Glycine decarboxylase activity drives non-small cell lung cancer tumor-initiating cells and tumorigenesis.

Identification of the factors critical to the tumor-initiating cell (TIC) state may open new avenues in cancer therapy. Here we show that the metabolic enzyme glycine decarboxylase (GLDC) is critical for TICs in non-small cell lung cancer (NSCLC). TICs from primary NSCLC tumors express high levels of the oncogenic stem cell factor LIN28B and GLDC, which are both required for TIC growth and tumorigenesis. Overexpression of GLDC and other glycine/serine enzymes, but not catalytically inactive GLDC, promotes cellular transformation and tumorigenesis. We found that GLDC induces dramatic changes in glycolysis and glycine/serine metabolism, leading to changes in pyrimidine metabolism to regulate cancer cell proliferation. In the clinic, aberrant activation of GLDC correlates with poorer survival in lung cancer patients, and aberrant GLDC expression is observed in multiple cancer types. This link between glycine metabolism and tumorigenesis may provide novel targets for advancing anticancer therapy.

A multimodal atlas of hepatocellular carcinoma reveals convergent evolutionary paths and 'bad apple' effect on clinical trajectory.

BACKGROUND & AIMS: Hepatocellular Carcinoma (HCC) is a highly fatal cancer characterized by high intra-tumor heterogeneity (ITH). A panoramic understanding of its tumor evolution, in relation to its clinical trajectory, may provide novel prognostic and treatment strategies. METHODS: Through the Asia-Pacific Hepatocellular Carcinoma (AHCC) trials group (NCT03267641), we recruited one of the largest prospective cohorts of HCC with over 600 whole genome and transcriptome samples from 123 treatment-naïve patients. RESULTS: Using a multi-region sampling approach, we revealed seven convergent genetic evolutionary paths governed by the early driver mutations, late copy number variations and viral integrations, which stratify patient clinical trajectories after surgical resection. Furthermore, such evolutionary paths shaped the molecular profiles, leading to distinct transcriptomic subtypes. Most significantly, although we found the coexistence of multiple transcriptomic subtypes within certain tumors, patient prognosis was best predicted by the most aggressive cell fraction of the tumor, rather than by overall degree of transcriptomic ITH level - a phenomenon we termed the 'bad apple' effect. Finally, we found that characteristics throughout early and late tumor evolution provide significant and complementary prognostic power in predicting patient survival. CONCLUSIONS: Taken together, our study generated a comprehensive landscape of evolutionary history for HCC and provided a rich multi-omics resource for understanding tumor heterogeneity and clinical trajectories. CLINICAL TRIAL NUMBER: NCT03267641 (Observational cohort) IMPACT AND IMPLICATIONS: This prospective study, utilizing comprehensive multi-sector, multi-omics sequencing and clinical data from surgically resected HCC, reveals critical insights into the role of tumor evolution and intra-tumor heterogeneity (ITH) in determining the prognosis of Hepatocellular Carcinoma (HCC). These findings are invaluable for oncology researchers and clinicians, as they underscore the influence of distinct evolutionary paths and the 'bad apple' effect, where the most aggressive tumor fraction dictates disease progression. These insights not only enhance prognostic accuracy post-surgical resection but also pave the way for developing personalized therapies tailored to specific tumor evolutionary and transcriptomic profiles. The co-existence of multiple sub-types within the same tumor prompts a re-appraisal of the utilities of depending on single samples to represent the entire tumor and suggests the need for clinical molecular imaging. This research thus marks a significant step forward in the clinical understanding and management of HCC, underscoring the importance of integrating tumor evolutionary dynamics and multi-omics biomarkers into therapeutic decision-making.

Hypoxia-driven immunosuppression by Treg and type-2 conventional dendritic cells in HCC.

BACKGROUND AND AIMS: Hypoxia is one of the central players in shaping the immune context of the tumor microenvironment (TME). However, the complex interplay between immune cell infiltrates within the hypoxic TME of HCC remains to be elucidated. APPROACH AND RESULTS: We analyzed the immune landscapes of hypoxia-low and hypoxia-high tumor regions using cytometry by time of light, immunohistochemistry, and transcriptomic analyses. The mechanisms of immunosuppression in immune subsets of interest were further explored using in vitro hypoxia assays. Regulatory T cells (Tregs) and a number of immunosuppressive myeloid subsets, including M2 macrophages and human leukocyte antigen-DR isotype (HLA-DRlo ) type 2 conventional dendritic cell (cDC2), were found to be significantly enriched in hypoxia-high tumor regions. On the other hand, the abundance of active granzyme Bhi PD-1lo CD8+ T cells in hypoxia-low tumor regions implied a relatively active immune landscape compared with hypoxia-high regions. The up-regulation of cancer-associated genes in the tumor tissues and immunosuppressive genes in the tumor-infiltrating leukocytes supported a highly pro-tumorigenic network in hypoxic HCC. Chemokine genes such as CCL20 (C-C motif chemokine ligand 20) and CXCL5 (C-X-C motif chemokine ligand 5) were associated with recruitment of both Tregs and HLA-DRlo cDC2 to hypoxia-high microenvironments. The interaction between Tregs and cDC2 under a hypoxic TME resulted in a loss of antigen-presenting HLA-DR on cDC2. CONCLUSIONS: We uncovered the unique immunosuppressive landscapes and identified key immune subsets enriched in hypoxic HCC. In particular, we identified a potential Treg-mediated immunosuppression through interaction with a cDC2 subset in HCC that could be exploited for immunotherapies.

Multi-region sampling with paired sample sequencing analyses reveals sub-groups of patients with novel patient-specific dysregulation in Hepatocellular Carcinoma.

BACKGROUND: Conventional differential expression (DE) testing compares the grouped mean value of tumour samples to the grouped mean value of the normal samples, and may miss out dysregulated genes in small subgroup of patients. This is especially so for highly heterogeneous cancer like Hepatocellular Carcinoma (HCC). METHODS: Using multi-region sampled RNA-seq data of 90 patients, we performed patient-specific differential expression testing, together with the patients' matched adjacent normal samples. RESULTS: Comparing the results from conventional DE analysis and patient-specific DE analyses, we show that the conventional DE analysis omits some genes due to high inter-individual variability present in both tumour and normal tissues. Dysregulated genes shared in small subgroup of patients were useful in stratifying patients, and presented differential prognosis. We also showed that the target genes of some of the current targeted agents used in HCC exhibited highly individualistic dysregulation pattern, which may explain the poor response rate. DISCUSSION/CONCLUSION: Our results highlight the importance of identifying patient-specific DE genes, with its potential to provide clinically valuable insights into patient subgroups for applications in precision medicine.

Extraction of intra-biliary hepatocellular carcinoma by endoscopic retrograde cholangiopancreatography.

BACKGROUND: Hepatocellular carcinoma with biliary ductal invasion is rare and associated with a significantly lower survival rate. CASE PRESENTATION: We present an unusual case of a patient with hepatocellular carcinoma and biliary invasion, who had his diagnosis confirmed by histological analysis from tissue extracted by endoscopic retrograde cholangiopancreatography. An 87-year-old male presented with a 1-day history of right upper quadrant pain and jaundice. His past medical history included recurrent gallstone cholangitis and a previous cholecystectomy. An abdominal CT demonstrated a dilated intrahepatic biliary tree with left proximal intrahepatic hyperdensities, as well as a 3 cm hepatocellular carcinoma. He was initially suspected to have concurrent gallstone cholangitis and a newly diagnosed hepatocellular carcinoma. Endoscopic retrograde cholangiopancreatography and balloon trawling of the intraductal lesions extracted necrotic tumour-like tissue which was histologically consistent with hepatocellular carcinoma. The extraction of the intra-biliary portion of HCC resulted in complete resolution of his jaundice, enabling further treatment with nivolumab, which would not have been possible if the obstruction was not cleared. The patient is currently well and has completed his 6th cycle of nivolumab. CONCLUSION: Obstructive jaundice is an uncommon presentation for patients with HCC. it is key for clinicians to be aware of the possibility of intrabiliary invasion in order obtain an early diagnosis and to reduce any delay in treatment.

Primary Renal Hybrid Low-grade Fibromyxoid Sarcoma-Sclerosing Epithelioid Fibrosarcoma: An Unusual Pediatric Case With EWSR1-CREB3L1 Fusion.

Low-grade fibromyxoid sarcoma (LGFMS) and sclerosing epithelioid fibrosarcoma (SEF) are rare tumors with distinct sets of morphological features, both characterized by MUC4 immunoreactivity. Tumors exhibiting features of both entities are considered hybrid LGFMS-SEF lesions. While the majority of LGFMS cases are characterized by FUS-CREB3L2 gene fusions, most cases of pure SEF show EWSR1 gene rearrangements. In the largest study of hybrid LGFMS-SEF tumors to date, all cases exhibited FUS rearrangements, a similar genetic profile to LGFMS. We herein describe the clinicopathological features and genetic findings of a case of primary renal hybrid LGFMS-SEF occurring in a 10-year-old child, with disseminated metastases. Fusion gene detection using a next-generation sequencing-based anchored multiplex PCR technique (Archer FusionPlex Sarcoma Panel) was performed on both the primary renal tumor that showed the morphology of a LGFMS, and a cervical metastasis that showed the morphology of SEF. An EWSR1-CREB3L1 gene fusion occurring between exon 11 of EWSR1 and exon 6 of CREB3L1 was present in both the LGFMS and SEF components. This unusual case provides evidence that a subset of hybrid LGFMS-SEF harbor EWSR1-CREB3L1 gene fusions. In this case, these features were associated with an aggressive clinical course, with disease-associated mortality occurring within 12 months of diagnosis.

Fine-needle aspiration of a pancreas with squamoid cyst of pancreatic duct and pancreatic duct stones: A case report of a rare pancreatic lesion and an unusual concurrent finding of stone-related crystals.

Squamoid cyst of pancreatic duct is a rare benign pancreatic lesion that is rarely encountered in fine-needle aspiration (FNA) and surgical resection specimens. Pancreatic stones can be seen in chronic pancreatitis, but stone-related crystals have previously not been described in pancreatic cytology. Presented here is a case report of a squamoid cyst of pancreatic duct with concurrent pancreatic duct stones. We describe the cytomorphology of this benign cyst, as well as the remarkable finding of polymorphous crystals on cyst fluid aspirate. We also describe the histology of the surgically resected cystic lesion. With the increase in detection of incidental pancreatic cysts on imaging, this case highlights the importance of awareness and recognition of benign non-neoplastic epithelial cysts on FNA sampling to avoid overtreatment. The presence of crystals on pancreatic FNA is an unusual finding, likely representing calcium carbonate crystals related to the formation of pancreatic duct stones.

IFNγ-IL-17+ CD8 T cells contribute to immunosuppression and tumor progression in human hepatocellular carcinoma.

IL-17-producing CD8 (Tc17) T cells have been shown to play an important role in infection and chronic inflammation, however their implications in hepatocellular carcinoma (HCC) remain elusive. In this study, we performed cytometry by time-of-flight (CyTOF) and revealed the distinctive immunological phenotypes of two IFNγ+ and IFNγ- Tc17 subsets that were preferentially enriched in human HCC. Single-cell RNA-sequencing analysis further revealed regulatory circuits governing the different phenotypes of these Tc17 subsets. In particular, we discovered that IFNγ- Tc17 subset demonstrated pro-tumoral characteristics and expressed higher levels of CCL20. This corresponded to increased tumor infiltration of T regulatory cells (Treg) validated by immunohistochemistry in another independent HCC cohort, demonstrating the immunosuppressive functions of IFNγ- Tc17 subset. Most importantly, higher intra-tumoral proportions of IFNγ- Tc17 were associated with poorer prognosis in patients with HCC and this was further validated in The Cancer Genome Atlas (TCGA) HCC cohort. Taken together, this compendium of transcriptomic and proteomic data of Tc17 subsets sheds light on the immunosuppressive phenotypes of IFNγ- Tc17 and its implications in HCC progression.

A genomic enhancer signature associates with hepatocellular carcinoma prognosis.

Background & Aims: Lifestyle and environmental-related exposures are important risk factors for hepatocellular carcinoma (HCC), suggesting that epigenetic dysregulation significantly underpins HCC. We profiled 30 surgically resected tumours and the matched adjacent normal tissues to understand the aberrant epigenetic events associated with HCC. Methods: We identified tumour differential enhancers and the associated genes by analysing H3K27 acetylation (H3K27ac) chromatin immunoprecipitation sequencing (ChIP-seq) and Hi-C/HiChIP data from the resected tumour samples of 30 patients with early-stage HCC. This epigenome dataset was analysed with previously reported genome and transcriptome data of the overlapping group of patients from the same cohort. We performed patient-specific differential expression testing using multiregion sequencing data to identify genes that undergo both enhancer and gene expression changes. Based on the genes selected, we identified two patient groups and performed a recurrence-free survival analysis. Results: We observed large-scale changes in the enhancer distribution between HCC tumours and the adjacent normal samples. Many of the gain-in-tumour enhancers showed corresponding upregulation of the associated genes and vice versa, but much of the enhancer and gene expression changes were patient-specific. A subset of the upregulated genes was activated in a subgroup of patients' tumours. Recurrence-free survival analysis revealed that the patients with a more robust upregulation of those genes showed a worse prognosis. Conclusions: We report the genomic enhancer signature associated with differential prognosis in HCC. Findings that cohere with oncofoetal reprogramming in HCC were underpinned by genome-wide enhancer rewiring. Our results present the epigenetic changes in HCC that offer the rational selection of epigenetic-driven gene targets for therapeutic intervention or disease prognostication in HCC. Impact and Implications: Lifestyle and environmental-related exposures are the important risk factors of hepatocellular carcinoma (HCC), suggesting that tumour-associated epigenetic dysregulations may significantly underpin HCC. We profiled tumour tissues and their matched normal from 30 patients with early-stage HCC to study the dysregulated epigenetic changes associated with HCC. By also analysing the patients' RNA-seq and clinical data, we found the signature genes - with epigenetic and transcriptomic dysregulation - associated with worse prognosis. Our findings suggest that systemic approaches are needed to consider the surrounding cellular environmental and epigenetic changes in HCC tumours.

Dynamic phenotypic heterogeneity and the evolution of multiple RNA subtypes in hepatocellular carcinoma: the PLANET study.

Intra-tumor heterogeneity (ITH) is a key challenge in cancer treatment, but previous studies have focused mainly on the genomic alterations without exploring phenotypic (transcriptomic and immune) heterogeneity. Using one of the largest prospective surgical cohorts for hepatocellular carcinoma (HCC) with multi-region sampling, we sequenced whole genomes and paired transcriptomes from 67 HCC patients (331 samples). We found that while genomic ITH was rather constant across stages, phenotypic ITH had a very different trajectory and quickly diversified in stage II patients. Most strikingly, 30% of patients were found to contain more than one transcriptomic subtype within a single tumor. Such phenotypic ITH was found to be much more informative in predicting patient survival than genomic ITH and explains the poor efficacy of single-target systemic therapies in HCC. Taken together, we not only revealed an unprecedentedly dynamic landscape of phenotypic heterogeneity in HCC, but also highlighted the importance of studying phenotypic evolution across cancer types.

Trajectory of immune evasion and cancer progression in hepatocellular carcinoma.

Immune evasion is key to cancer initiation and later at metastasis, but its dynamics at intermediate stages, where potential therapeutic interventions could be applied, is undefined. Here we show, using multi-dimensional analyses of resected tumours, their adjacent non-tumour tissues and peripheral blood, that extensive immune remodelling takes place in patients with stage I to III hepatocellular carcinoma (HCC). We demonstrate the depletion of anti-tumoural immune subsets and accumulation of immunosuppressive or exhausted subsets along with reduced tumour infiltration of CD8 T cells peaking at stage II tumours. Corresponding transcriptomic modification occur in the genes related to antigen presentation, immune responses, and chemotaxis. The progressive immune evasion is validated in a murine model of HCC. Our results show evidence of ongoing tumour-immune co-evolution during HCC progression and offer insights into potential interventions to reverse, prevent or limit the progression of the disease.

Predicting HCC Response to Multikinase Inhibitors With In Vivo Cirrhotic Mouse Model for Personalized Therapy.

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) arises in a cirrhotic, pro-angiogenic microenvironment. Inhibiting angiogenesis is a key mode of action of multikinase inhibitors and current non-cirrhotic models are unable to predict treatment response. We present a novel mouse cirrhotic model of xenotransplant that predicts the natural biology of HCC and allows personalized therapy. METHODS: Cirrhosis was induced in NOD Scid gamma mice with 4 months of thioacetamide administration. Patient derived xenografts (PDXs) were created by transplant of human HCC subcutaneously into non-cirrhotic mice and intra-hepatically into both cirrhotic and non-cirrhotic mice. The applicability of cirrhotic PDXs for drug testing was tested with 16 days of either sorafenib or lenvatinib. Treatment response was evaluated by MRI. RESULTS: 8 out of 19 (42%) human HCC engrafted in the cirrhotic model compared with only 3 out of 19 (16%) that engrafted in the subcutaneous non-cirrhotic model. Tumor vasculature was preserved in the cirrhotic model but was diminished in the non-cirrhotic models. Metastasis developed in 3 cirrhotic PDX lines and was associated with early HCC recurrence in all 3 corresponding patients (100%), compared with only 5 out of 16 (31%) of the other PDX lines, P = .027. The cirrhotic model was able to predict response and non-response to lenvatinib and sorafenib respectively in the corresponding patients. Response to lenvatinib in the cirrhotic PDX was associated with reduction in CD34, VEGFR2 and CLEC4G immunofluorescence area and intensity (all P ≤ .03). CONCLUSIONS: A clinically relevant cirrhotic PDX model preserves tumor angiogenesis and allows prediction of response to multikinase inhibitors for personalized therapy.

Intratumoural immune heterogeneity as a hallmark of tumour evolution and progression in hepatocellular carcinoma.

The clinical relevance of immune landscape intratumoural heterogeneity (immune-ITH) and its role in tumour evolution remain largely unexplored. Here, we uncover significant spatial and phenotypic immune-ITH from multiple tumour sectors and decipher its relationship with tumour evolution and disease progression in hepatocellular carcinomas (HCC). Immune-ITH is associated with tumour transcriptomic-ITH, mutational burden and distinct immune microenvironments. Tumours with low immune-ITH experience higher immunoselective pressure and escape via loss of heterozygosity in human leukocyte antigens and immunoediting. Instead, the tumours with high immune-ITH evolve to a more immunosuppressive/exhausted microenvironment. This gradient of immune pressure along with immune-ITH represents a hallmark of tumour evolution, which is closely linked to the transcriptome-immune networks contributing to disease progression and immune inactivation. Remarkably, high immune-ITH and its transcriptomic signature are predictive for worse clinical outcome in HCC patients. This in-depth investigation of ITH provides evidence on tumour-immune co-evolution along HCC progression.

Bowel ulceration following tocilizumab administration in a COVID-19 patient.

Tocilizumab, a monoclonal antibody against interleukin-6, has been used to treat cytokine release syndrome (CRS) in a subset of patients with severe COVID-19 disease. Acute ulcerative bowel disease has been only rarely documented in patients treated for rheumatological conditions. The gastrointestinal side effects seen when used in the context of COVID-19 are unknown. We present a case of COVID-19 CRS in which acute terminal ileum and perforated caecal ulceration evolved after tocilizumab exposure. We raise awareness of a possible causal relationship between even a single dose of tocilizumab and gut ulceration in patients with COVID-19. Any such drug enteropathy relationship requires watchful monitoring during upcoming trials of tocilizumab in patients with COVID-19.

Anaplastic Oligodendroglioma with Transdural Extension.

Oligodendrogliomas, the third most common primary gliomas, have a strict molecular definition, characterized by the combined presence of isocitrate dehydrogenase mutation and 1p19q codeletion. Herein, we describe an extremely unusual case of molecularly defined anaplastic oligodendroglioma with transdural extension into the frontal and ethmoid sinuses, without prior neurosurgical intervention or radiotherapy. The molecular profile of the tumor is also provided. To the best of our knowledge, this has never been reported before. Most of the previously reported glial tumors with transdural extension were cases of histologically proven glioblastomas and gliosarcomas, typically seen in the context of prior neurosurgical intervention and/or radiotherapy. This case adds to the limited literature on oligodendrogliomas with transdural extension. Further studies are necessary to elucidate the relationship between the incidence of transdural extension and molecular subtypes of oligodendrogliomas.

Two Extraordinary Sellar Neuronal Tumors: Literature Review and Comparison of Clinicopathologic Features.

Objectives: The list of tumors involving the pituitary gland has been expanded to include a variety of neuronal and paraneuronal tumors in the 2017 World Health Organization tumor classification of endocrine organs. All the entities included in this category are distinctly rare, with limited case reports in the literature. Methods: We illustrate two extraordinary sellar tumors with neuronal differentiation: a sellar paraganglioma and a sellar neurocytoma, with thorough literature review and comparison of the clinicopathologic features of these entities. Results: Both entities are exceptionally rare and tend to be misdiagnosed as pituitary adenoma preoperatively. Both entities demonstrate frequent clinical recurrence compared with pituitary adenoma, as well as the rare occurrence of metastatic disease. Conclusions: In evaluating a sellar tumor with an uncommon morphology and neuroendocrine differentiation, an increased awareness of the unusual entities that may involve the sellar region and a judicious panel of immunohistochemical studies should improve the diagnosis.

Unusual presentation of osteoblastoma as vertebra plana-a case report and review of literature.

BACKGROUND: Osteoblastoma is rare and accounts for 3% of all benign tumors and 1% of all bone tumors. The spine is the most common site of occurrence, constituting 32% to 45% of all osteoblastomas. It has a strong predilection for the posterior elements, most often occurring in the lumbar spine. METHOD: In this case report, we describe an unusual presentation of spinal osteoblastoma presenting as thoracic T9 vertebra plana in a 20-year-old female. She presented with discomfort over the midback with unsteadiness of gait. The patient underwent detailed investigations including computed tomography (CT), magnetic resonance imaging, and CT-guided biopsy. To our knowledge, this is the first case report of vertebra plana due to spinal osteoblastoma in the English literature. RESULT: The patient successfully underwent posterior decompression of T9 with laminectomy followed by minimally invasive surgery posterior instrumentation from T7 to T11. Histopathology of the intraoperative specimen was consistent with osteoblastoma. The patient had an uneventful postoperative recovery and no evidence of tumor recurrence could be demonstrated on positron emission tomography scan at 15 months' follow-up. CONCLUSION: In conclusion, the differential diagnosis for vertebra plana is extensive and we add spinal osteoblastoma as another etiology to the existing list. Diagnosis and treatment of vertebra plana involve multimodality radiological imaging, and careful histological and surgical evaluation to identify the underlying etiology.

Hybrid Intercalated Duct Lesion of the Parotid: Diagnostic Challenges of a Recently Described Entity with Fine Needle Aspiration Findings.

Intercalated duct lesions (IDL) of the salivary glands are recently described, and encompass both hyperplasia and benign neoplasms that remain incompletely understood. IDLs have been linked to various benign and low-grade malignant salivary gland neoplasms. We herein present a case of a 77 year old woman with an IDL of the parotid composed of both a hyperplastic and an adenomatous component and report, for the first time, the fine needle aspiration findings of such a lesion. This case illustrates the morphologic spectrum of an IDL, as well as challenges in rendering an accurate cytological and histologic diagnosis. The potential diagnostic pitfalls presented by the hybrid pattern of this lesion are also discussed.

Tumour-initiating cell-specific miR-1246 and miR-1290 expression converge to promote non-small cell lung cancer progression.

The tumour-initiating cell (TIC) model accounts for phenotypic and functional heterogeneity among tumour cells. MicroRNAs (miRNAs) are regulatory molecules frequently aberrantly expressed in cancers, and may contribute towards tumour heterogeneity and TIC behaviour. More recent efforts have focused on miRNAs as diagnostic or therapeutic targets. Here, we identified the TIC-specific miRNAs, miR-1246 and miR-1290, as crucial drivers for tumour initiation and cancer progression in human non-small cell lung cancer. The loss of either miRNA impacted the tumour-initiating potential of TICs and their ability to metastasize. Longitudinal analyses of serum miR-1246 and miR-1290 levels across time correlate their circulating levels to the clinical response of lung cancer patients who were receiving ongoing anti-neoplastic therapies. Functionally, direct inhibition of either miRNA with locked nucleic acid administered systemically, can arrest the growth of established patient-derived xenograft tumours, thus indicating that these miRNAs are clinically useful as biomarkers for tracking disease progression and as therapeutic targets.