Identification and Characterization of Chemotherapy-Resistant High-Risk Neuroblastoma Persister Cells.

Relapse rates in high-risk neuroblastoma remain exceedingly high. The malignant cells that are responsible for relapse have not been identified, and mechanisms of therapy resistance remain poorly understood. In this study, we used single-nucleus RNA sequencing and bulk whole-genome sequencing to identify and characterize the residual malignant persister cells that survive chemotherapy from a cohort of 20 matched diagnosis and definitive surgery tumor samples from patients treated with high-risk neuroblastoma induction chemotherapy.

Nuclear factor-κB activation by transforming growth factor-β1 drives tumour microenvironment-mediated drug resistance in neuroblastoma.

Background: Intrinsic and extrinsic factors in the tumour microenvironment (TME) contribute to therapeutic resistance. Here we demonstrate that transforming growth factor (TGF)-β1 produced in the TME increased drug resistance of neuroblastoma (NB) cells.

Methods: Human NB cell lines were tested in vitro for their sensitivity to Doxorubicin (DOX) and Etoposide (ETOP) in the presence of tumour-associated macrophages (TAM) and mesenchymal stromal cells/cancer-associated fibroblasts (MSC/CAF).

Envision the future of precision medicine in pediatric cancer.

Exploring the diversity within the tumor microenvironment (TME) can offer crucial insights to steer cancer therapy toward precision medicine. In this issue of Cancer Cell, Wienke et al. undertake a comprehensive single-cell analysis of neuroblastoma, unveiling its immune landscape and identifying NECTIN2-TIGIT as a promising target for immunotherapy.

The capture of extracellular vesicles endogenously released by xenotransplanted tumours induces an inflammatory reaction in the premetastatic niche.

The capture of tumour-derived extracellular vesicles (TEVs) by cells in the tumour microenvironment (TME) contributes to metastasis and notably to the formation of the pre-metastatic niche (PMN). However, due to the challenges associated with modelling release of small EVs in vivo, the kinetics of PMN formation in response to endogenously released TEVs have not been examined. Here, we have studied the endogenous release of TEVs in mice orthotopically implanted with metastatic human melanoma (MEL) and neuroblastoma (NB) cells releasing GFP-tagged EVs (GFTEVs) and their capture by host cells to demonstrate the active contribution of TEVs to metastasis.

Fibroblasts and macrophages cooperate to create a pro-tumorigenic and immune resistant environment via activation of TGF-β/IL-6 pathway in neuroblastoma.

Tumor-associated macrophages (TAM) and cancer-associated fibroblasts (CAF) and their precursor mesenchymal stromal cells (MSC) are often detected together in tumors, but how they cooperate is not well understood. Here, we show that TAM and CAF are the most abundant nonmalignant cells and are present together in untreated human neuroblastoma (NB) tumors that are also poorly infiltrated with T and natural killer (NK) cells. We then show that MSC and CAF-MSC harvested from NB tumors protected human monocytes (MN) from spontaneous apoptosis in an interleukin (IL)-6 dependent mechanism.

Cancer-Associated Fibroblasts: Understanding Their Heterogeneity.

The tumor microenvironment (TME) plays a critical role in tumor progression. Among its multiple components are cancer-associated fibroblasts (CAFs) that are the main suppliers of extracellular matrix molecules and important contributors to inflammation. As a source of growth factors, cytokines, chemokines and other regulatory molecules, they participate in cancer progression, metastasis, angiogenesis, immune cell reprogramming and therapeutic resistance.

The Tumor Microenvironment in Neuroblastoma: New Players, New Mechanisms of Interaction and New Perspectives.

The contribution of the tumor microenvironment (TME) to cancer progression has been well recognized in recent decades. As cancer therapeutic strategies are increasingly precise and include immunotherapies, knowledge of the nature and function of the TME in a tumor becomes essential. Our understanding of the TME in neuroblastoma (NB), the second most common solid tumor in children, has significantly progressed from an initial focus on its Schwannian component to a better awareness of its complex nature, which includes not only immune but also non-immune cells such as cancer-associated fibroblasts (CAFs), the contribution of which to inflammation and interaction with tumor-associated macrophages (TAMs) is now recognized.

Extracellular Vesicle and Particle Biomarkers Define Multiple Human Cancers.

There is an unmet clinical need for improved tissue and liquid biopsy tools for cancer detection. We investigated the proteomic profile of extracellular vesicles and particles (EVPs) in 426 human samples from tissue explants (TEs), plasma, and other bodily fluids. Among traditional exosome markers, CD9, HSPA8, ALIX, and HSP90AB1 represent pan-EVP markers, while ACTB, MSN, and RAP1B are novel pan-EVP markers.

The plasminogen activator inhibitor-1 paradox in cancer: a mechanistic understanding.

The paradoxical pro-tumorigenic function of plasminogen activator inhibitor 1 (PAI-1, aka Serpin E1) in cancer progression and metastasis has been the subject of an abundant scientific literature that has pointed to a pro-angiogenic role, a growth and migration stimulatory function, and an anti-apoptotic activity, all directed toward promoting tumor growth, cancer cell survival, and metastasis. With uPA, PAI-1 is among the most reliable biomarkers and prognosticators in many cancer types. More recently, a novel pro-tumorigenic function of PAI-1 in cancer-related inflammation has been demonstrated.

Anti-CD105 Antibody Eliminates Tumor Microenvironment Cells and Enhances Anti-GD2 Antibody Immunotherapy of Neuroblastoma with Activated Natural Killer Cells.

Purpose: We determined whether elimination of CD105 cells in the tumor microenvironment (TME) with anti-CD105 antibodies enhanced anti-disialoganglioside (GD2) antibody dinutuximab therapy of neuroblastoma when combined with activated natural killer (aNK) cells.

Experimental Design: The effect of MSCs and monocytes on antibody-dependent cellular cytotoxicity (ADCC) mediated by dinutuximab with aNK cells against neuroblastoma cells was determined . ADCC with anti-CD105 mAb TRC105 and aNK cells against MSCs, monocytes, and endothelial cells, which express CD105, was evaluated.

Plasminogen Activator Inhibitor-1 Promotes the Recruitment and Polarization of Macrophages in Cancer.

Plasminogen activator inhibitor-1 (PAI-1) has a pro-tumorigenic function via its pro-angiogenic and anti-apoptotic activities. Here, we demonstrate that PAI-1 promotes the recruitment and M2 polarization of monocytes/macrophages through different structural domains. Its LRP1 interacting domain regulated macrophage migration, while its C-terminal uPA interacting domain promoted M2 macrophage polarization through activation of p38MAPK and nuclear factor κB (NF-κB) and induction of an autocrine interleukin (IL)-6/STAT3 activation pathway.

Conditional Knockdown of Gene Expression in Cancer Cell Lines to Study the Recruitment of Monocytes/Macrophages to the Tumor Microenvironment.

siRNA and shRNA-mediated knock down (KD) methods of regulating gene expression are invaluable tools for understanding gene and protein function. However, in the case that the KD of the protein of interest has a lethal effect on cells or the anticipated effect of the KD is time-dependent, unconditional KD methods are not appropriate. Conditional systems are more suitable in these cases and have been the subject of much interest.

Tumor-associated macrophages promote neuroblastoma via STAT3 phosphorylation and up-regulation of c-MYC.

Tumor-associated macrophages (TAMs) are strongly associated with poor survival in neuroblastomas that lack amplification. To study TAM action in neuroblastomas, we used a novel murine model of spontaneous neuroblastoma lacking amplification, and observed recruitment and polarization of TAMs, which in turn enhanced neuroblastoma proliferation and growth. In both murine and human neuroblastoma cells, we found that TAMs increased STAT3 activation in neuroblastoma cells and transcriptionally up-regulated the oncogene.

Contribution of neuroblastoma-derived exosomes to the production of pro-tumorigenic signals by bone marrow mesenchymal stromal cells.

The bone marrow (BM) niche is a microenvironment promoting survival, dormancy and therapeutic resistance in tumor cells. Central to this function are mesenchymal stromal cells (MSCs). Here, using neuroblastoma (NB) as a model, we demonstrate that NB cells release an extracellular vesicle (EVs) whose protein cargo is enriched in exosomal proteins but lacks cytokines and chemokines.

Sphingosine-1-Phosphate Receptor-1 Promotes Environment-Mediated and Acquired Chemoresistance.

Drug resistance is a major barrier for the development of effective and durable cancer therapies. Overcoming this challenge requires further defining the cellular and molecular mechanisms underlying drug resistance, both acquired and environment-mediated drug resistance (EMDR). Here, using neuroblastoma (NB), a childhood cancer with high incidence of recurrence due to resistance to chemotherapy, as a model we show that human bone marrow-mesenchymal stromal cells induce tumor expression of sphingosine-1-phosphate receptor-1 (S1PR1), leading to their resistance to chemotherapy.

Cancer-Associated Fibroblasts Share Characteristics and Protumorigenic Activity with Mesenchymal Stromal Cells.

Cancer-associated fibroblasts (CAF) have been suggested to originate from mesenchymal stromal cells (MSC), but their relationship with MSCs is not clear. Here, we have isolated from primary human neuroblastoma tumors a population of αFAP- and FSP-1-expressing CAFs that share phenotypic and functional characteristics with bone marrow-derived MSCs (BM-MSC). Analysis of human neuroblastoma tumors also confirmed the presence of αFAP- and FSP-1-positive cells in the tumor stroma, and their presence correlated with that of M2 tumor-associated macrophages.

Paternal Risk Factors for Oral Clefts in Northern Africans, Southeast Asians, and Central Americans.

While several studies have investigated maternal exposures as risk factors for oral clefts, few have examined paternal factors. We conducted an international multi-centered case-control study to better understand paternal risk exposures for oral clefts (cases = 392 and controls = 234). Participants were recruited from local hospitals and oral cleft repair surgical missions in Vietnam, the Philippines, Honduras, and Morocco.

The Tumor Microenvironment at a Turning Point Knowledge Gained Over the Last Decade, and Challenges and Opportunities Ahead: A White Paper from the NCI TME Network.

Over the past 10 years, the Tumor Microenvironment Network (TMEN), supported by the NCI (Bethesda, MD), has promoted collaborative research with the explicit goal of fostering multi-institutional and transdisciplinary groups that are capable of addressing complex issues involving the tumor microenvironment. The main goal of the TMEN was to generate novel information about the dynamic complexity of tumor-host interactions in different organ systems with emphasis on using human tissues and supplemented by experimental models. As this initiative comes to a close, members of the TMEN took time to examine what has been accomplished by the Network and importantly to identify the challenges and opportunities ahead.

More than the genes, the tumor microenvironment in neuroblastoma.

Neuroblastoma is the second most common solid tumor in children. Since the seminal discovery of the role of amplification of the MYCN oncogene in the pathogenesis of neuroblastoma in the 1980s, much focus has been on the contribution of genetic alterations in the progression of this cancer. However it is now clear that not only genetic events play a role but that the tumor microenvironment (TME) substantially contributes to the biology of neuroblastoma.

Small Molecule Inhibitors of Plasminogen Activator Inhibitor-1 Elicit Anti-Tumorigenic and Anti-Angiogenic Activity.

Numerous studies have shown a paradoxical positive correlation between elevated levels of plasminogen activator inhibitior-1 (PAI-1) in tumors and blood of cancer patients with poor clinical outcome, suggesting that PAI-1 could be a therapeutic target. Here we tested two orally bioavailable small molecule inhibitors of PAI-1 (TM5275 and TM5441) for their efficacy in pre-clinical models of cancer. We demonstrated that these inhibitors decreased cell viability in several human cancer cell lines with an IC50 in the 9.

Plasminogen Activator Inhibitor-1 in Cancer: Rationale and Insight for Future Therapeutic Testing.

Despite its function as an inhibitor of urokinase and tissue-type plasminogen activator (PA), PA inhibitor-1 (PAI-1) has a paradoxical protumorigenic role in cancer, promoting angiogenesis and tumor cell survival. In this review, we summarize preclinical evidence in support of the protumorigenic function of PAI-1 that has led to the testing of small-molecule PAI-1 inhibitors, initially developed as antithrombotic agents, in animal models of cancer. The review discusses the challenges and the opportunities that lay ahead to the development of efficacious and nontoxic PAI-1 inhibitors as anticancer agents.

Interaction between bone marrow stromal cells and neuroblastoma cells leads to a VEGFA-mediated osteoblastogenesis.

The potential role of osteoblasts in bone and bone marrow (BM) metastases in neuroblastoma (NBL) remains unclear. In this study, we examined the effect of NBL cells on the osteoblastic differentiation of BM-derived mesenchymal stromal cells (BMMSC). We show that the presence of NBL cells enhanced the osteoblastic differentiation of BMMSC driven by bone morphogenetic protein (BMP)-4, in the absence of any effect on NBL cell proliferation.

MYCN-dependent expression of sulfatase-2 regulates neuroblastoma cell survival.

Heparan sulfate proteoglycans (HSPG) play a critical role in the interaction of tumor cells and their microenvironment. HSPG activity is dictated by sulfation patterns controlled by sulfotransferases, which add sulfate groups, and sulfatases (Sulf), which remove 6-O-sulfates. Here, we report altered expression of these enzymes in human neuroblastoma cells with higher levels of Sulf-2 expression, a specific feature of MYCN-amplified cells (MYCN-A cells) that represent a particularly aggressive subclass.

[Tumor microenvironment and therapeutic resistance process].

Over the last decade, it has become clear that cancer is not just a disease of the genes, and that the tumor microenvironment (TME) plays an important role in cancer progression. Interactions between tumor cells and the TME, made of the extracellular matrix (ECM) and of non-transformed cells (designated here as stromal cells), promote cancer cell survival and drug resistance. Many of the mechanisms involved are known and are either contact-dependent or contact-independent.