Citations

AbstractMAPK pathway activation has been recurrently observed in desmoplastic infantile ganglioglioma/astrocytoma (DIG/DIA) with reported disproportionally low mutation allele frequencies relative to the apparent high tumor content, suggesting that MAPK pathway alterations may be subclonal. We sought to expand the number of molecularly profiled cases and investigate if tumor cell composition could account for the observed low mutation allele frequencies. Molecular (targeted neuro-oncology next-generation sequencing/RNA sequencing and OncoScan microarray) and immunohistochemical (CD68-PGM1/CD163/CD14/CD11c/lysozyme/CD3/CD20/CD34/PD-L1) studies were performed in 7 DIG. Activating MAPK pathway alterations were identified in 4 (57%) cases: 3 had a BRAF mutation (V600E/V600D/V600_W604delinsDQTDG, at 8%–27% variant allele frequency) and 1 showed a TPM3-NTRK1 fusion. Copy number changes were infrequent and nonrecurrent. All tumors had at least 30% of cells morphologically and immunophenotypically consistent with microglial/macrophage lineage. Two subtotally resected tumors regrew; 1 was re-excised and received adjuvant treatment (chemotherapy/targeted therapy), with clinical response to targeted therapy only. Even with residual tumor, all patients are alive (median follow-up, 83 months; 19–139). This study further supports DIG as another MAPK pathway-driven neuroepithelial tumor, thus expanding potential treatment options for tumors not amenable to surgical cure, and suggests that DIG is a microglia/macrophage-rich neuroepithelial tumor with frequent low driver mutation allele frequencies.

AbstractPrimary aneurysmal bone cyst (ABC) is a benign multiloculated cystic lesion of bone that is defined cytogenetically by USP6 gene rearrangements. Rearrangements involving USP6 are promoter swaps, usually generated by fusion of the noncoding upstream exons of different partner genes with exon 1 or 2 of USP6, thus leading to transcriptional upregulation of full‐length USP6 coding sequence. Testing for USP6 rearrangements is used diagnostically to distinguish it from secondary ABC and other giant cell‐rich primary bone tumors. In this report, we present a case of a 16‐year‐old male with a primary ABC of the left distal femur. USP6 break apart fluorescence in situ hybridization was positive for a rearrangement and conventional chromosome analysis identified a reciprocal X;17 translocation. In order to identify the putative USP6 fusion partner, we performed RNA sequencing and uncovered a novel USP9X‐USP6 promoter swap fusion. This result was confirmed by reverse transcription‐polymerase chain reaction (RT‐PCR) and by mate pair sequencing thus showing the utility of these alternative methodologies in identifying novel fusion candidates. Ubiquitin‐specific protease 9X (USP9X), like USP6, encodes a highly conserved substrate‐specific deubiquitylating enzyme. USP9X is highly expressed in a number of tissue types and acts as both an oncogene and tumor suppressor in several human cancers. We conclude that oncogenic activation of USP6 via USP9X promoter exchange represents a novel driver of primary ABC formation.

Abstract In lung adenocarcinoma, gene fusion involving the anaplastic lymphoma kinase (ALK) gene leads to overexpression of its tyrosine kinase domain, which serves as the oncogenic driver in lung tumors carrying such a molecular alteration. Similar fusions involving ROS1, RET and NTRK are also found in lung cancer and tumors with oncogenic fusions often respond well to targeted therapy. However, these tumors do develop resistance to the targeting drug and patients eventually relapse while on treatment. Although newer drugs have been developed, it remains a challenge to effectively control cancer recurrence after targeted therapy. In vivo tumor models carrying these fusions, particularly those that have progressed while on targeted treatment, will help to facilitate further drug evaluations prior to their administration to the patient as well as offer an opportunity to better understand the molecular mechanisms driving disease onset, progression, and drug resistance in tumors carrying the fusion of interest. Here, we describe the first patient-derived tumor xenograft (PDX) carrying an STRN-ALK fusion. The patient was diagnosed with stage IV lung adenocarcinoma in 2011. She was initially treated with crizotinib and remained stable until 2015. At the time of disease progression, a biopsy was taken with patient consent under CT guidance from a chest wall mass using an 18-gauge needle and then implanted into two SCID-beige mice subcutaneously and into the fat pad of the kidney. Palpable tumor was observed in both mice at about 28 weeks post implantation and passaged at 36 weeks. They have been passaged two successive times and viably frozen. Pathologic analysis of the PDX showed that both tumors have the same mucinous morphology as observed in the original biopsy sample. RNA-seq and matepair analysis revealed that the tumors carry a rare fusion involving the first three exons of STRN and exon 20 to 3’ end of the ALK gene. The patient has been on ceritinib since January 2016 and remains stable as of last follow up. Several unique features contributed to the success of this study. 1) The metastatic chest wall lesion used for the PDX developed was easily accessible on the chest wall using a relative large 18-gauge needle for CT-guided biopsies. 2) The slow growth of the tumor biopsy mirrors the clinical course of the disease observed in the patient. 3) The patient remains clinically stable and continues to respond to TKi. The availability of a viable PDX tumor model for this rare STRN-ALK fusion offers a unique opportunity to identify the most appropriate drug for the patient at the time of disease progression as well as the opportunity to examine the unique molecular features associated with this rare ALK fusion. This work was supported in part by the National Foundation for Cancer Research Hillsberg Lung Cancer Translational Research Grant and by the Biomarker Discovery Program at the Mayo Clinic Center for Individualized Medicine. Citation Format: HongZheng Res, XiaoNan Hou, Karlyn E. Pierson, Patric Eiken, Asha Nair, PingYang Yang, Joanne Eunhee Yi, Aaron S. Mansfield, Jin Jen. Establishment of a unique patient-derived tumor model positive for STRN-ALK fusion from a patient with stage IV lung adenocarcinoma [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr A17.

AimsRhabdomyosarcomas of bone are extremely rare, with fewer than 10 reported cases. A very rare subtype of spindle cell/sclerosing rhabdomyosarcoma harbouring a FUS–TFCP2 fusion and involving both soft tissue and bone locations has been reported very recently. We report only the fourth case of this unusual, clinically aggressive rhabdomyosarcoma.Material and resultsA previously well 72‐year‐old male presented with a destructive lesion of the mandible. Morphological and immunohistochemical study of a needle biopsy and the subsequent resection showed a spindle cell rhabdomyosarcoma. RNA‐seq, RT–PCR and FISH confirmed the presence of the FUS–TFCP2 fusion.ConclusionsSpindle cell rhabdomyosarcomas carrying the FUS–TFCP2 fusion are very rare rhabdomyosarcoma variants with osseous predilection. The classification and differential diagnosis of this unusual molecular variant of spindle cell/sclerosing rhabdomyosarcoma are discussed.

Abstract Human immunity exhibits remarkable heterogeneity among individuals, which engenders variable responses to immune perturbations in human populations. Population studies reveal that, in addition to interindividual heterogeneity, systemic immune signatures display longitudinal stability within individuals, and these signatures may reliably dictate how given individuals respond to immune perturbations. We hypothesize that analyzing relationships among these signatures at the population level may uncover baseline immune phenotypes that correspond with response outcomes to immune stimuli. To test this, we quantified global gene expression in peripheral blood CD4+ cells from healthy individuals at baseline and following CD3/CD28 stimulation at two time points 1 mo apart. Systemic CD4+ cell baseline and poststimulation molecular immune response signatures (MIRS) were defined by identifying genes expressed at levels that were stable between time points within individuals and differential among individuals in each state. Iterative differential gene expression analyses between all possible phenotypic groupings of at least three individuals using the baseline and stimulated MIRS gene sets revealed shared baseline and response phenotypic groupings, indicating the baseline MIRS contained determinants of immune responsiveness. Furthermore, significant numbers of shared phenotype-defining sets of determinants were identified in baseline data across independent healthy cohorts. Combining the cohorts and repeating the analyses resulted in identification of over 6000 baseline immune phenotypic groups, implying that the MIRS concept may be useful in many immune perturbation contexts. These findings demonstrate that patterns in complex gene expression variability can be used to define immune phenotypes and discover determinants of immune responsiveness.

Long intergenic non-coding RNA (lincRNA) is a family of gene transcripts, the functions of which are largely unknown. Although cigarette smoking is the main cause for lung cancer, lung cancer in non-smokers is a separate entity and its underlying cause is little known. Growing evidence suggests lincRNAs play a significant role in cancer development and progression; however, such data is lacking for lung cancer in non-smokers, or those who have never smoked. This study conducted comprehensive profiling of lincRNAs from RNA sequencing (RNA-seq) data of non-smoker patients with lung adenocarcinoma. Both known and novel lincRNAs distinctly segregated tumors from normal tissues. Approximately one third of lincRNAs were differentially expressed between tumors and normal samples and most of them were coordinated with their putative protein gene targets. More importantly, lincRNAs defined two clusters of tumors that were associated with tumor aggressiveness and patient survival. We identified a subset of lincRNAs that were differentially expressed and also associated with patient survival. Very high concordance (R2 = 0.9) was observed for the differentially expressed lincRNAs in the Cancer Genome Atlas (TCGA) validation set of 85 transcriptomes and the lincRNAs associated with survival from the discovery set were similarly predictive in the validation set. These lincRNAs warrant further investigation as potential diagnostic and prognostic markers.

Abstract By regulating gene expression networks that mediate neoplastic behavior, epigenetic protein complexes are the ultimate effectors of oncogenic pathways. This hierarchical position of epigenetic pathways within the mechanisms that regulate cancer makes them ideal candidates for therapeutic targets. The current work was designed to characterize the role of the K9H3 Histone Methyltransferase (HMT) pathway in mediating oncogenic signals downstream of Aurora Kinase A (AurkA), with the goal of designing efficient combinatorial therapies against PDAC. Affinity protein purification, combined with mass spectrometry, demonstrates that HP1γ isolated from mitotic cells interacts with AurkA and the HMTs, G9a and GLP. We also find that this complex is critical for mitotic progression and cell proliferation. Congruently, treatment of PDAC cells with individual drugs against AurkA or HP1-HMT inhibits cell growth by inducing senescence, a cytostatic response. However, the combination of these agents has a synergistic effect of reducing cell growth in both, monolayer and spheroid cultures to result in a cytotoxic effect. Confocal and electron microscopy, along with cell cycle analysis, demonstrate that the cytotoxic effect of this combination is due to induction of mitotic catastrophe, a distinct form of cell death that occurs during mitosis. Molecularly, the combination of AurkA and HP1-HMT inhibition bypasses G2/M arrest with downregulation of the Chk1-Cdc25c pathway. In vivo treatment of PDAC orthotopic xenografts with the HP1-HMT inhibitor alone demonstrated reduced PDAC growth, and increased efficacy in PDAC growth reduction was observed in combination with the AurkA inhibitor over either individual treatment. Thus, our data demonstrate a novel AurkA-HP1-HMT mitotic pathway that holds promise for potential pharmacological targeting in combinatorial therapy for this malignancy. Citation Format: Angela Mathison, Ann Salmonson, Mckenna Missfeldt, Thiago de Assuncao, Jennifer Bintz, Trace Christensen, Sarah Kossak, Asha Nair, Juan Iovanna, Robert Huebert, Gwen Lomberk. Pharmacological targeting of the Aurora A and Histone 3 lysine 9 methyltransferase pathways in pancreatic cancer induces mitotic catastrophe [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 4060. doi:10.1158/1538-7445.AM2017-4060