Citations

Abstract Lung cancer is the most lethal malignancy around the world. In Hong Kong, chronic tobacco use is prevalent in male but <20% of female patients smoke. Epidermal growth factor receptor (EGFR) mutations affect 30% and 67% of male and female lung adenocarcinomas, respectively. Treatment responses to tyrosine kinase inhibitors (TKI) are not uniform and all patients eventually develop drug resistance. Mutation screening of immediate pre-treatment tumors is the most ideal for assessing drug targets but in practice, excision specimens of primary or metastatic sites are more efficient in terms of tumor quantity, timeliness and patient acceptance. However, information on the applicability of this approach and correlation with the clinical outcome is limited. For this purpose, we compared the whole exome mutation profiles of 39 excised EGFR mutant cancers treated with first line TKI with the clinical response type, including 15 stage I/II and 24 stage III/IV tumors. They included 16 non-responders (NR) defined clinically as those with stable or progressive disease, and 23 responders (R) defined by partial or complete tumor shrinkage. Considering only the non-synonymous SNV and INDEL mutations of coding regions of known actionable targets and genes listed in both the CGC and COSMIC cancer databases, the tumors harbored 54 mutated genes including 26 recurrently mutated and 28 genes involving only 1 tumor. Excluding EGFR, TP53 was the most commonly mutated gene occurring in 25/39 (64.1%) cases. Most known resistant genes involved in the EGFR and bypass signaling network showed mutations only in the NR group including recurrent mutations of PTEN (3/16), PIK3CA (2/16) and NF1 (2/16), and single case mutations of AKT1, ALK, RAF1 and KDR. Two EGFR network candidates showed mutations in both the NR and R groups, including HGF (2/16 NR, 1/23 R) and ROS1 (1 case in either group). Four post-treatment (post-TKI) tumors of acquired resistance were also analyzed, all of which showed EGFR T790M while no pre-treatment NR or R tumor harbored this mutation. Notably, mutations in the β-catenin pathway were prominent in the NR and post-TKI tumors, including APC (2/16 NR), CTNNB1 (1/16 NR, 1/4 post-TKI) and c-MYC (1/16 NR, 1/4 post-TKI) while they were not detected in the R tumors. Also, nonsense mutations of ARID1A was observed in 2/16 NR but none of the R tumors. In summary, our findings revealed candidate TKI resistant mutations involving the EGFR and bypass signaling networks in pre-treatment excision specimens particularly in non-responding patients. The EGFR T790M was not detectable in pre-treatment samples but was prevalent in post-TKI treated cancers. While this study is limited by its small cohort size, the findings indicate deep sequencing analysis of pre-treatment excision specimens of EGFR-mutant lung adenocarcinomas is warranted for detection of resistant mutations and predicting treatment response. Note: This abstract was not presented at the meeting. Citation Format: Xu-yuan Gao, Hang Xu, James CM Ho, Oscar SH Chan, Feng Xu, Junwen Wang, Victor HF Lee, Vicky PC Tin, Zhijie Xiao, Siqi Wang, Judy WP Yam, Maria P. Wong. Whole-exome sequencing reveals tyrosine kinase-resistant mutations in pretreatment EGFR-mutant lung adenocarcinomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-274. doi:10.1158/1538-7445.AM2017-LB-274

Abstract Oncogenic role of p21-activated protein 4 (PAK4) in relation to cell growth and migration through various signalling cascades has been well studied in multiple cancer types. Such role might be contributed largely by cytoplasmic PAK4 as most of the known PAK4 signalling events occur in the cytoplasm. However, recent studies have demonstrated that PAK4 is capable of shuttling between nucleus and cytoplasm. The precise function of nuclear PAK4 (nPAK4) remains substantially unexplored. We have previously shown that PAK4 can act as a transcription factor. Herein, we aim to identify nPAK4-regulated genes by delineating the PAK4-binding landscape in ovarian cancer genome and mapping the PAK4 transcriptome in ovarian cancer cells. PAK4-enriched genomic regions were identified by chromatin immunoprecipitation with a PAK4 antibody followed by next-generation sequencing (ChIP-seq) in OVCAR3 cells. PAK4 transcriptome data was generated using RNA extracted from SKOV3 cells stably expressing PAK4 in the Affymetrix GeneChIP® Human Transcriptome Array 2.0 system. Gene lists generated from both ChIP-seq and microarray dataset were subjected to pathway enrichment analysis by Partek® PathwayTM. Among the 20 most significantly enriched pathways from microarray studies, several pathways in which PAK4 has no known reported function by far have been identified. Notably, DNA repair pathways that are known to influence drug chemosensitivity in ovarian cancer, such as the nucleotide excision repair (p=0.008), homologous recombination (p=0.012), mismatch repair (p=0.018) and DNA replication (p=0.018) pathways were significantly enriched. In concordance with the findings, the nucleotide excision repair pathway was also significantly enriched (p<0.006) in the gene list derived from promoter-restricted PAK4 binding sites in the ovarian cancer genome. Among the many candidates that were deregulated by PAK4, cyclin-dependent kinase 7 (CDK7), a core component in the CAK subcomplex crucial to nucleotide excision repair pathway, was further validated. The transcription start site (TSS) of cyclin-dependent kinase 7 (CDK7) was bound by PAK4. CDK7 mRNA expression was upregulated upon PAK4 overexpression. Quantitative-PCR showed downregulated CDK7 mRNA expression upon siRNA-mediated knockdown of PAK4 in OVCAR3 and TUOS3. This study points to the possible role of PAK4 in regulating gene transcription as PAK4 is mapped to several core promoter/TSS regions in the ovarian cancer genome. Our findings also shed light into the potential role of PAK4 in regulating drug chemosensitivity via its transcriptional control on genes implicated in DNA repair pathways. We postulate CDK7 as a direct transcriptional target of PAK4 and thereby contributes to chemoresistance in ovarian cancer cells. Citation Format: Ivy Tsz-Lo Wong, Oscar Gee-Wan Wong, Yiming Qin, Junwen Wang, Annie Nga-Yin Cheung. Genome-wide profiling of PAK4 DNA-binding sites and transcriptome reveals its potential transcriptional control on DNA repair-related genes in ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2419. doi:10.1158/1538-7445.AM2017-2419

AbstractRenal agenesis is a devastating birth defect, and although genes encoding retinoic acid signaling components have been shown to be important for renal...Renal agenesis (RA) is one of the more extreme examples of congenital anomalies of the kidney and urinary tract (CAKUT). Bilateral renal agenesis is almost invariably fatal at birth, and unilateral renal agenesis can lead to future health issues including end-stage renal disease. Genetic investigations have identified several gene variants that cause RA, including EYA1, LHX1, and WT1. However, whereas compound null mutations of genes encoding α and γ retinoic acid receptors (RARs) cause RA in mice, to date there have been no reports of variants in RAR genes causing RA in humans. In this study, we carried out whole exome sequence analysis of two families showing inheritance of an RA phenotype, and in both identified a single candidate gene, GREB1L. Analysis of a zebrafish greb1l loss-of-function mutant revealed defects in the pronephric kidney just prior to death, and F0 CRISPR/Cas9 mutagenesis of Greb1l in the mouse revealed kidney agenesis phenotypes, implicating Greb1l in this disorder. GREB1L resides in a chromatin complex with RAR members, and our data implicate GREB1L as a coactivator for RARs. This study is the first to associate a component of the RAR pathway with renal agenesis in humans.

Tumor-initiating cells (TIC) are dynamic cancer cell subsets that display enhanced tumor functions and resilience to treatment but the mechanism of TIC induction or maintenance in lung cancer is not fully understood. In this study, we show the calcium pathway transcription factor NFATc2 is a novel regulator of lung TIC phenotypes, including tumorspheres, cell motility, tumorigenesis, as well as in vitro and in vivo responses to chemotherapy and targeted therapy. In human lung cancers, high NFATc2 expression predicted poor tumor differentiation, adverse recurrence-free and cancer-specific overall survivals. Mechanistic investigations identified NFATc2 response elements in the 3’ enhancer region of SOX2, and NFATc2/SOX2 coupling upregulates ALDH1A1 by binding to its 5’ enhancer. Through this axis, oxidative stress induced by cancer drug treatment is attenuated, leading to increased resistance in a mutation-independent manner. Targeting this axis provides a novel approach for the long-term treatment of lung cancer through TIC elimination.

Significance Deposition of amyloid-β (Aβ) peptide in the form of parenchymal plaques and Aβ accumulation in the walls of cerebral vessels as cerebral amyloid angiopathy (CAA) are pathological hallmarks of Alzheimer’s disease (AD). The clusterin ( CLU ) gene, which confers AD risk, is associated with amyloid deposition. Here we show that loss of CLU promotes cerebrovascular CAA, yet significantly reduces the amount of parenchymal plaques by altering perivascular drainage of Aβ in the APP/PS1 mouse model of AD. The absence of CLU in these mice is associated with a lower number of hemorrhages and a decrease in inflammation. These results suggest that CLU functions as a major Aβ chaperone to maintain Aβ solubility along interstitial fluid drainage pathways and prevent CAA formation.

Background: Serious adverse events (SAEs) and subject replacements occur frequently in phase 1 oncology clinical trials. Whether baseline quality-of-life (QOL) or social support can predict risk for SAEs or subject replacement among these patients is not known. Methods: Between 2011–2013, 92 patients undergoing screening for enrollment into one of 22 phase 1 solid tumor clinical trials at Roswell Park Cancer Institute were included in this study. QOL Questionnaires (EORTC QLQ-C30 and FACT-G), Medical Outcomes Study Social Support Survey (MOSSSS), Charlson comorbidity scores (CCS) and Royal Marsden scores (RMS) were obtained at baseline. Frequency of dose limiting toxicities (DLTs), subject replacement and SAEs that occurred within the first 4 cycles of treatment were recorded. Fisher’s exact test and Mann-Whitney-Wilcoxon test were used to study the association between categorical and continuous variables, respectively. A linear transformation was used to standardize QOL scores. p-value ≤ 0.05 was considered statistically significant. Results: Baseline QOL, MOSSSS, CCS and RMS were not associated with subject replacement nor DLTs. Baseline EORTC QLQ-C30 scores were significantly lower among patients who encountered SAEs within the first 4 cycles (p = 0.04). Conclusions: Lower (worse) EORTC QLQ-C30 score at baseline is associated with SAE occurrence during phase 1 oncology trials.

The failure of the polycystins (PCs) to function in primary cilia is thought to be responsible for autosomal dominant polycystic kidney disease (ADPKD). Primary cilia integrate multiple cellular signaling pathways, including calcium, cAMP, Wnt, and Hedgehog, which control cell proliferation and differentiation. It has been proposed that mutated PCs result in reduced intracellular calcium, which in turn upregulates cAMP, protein kinase A (PKA) signaling, and subsequently other proliferative signaling pathways. However, the role of PKA in ADPKD has not been directly ascertained in vivo, although the expression of the main regulatory subunit of PKA in cilia and other compartments (PKA-RIα, encoded by PRKAR1A) is increased in a mouse model orthologous to ADPKD. Therefore, we generated a kidney-specific knockout of Prkar1a to examine the consequences of constitutive upregulation of PKA on wild-type and Pkd1 hypomorphic ( Pkd1 RC ) backgrounds. Kidney-specific loss of Prkar1a induced renal cystic disease and markedly aggravated cystogenesis in the Pkd1 RC models. In both settings, it was accompanied by upregulation of Src, Ras, MAPK/ERK, mTOR, CREB, STAT3, Pax2 and Wnt signaling. On the other hand, Gli3 repressor activity was enhanced, possibly contributing to hydronephrosis and impaired glomerulogenesis in some animals. To assess the relevance of these observations in humans we looked for and found evidence for kidney and liver cystic phenotypes in the Carney complex, a tumoral syndrome caused by mutations in PRKAR1A. These observations expand our understanding of the pathogenesis of ADPKD and demonstrate the importance of PRKAR1A highlighting PKA as a therapeutic target in ADPKD.