The novel finding of an FGFR1::TACC1 fusion in an undifferentiated spindle cell sarcoma of soft tissue with aggressive clinical course.

We report an aggressive soft tissue sarcoma with FGFR1-TACC1 fusion occurring in the thigh of an 83-year-old man. Microscopically, the tumor was composed of monomorphic spindle cells arranged in intersecting fascicles. The tumor cells showed ovoid nuclei, fine chromatin, indistinct nucleoli, and elongated eosinophilic cytoplasm. Focal increase in nuclear atypia was noted. Immunohistochemistry showed only focal rare positivity to S100, but negative to SOX10, CD34, STAT6, TLE-1, SMA, and other myogenic markers. An extensive panel of immunostains did not reveal a definite lineage of cellular differentiation. The fusion junction of the chimeric transcript involved FGFR1 exon 16 and TACC1 exon 7, which was similar to those reported in other types of neoplasms such as glioblastomas and epithelial cancers. The transcript was predicted to be in-frame and confirmed by Sanger sequencing after reverse transcriptase-polymerase chain reaction. The patient was treated by marginal excision of tumor without receiving adjuvant therapy. He experienced rapid tumor recurrence with distant metastases and succumbed at 3.5 months after presentation. The finding of FGFR1-TACC1 fusion in a high-grade, undifferentiated spindle cell sarcoma of soft tissue is novel and may offer a potential therapeutic target in the near future.

Hepatitis B Virus-Telomerase Reverse Transcriptase Promoter Integration Harnesses Host ELF4, Resulting in Telomerase Reverse Transcriptase Gene Transcription in Hepatocellular Carcinoma.

BACKGROUND AND AIMS: Hepatitis B virus (HBV) integrations are common in hepatocellular carcinoma (HCC). In particular, alterations of the telomerase reverse transcriptase (TERT) gene by HBV integrations are frequent; however, the molecular mechanism and functional consequence underlying TERT HBV integration are unclear. APPROACH AND RESULTS: We adopted a targeted sequencing strategy to survey HBV integrations in human HBV-associated HCCs (n = 95). HBV integration at the TERT promoter was frequent (35.8%, n = 34/95) in HCC tumors and was associated with increased TERT mRNA expression and more aggressive tumor behavior. To investigate the functional importance of various integrated HBV components, we employed different luciferase reporter constructs and found that HBV enhancer I (EnhI) was the key viral component leading to TERT activation on integration at the TERT promoter. In addition, the orientation of the HBV integration at the TERT promoter further modulated the degree of TERT transcription activation in HCC cell lines and patients' HCCs. Furthermore, we performed array-based small interfering RNA library functional screening to interrogate the potential major transcription factors that physically interacted with HBV and investigated the cis-activation of host TERT gene transcription on viral integration. We identified a molecular mechanism of TERT activation through the E74 like ETS transcription factor 4 (ELF4), which normally could drive HBV gene transcription. ELF4 bound to the chimeric HBV EnhI at the TERT promoter, resulting in telomerase activation. Stable knockdown of ELF4 significantly reduced the TERT expression and sphere-forming ability in HCC cells. CONCLUSIONS: Our results reveal a cis-activating mechanism harnessing host ELF4 and HBV integrated at the TERT promoter and uncover how TERT HBV-integrated HCCs may achieve TERT activation in hepatocarcinogenesis.

RSK2-inactivating mutations potentiate MAPK signaling and support cholesterol metabolism in hepatocellular carcinoma.

BACKGROUND & AIMS: Mutational profiling of patient tumors has suggested that hepatocellular carcinoma (HCC) development is mainly driven by loss-of-function mutations in tumor suppressor genes. p90 ribosomal S6 kinase 2 (RSK2) functions as a direct downstream kinase of ERK1/2 and elevated RSK2 expression has been reported to support oncogenic functions in some cancers. We investigated if RSK2 was also dysregulated by inactivating mutations in cancers including HCC. METHODS: We performed exome sequencing and targeted DNA sequencing on HBV-associated HCCs to examine recurrent RSK2 mutations. The functional significance and mechanistic consequences of RSK2 mutations were examined in natural RSK2-null HCC cells, and RSK2-knockout HCC cells. The potential downstream pathways underlying RSK2 mutations were investigated by RNA sequencing, qRT-PCR and mass spectrometry. RESULTS: We detected recurrent somatic RSK2 mutations at a rate of 6.3% in our HCC cohorts and revealed that, among many cancer types, HCC was the cancer most commonly harboring RSK2 mutations. The RSK2 mutations were inactivating and associated with a more aggressive tumor phenotype. We found that, functionally, restoring RSK2 expression in natural RSK2-null HBV-positive Hep3B cells suppressed proliferation and migration in vitro and tumorigenicity in vivo. Mechanistically, RSK2-inactivating mutations attenuated a SOS1/2-dependent negative feedback loop, leading to the activation of MAPK signaling. Of note, this RSK2 mutation-mediated MAPK upregulation rendered HCC cells more sensitive to sorafenib, a first-line multi-kinase inhibitor for advanced HCC. Furthermore, such activation of MAPK signaling enhanced cholesterol biosynthesis-related gene expression in HCC cells. CONCLUSIONS: Our findings reveal the mechanistic and functional significance of RSK2-inactivating mutations in HCC. These inactivating mutations may serve as an alternative route to activate MAPK signaling and cholesterol metabolism in HCC. LAY SUMMARY: In this study, we identified and functionally characterized RSK2-inactivating mutations in human hepatocellular carcinoma and demonstrated their association with aggressive tumor behavior. Mutations in RSK2 drive signaling pathways with known oncogenic potential, leading to enhanced cholesterol biosynthesis and potentially sensitizing tumors to sorafenib treatment.

Round Cell Sarcoma with EWSR1-PATZ1 Fusion in the Face of a Five-Year-Old Boy: Report of a Case with Unusual Histologic Features.

Round cell sarcomas with EWSR1-PATZ1 fusion are rare polyphenotypic sarcomas that typically show both neural and myogenic differentiation on immunohistochemistry. The histology features lobular admixture of cellular fascicles of relatively monotonous spindle cells and small blue round cells separated by fibrotic stroma. The clinical behavior of EWSR1-PATZ1 sarcoma is uncertain currently with mixed outcomes reported even in cases with metastases. We herein report an additional case of EWSR1-PATZ1 fusion-related round cell sarcoma in the face of a 5-year-old boy with unusual histologic features of pale zones, rosette/gland-like structures and expression of epithelial markers. Fluorescent in-situ hybridization study (FISH) using EWSR1 breakapart probes was negative and molecular study with RNA sequencing was required to confirm the diagnosis. These findings highlight the diagnostic challenge and potential pitfall of FISH study in EWSR1-PATZ1 sarcoma. Further studies are required to increase the understanding of their behavior, morphologic spectrum and molecular features that will help devise new treatment strategies to these rare tumours.

Single-cell RNA sequencing shows the immunosuppressive landscape and tumor heterogeneity of HBV-associated hepatocellular carcinoma.

Interaction between tumor cells and immune cells in the tumor microenvironment is important in cancer development. Immune cells interact with the tumor cells to shape this process. Here, we use single-cell RNA sequencing analysis to delineate the immune landscape and tumor heterogeneity in a cohort of patients with HBV-associated human hepatocellular carcinoma (HCC). We found that tumor-associated macrophages suppress tumor T cell infiltration and TIGIT-NECTIN2 interaction regulates the immunosuppressive environment. The cell state transition of immune cells towards a more immunosuppressive and exhaustive status exemplifies the overall cancer-promoting immunocellular landscape. Furthermore, the heterogeneity of global molecular profiles reveals co-existence of intra-tumoral and inter-tumoral heterogeneity, but is more apparent in the latter. This analysis of the immunosuppressive landscape and intercellular interactions provides mechanistic information for the design of efficacious immune-oncology treatments in hepatocellular carcinoma.

Bone Sarcoma With EWSR1-NFATC2 Fusion: Sarcoma With Varied Morphology and Amplification of Fusion Gene Distinct From Ewing Sarcoma.

Ewing sarcomas are typified by EWSR1 fusion to ETS gene family members. Tumors with fusion partners other than ETS family members and atypical histologic features pose significant diagnostic challenges and controversies as to their classification. In this article, we report a tumor with EWSR1-NFATC2 fusion in the left femur of a 43-year-old man and with unusual morphologic features that resemble undifferentiated high-grade sarcoma. Analysis together with reported cases in the literature shows that tumors with EWSR1-NFATC2 exhibit distinctive clinicopathologic features, including predilection for young male adults, highly variable histology that varies from round cell tumors frequently associated with nuclear irregularity, short spindle cells with nuclear pleomorphism, to myoepithelial tumor-like with or without myxohyaline matrix. They show variable positivity to CD99, frequent expression of cytokeratins, and consistent high-level amplification of EWSR1-NFATC2 fusion gene with distinctive gene expression profile. These tumors thus deserve classification separate from Ewing sarcoma.