria in Solid Tumors (RECIST), version 1.0. Tolerability was assessed at least twice per cycle until crizotinib was discontinued. The patients tended to be young (mean age 55 years, range 31–83 years), never or light smokers (smoking index

Despite the promising results from initial studies, there are significant limitations in the application of MET-targeted therapy in gastric cancer. Intrinsic resistance is one of the major obstacles. The aim of this study is to identify the responsible receptor tyrosine kinases (RTKs) that determine the unresponsiveness of MET inhibitor in MET-positive gastric cancer. through an RNA-interference-based functional screen targeting most human RTKs, we identified that activation of the fibroblast growth factor receptor 2 (FGFR2) and recepteur d'origine nantais (RON) pathways attenuated MET inhibitor-induced suppression of cell proliferation and migration. Notably, in the two forms of RON pathway activation, only upregulation of short-form RON (sf-RON), but not stimulation of full length RON with macrophage stimulating protein, conferred MET inhibitor resistance in vitro and in vivo. Furthermore, the profile of the gastric cancer samples observed that sf-RON was frequently upregulated in MET-positive gastric cancer. Our findings indicate that activation of the sf-RON signaling pathway represents a novel mechanism underlying MET inhibitor unresponsiveness. A combination strategy with drugs targeting both RON and MET pathways is believed to improve the efficacy of MET-targeted therapy.

Growing evidence indicates that microRNA (miRNA) plays a vital role in progression and metastasis of gastric cancer (GC). However, the underlying mechanism of miRNA-mediated metastasis has not been fully understood. Recently, miRNA-940 (miR-940) was found to be overexpressed in GC, which correlated with malignant progression and poor survival. Mechanistically, we found that miR-940 promoted GC cell migration, invasion, and metastasis in vivo by directly and functionally repressing the expression of Zinc Finger Transcription Factor 24 (ZNF24). Importantly, upregulation of ZNF24 could re-inhibit miR-940-induced migration and invasion. Hence, we demonstrated the oncogenic role of miR-940 in GC, finding that miR-940 promoted GC progression by directly downregulating ZNF24 expression, and targeting miR-940 could serve as a novel strategy for future GC therapy.

Small cell lung cancer (SCLC) is an aggressive disease with poor survival. A few sequencing studies performed on limited number of samples have revealed potential disease-driving genes in SCLC, however, much still remains unknown, particularly in the Asian patient population. Here we conducted whole exome sequencing (WES) and transcriptomic sequencing of primary tumors from 99 Chinese SCLC patients. Dysregulation of tumor suppressor genes TP53 and RB1 was observed in 82% and 62% of SCLC patients, respectively, and more than half of the SCLC patients (62%) harbored TP53 and RB1 mutation and/or copy number loss. Additionally, Serine/Arginine Splicing Factor 1 (SRSF1) DNA copy number gain and mRNA over-expression was strongly associated with poor survival using both discovery and validation patient cohorts. Functional studies in vitro and in vivo demonstrate that SRSF1 is important for tumorigenicity of SCLC and may play a key role in DNA repair and chemo-sensitivity. These results strongly support SRSF1 as a prognostic biomarker in SCLC and provide a rationale for personalized therapy in SCLC., Author Summary SCLC patients are initially highly chemo-sensitive with response rates of greater than 80% in both limited and extensive diseases, but suffer uniform disease recurrence or progression in a very short period of time. In the absence of well-defined genomic biomarkers and insights into the resistance mechanism, many targeted treatments have yielded negative results in the last decade Using integrated next generation sequencing (NGS) technology in combination with a high quality surgical sample set with comprehensive clinical annotation, our study not only identified novel recurrent genetic alterations in genes such as CDH10 and DNA repair pathways which may influence outcomes in SCLC patients, but also discovered the expression of SRSF1, an RNA-splicing factor which can both regulate key oncogenic and survival pathways such as BCL2, and play a critical role in patient survival.

Small cell lung cancer (SCLC) is an aggressive disease with poor survival. Large-scale sequencing studies have revealed potential disease-driving genes in various cancers, although in SCLC, much still remains unknown, particularly in the Asian patient population. Here we conducted whole exome sequencing (WES) and transcriptomic sequencing of primary tumors from 99 Chinese SCLC patients. It revealed 34,099 somatic nonsilent single nucleotide variants (SNVs) or insertion/deletions (indels), an average of 308 per patient and nonsilent/silent ratio of 1.66. Genes harboring the most recurrent SNVs or indels were TP53 (81%), CSMD3 (43%), RB1 (46%), LRP1B (38%) and OBSCN (41%). Somatic copy number variants (CNVs) were also identified including MYC (8%), KIT (17%), and SOX4 (19%), with SOX2 (67%) and multiple other genes located across a segment on chromosome 3q27.1. Genes with CN losses include RB1 (34%), RASSF1 (57%), FHIT (54%), KIF2A (16%), and CNTN3 (53%), as well as a long segment along chromosome 3p22.1. Recurrence rates of these genes affected by SNVs and CNVs were comparable to those reported previously. In addition, we found recurrent gains of Serine/Arginine Splicing Factor 1 (SRSF1) in 51% (49/96) patients as well as concordant over-expression of mRNA for those patients with gains (p = 0.005; two-tailed two-sided Welch's t-test;). Among these 96 Chinese patients, 28% had both CN gain and mRNA over-expression of SRSF1; SRSF1 CN status was evaluated by FISH assay (N = 34), the positive and negative predictive values were 57% and 69% respectively. Patients with both RNASeq and survival data were interrogated (N = 49), SRSF1 was the only gene correlating between both CN gain and mRNA over-expression as well as between over-expression and survival using a Cox proportion hazard (PH) regression model adjusting for age, gender, tumor stage, and chemotherapy status (p = 0.034; HR = 3.0). Patients with SRSF1 mRNA over-expression or CN gain demonstrated significantly worse survival. The discovery (log-rank test p = 0.062), validation (log-rank test p = 0.03), and combined patient cohort (Cox PH p = 0.021; HR = 2.1; log-rank test p = 0.005). On target in vivo studies demonstrated that SRSF1 is essential for tumorigenecity of SCLC. siRNAs (control/SRSF1)were transfected to cell lines with DNA amplification/mRNA overexpression (DMS114 and SHP-77SRSF1). Equal numbers of viable transfected cells were injected in immunocompromised mice and tumor growth was monitored for up to three weeks. SRSF1 knockdown completely suppressed the tumor growth These data strongly support SRSF1 as a therapeutic target in SCLC and provide a rationale for personalized therapy in SCLC. Citation Format: Zheng Liu, Jiaqi Huang, Brandon W. Higgs, Haihong Zhong, Dong Shen, Zhan Xiao, Xin Yao, Philip Brohawn, Xiaoxiao Ge, Zhibing Hu, Yue Jiang, Chris Morehouse, Wei Zhu, Yinong Sebastian, Meggan Czapiga, Vaheh Oganesyan, Haihua. Fu, Xinying Su, Yi Gu, Baohui Han, Ronald Herbst, Liayang Jiang, Hongbing Shen, Yihong Yao. Genomic landscape survey identifies SRSF1 as a key oncodriver in small cell lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2032. doi:10.1158/1538-7445.AM2015-2032

Small cell lung cancer (SCLC) is the most aggressive subtype of lung cancer. Despite a high response rate to chemotherapy, more than 95% of patients eventually die from SCLC. We have identified that Serine/Arginine Splicing Factor 1 (SRSF1) DNA copy number gain and mRNA over-expression in tumor is strongly associated with poor survival based on whole exome sequencing (WES) and transcriptomic sequencing of primary tumors from 99 Chinese SCLC patients. Here, SRSF1 is evaluated as a tumor driver in SCLC. Treatment of SCLC cell lines in vitro with a low dose of cisplatin or topotecan (two of the most common standard of care in SCLC) only induced a modest decrease of cell growth. However, knockdown of SRSF1 with siRNA along with a sub-lethal dose of cisplatin or topotecan enhanced the overall growth inhibition effect compared to the chemotherapy alone. SRSF1 siRNA alone induced modest but significant caspase-3 activation, similar to cisplatin or topotecan treatment alone. The combination of SRSF1 siRNA with chemotherapy treatments produced a significantly higher caspase induction. DNA-damage induction as a potential mechanism of SRSF1 knockdown was assessed. Phosphorylation of H2AX and Chk2, established markers of DNA-strand breaks and DNA-repair response, was consistently induced upon SRSF1 abrogation in cells, and further increased the phosphorylation of these proteins when combined with cisplatin or topotecan treatment. The knockdown of SRSF1 in SCLC cells also resulted in significant growth inhibition when cells were grown as 3D spheroids. Cells transfected with non-targeting siRNA produced large and well-organized spheroids; in contrast, cells transfected with SRSF1 siRNA did not form well-organized structures but mainly existed as single cells with poor viability. Results were confirmed by colony formation assays and could be rescued with a siRNA-resistant SRSF1 expression construct. Finally, we investigated the impact of SRSF1 loss on kinase signaling pathways in SCLC cells through phospho-kinase array profiling. Strong phospho-AKT and ERK signals were observed in control siRNA-transfected cells, and were abrogated by SRSF1 siRNA. Western blot confirmed this in several SCLC cell lines. These targeting studies demonstrate that SRSF1 plays a key role in DNA repair, chemo-sensitivity and cell proliferation. Together, these data reveal SRSF1 as a therapeutic target in SCLC and provide a rationale for personalized therapy in SCLC. Citation Format: Sarah J. Conley, Xin Yao, Jiaqi Huang, Brandon Higgs, Zhibin Hu, Zhan Xiao, Haihong Zhong, Zheng Liu, Philip Brohawn, Xiaoxiao Ge, Meggan Czapiga, Vaheh Oganesyan, Haihua Fu, David Tice, Ronald Herbst, Xinying Su, Yi Gu, Jianren Gu, Baohui Han, Laura Richman, Bahija Jallal, Liyan Jiang, Hongbing Shen, Yihong Yao. Serine/arginine splicing factor 1 (SRSF1) mediates DNA repair and chemo-sensitivity and drives growth in small cell lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4954. doi:10.1158/1538-7445.AM2015-4954

To evaluate the insulin receptor isoform mRNA expression status in NSCLC patients RNA-seq data from 614 NSCLC [355 adenocarcinomas(LUAD) and 259 squamous cell carcinomas (LUSC)] and 92 normal lung specimens were obtained from The Cancer Genome Atlas (TCGA) to evaluate the mRNAexpression of insulin receptor isoform A (IR-A) and insulin receptor isoform B(IR-B). The differential expression status of the insulin receptor isoforms in NSCLC patients was confirmed using qRT-PCR assays with lung cancer cDNA arrays and primary tumor samples. The mRNA expression levels of IR-B were significantly lower in some NSCLC samples compared to normal lung specimens,including both LUAD and LUSC. Notably, no IR-B transcripts were detected - only the IR-A isoform was expressed in 11% of NSCLC patients. This decrease in IR-B expression contributed to an elevated IR-A/IR-B ratio, which was also associated with lower epithelial-mesenchymal transition gene signatures in NSCLC and longer patient survival under standard of care in LUSC. In addition to NSCLC, RNA-seq data from TCGA revealed a similar increase in IR-A/IR-B ratio in many other cancer types, with high prevalence in acute myeloid leukemia,glioblastoma multiforme, and brain lower grade glioma. Our results indicate a common reduction of the mRNA expression level of IR-B and an increased IR-A/IR-B mRNA ratio in NSCLC and other tumor types. The relationship of altered IR-A/IR-B ratios with cancer progression and patient survival should be prospectively explored in future studies. Citation Format: Jiaqi Huang, Wei Zhu, Liyan Jiang, Katie Streicher, Chris Morehouse, Philip Brohawn, XiaoXiao Ge, Zhengwei Dong, Xiaolu Yin, Guanshan Zhu, Yi Gu, Parthiv Mahadevia, Brandon W. Higgs, Yihong Yao. Increased IR-A/IR-B ratio in non-small cell lung cancers associates with lower epithelial-mesenchymal transition signature and longer survival in squamous cell lung carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4909. doi:10.1158/1538-7445.AM2015-4909

Intratumor heterogeneity can confound the results of mutation analyses in oncodriver genes using traditional methods thereby challenging the application of targeted cancer therapy strategies for patients. Ultradeep sequencing can detect low frequency and expanded clonal mutations in primary tumors to better inform treatment decisions. KRAS coding exons in 61 treatment-naïve colorectal cancer (CRC) tumors and KRAS, EGFR, ALK, and MET in lung tumors from three Chinese non-small cell lung cancer (NSCLC) patients were sequenced using ultradeep sequencing methods. Forty-one percent of CRC patients (25/61) harbored mutations in the KRAS active domain, eight of which (13%) were not detected by Sanger sequencing. Three (of eight) had frequencies less than 10% and one patient harbored more than one mutation. Low frequency KRAS active (G12R) and EGFR kinase domain mutations (G719A) were identified in one NSCLC patient. A second NSCLC patient showed an EML4-ALK fusion with ALK, EGFR, and MET mutations. A third NSCLC patient harbored multiple low frequency mutations in KRAS, EGFR, and MET as well as ALK gene copy number increases. Within the same patient, multiple low frequency mutations occurred within a gene. A complex pattern of intrinsic low frequency driver mutations in well-known tumor oncogenes may exist prior to treatment, resulting in resistance to targeted therapies. Current targeted cancer therapies usually lack durability and demonstrate limited overall efficacy in patients. The types of low frequency concurrent mutations in candidate oncogenes presented here suggest necessary modifications both to methods for detection of these variants and to general treatment strategies. To date, Sanger sequencing has been effectively used for detection of treatment-relevant somatic mutations. However, in a heterogeneous mixture of cancerous and normal tissue, Sanger sequencing will likely fail to detect low frequency mutations. More sensitive and cost-effective sequencing methods are required to systematically assess the mutation status within cancer pathway genes or at the whole genome level. Furthermore, because patients often develop resistance to targeted therapy over time that is due to the preexistence of low frequency mutations in oncogenes, treatment strategies based on combination therapy might prove to be the most optimal treatment approach for cancer patients. Ultradeep sequencing can characterize intratumor heterogeneity and identify such mutations to ultimately affect treatment decisions. Citation Format: Christopher Morehouse, Liyan Jiang, Jiagi Huang, Wei Zhu, Susana Korolevich, Xiaoxiao Ge, Kim Lehmann, Zheng Lui, Christine Kiefer, Meggan Czapiga, Xinying Su, Philip Brohawn, Yi Gu, Brandon Higgs, Yihong Yao. Low frequency KRAS mutations in colorectal cancer patients and the presence of multiple mutations in oncogenic drivers in non-small cell lung cancer patients. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2377. doi:10.1158/1538-7445.AM2014-2377

Introduction: Salivary gland–type cancers are rare lung neoplasms involving mucoepidermoid carcinoma (MEC), adenoid cystic carcinoma (ACC), and epithelial–myoepithelial carcinoma (EMC). Their behavior and prognostic features are not clearly defined because of their low incidence. We retrospectively analyzed the clinicopathologic profiles of these tumors in a large series. Methods: Eighty-eight patients confirmed as having primary salivary gland–type lung cancer between May 2001 and January 2013 were included from the archives of two thoracic oncology center institutions in China and retrospectively evaluated. Results: Of the total 88 patients, 69 were MEC, 12 ACC, and seven EMC. Overall survival (OS) at 3, 5, and 10 years was 91.3%, 86%, and 80.6% in all cases, respectively, and disease-free survival (DFS) was 90.1%, 78.6%, and 55%, respectively. No significant difference was found among MEC, ACC, and EMC groups regarding OS (p = 0.518) and DFS (p = 0.082). Tumor-node-metastasis stage, lymph node involvement, intrathoracic invasion, and margin status were found to be related with OS (p = 0.000, 0.029, 0.000, 0.004) and DFS (p = 0.018, 0.042, 0.002, 0.002). Intrathoracic invasion was an independent predictor for OS (hazard ratio [HR], 1.129; p = 0.039) and DFS (HR, 1.071; p = 0.011). For patients with MEC, pathological grade also was an independent predictor of OS (HR, 0.045; p = 0.006) and DFS (HR, 0.067; p = 0.001). Conclusions: Salivary gland–type lung cancers are a group of low-aggressive entities with higher tendency to recurrence/metastasis. Intensive clinical, radiological, and pathological examinations are essential to estimation of the risk stratification and management.

Background To evaluate the insulin receptor isoform mRNA expression status in non-small cell lung cancer (NSCLC) patients. Methods RNA-seq data from 614 NSCLC [355 adenocarcinomas (LUAD) and 259 squamous cell carcinomas (LUSC)] and 92 normal lung specimens were obtained from The Cancer Genome Atlas (TCGA) to evaluate the mRNA expression of insulin receptor isoform A (IR-A) and insulin receptor isoform B (IR-B). The differential expression status of the insulin receptor isoforms in NSCLC patients was confirmed using qRT-PCR assays with lung cancer cDNA arrays and primary tumor samples. Results The mRNA expression levels of IR-B were significantly lower in some NSCLC samples compared to normal lung specimens, including both LUAD and LUSC. Notably, no IR-B transcripts were detected - only the IR-A isoform was expressed in 11% of NSCLC patients. This decrease in IR-B expression contributed to an elevated IR-A/IR-B ratio, which was also associated with lower epithelial-mesenchymal transition gene signatures in NSCLC and longer patient survival under standard of care in LUSC. In addition to NSCLC, RNA-seq data from TCGA revealed a similar increase in IR-A/IR-B ratio in many other cancer types, with high prevalence in acute myeloid leukemia, glioblastoma multiforme, and brain lower grade glioma. Conclusions Our results indicate a common reduction of the mRNA expression level of IR-B and an increased IR-A/IR-B mRNA ratio in NSCLC and other tumor types. The relationship of altered IR-A/IR-B ratios with cancer progression and patient survival should be prospectively explored in future studies.