Pre-metastatic Niche Research Articles

Vaping and tumor metastasis: current insights and progress.

Tumor metastasis is the primary cause of cancer-related mortality and remains a major hurdle in cancer treatment. Traditional cigarette smoking has been extensively studied for its role in promoting metastasis. However, the impact of e-cigarette (e-cig) on cancer metastasis is not well understood despite their increasing popularity as a supposedly safer alternative. This mini review synthesizes current literature on the effects of e-cig on cancer metastasis, focusing on the processes of dissemination, dormancy, and colonization. It also incorporates recent findings from our laboratory regarding the role of e-cig in tumor progression. E-cig exposure enhances metastatic potential through various mechanisms: it induces epithelial-mesenchymal transition (EMT), increasing cell migratory and invasive capabilities; promotes lymphangiogenesis, aiding tumor cell spread; and alters the pre-metastatic niche to support dormant tumor cells, enhancing their reactivation and colonization. Furthermore, e-cig induce significant epigenetic changes, such as DNA methylation and histone modifications, which regulate genes involved in metastasis. Our data suggest that e-cig upregulate histone demethylases like KDM6B in macrophages, impacting the TME and promoting metastasis. These findings underscore the need for further research to understand the long-term health implications of e-cig use and inform public health policies to reduce e-cig use.

Pre-metastatic niche in oral squamous cell carcinoma: Insights from a transcriptomic meta-analysis

Primary tumors can precondition a pre-metastatic niche, promoting the colonization of circulating neoplastic cells and influencing the secondary tumor microenvironment. Nevertheless, the mechanisms underlying the formation of this niche, as well as perineural invasion in oral squamous cell carcinoma (OSCC), are not well-elucidated. The study aims to identify differentially expressed genes (DEGs) and related pathways associated with pre-metastatic niche and perineural invasion in OSCC. We evaluated metastatic and non-metastatic primary tumor samples, healthy oral tissues, OSCC samples, metastatic lymph nodes from patients with OSCC, and normal lymph node samples. The GEO2R tool was applied to identify mRNAs differentially expressed between tissues exhibiting features of a pre-metastatic niche and normal tissue samples, including selected non-metastatic and metastatic OSCC samples. We also performed an analysis of perineural invasion-negative and perineural invasion-positive tumor samples. Our data revealed that SERPINE1, SPP1, CALCA, and MMP13 genes were upregulated. These upregulated genes are associated with several cancer-related pathways, while downregulated genes are mainly associated with immune responses, axon guidance, and the neurotrophin signaling pathway. Given the upregulation of the circadian rhythm pathway in metastatic lymph nodes, we also performed a correlation analysis that allows users to compute function-specific parameters, with resulting figures dynamically displayed to conveniently access the tumor’s immunological, clinical, and genomic features. Downregulation of the circadian rhythm gene PER3 and upregulation of Bhlhe40 were associated with poor survival outcomes. In conclusion, we postulate that during lymph node invasion, OSCCs activate axonal guidance genes, such as SERPENE1, L1AM, CXCR4, and SPP1. As neoplastic cells establish themselves, circadian rhythm genes are upregulated, contributing to immune evasion and promoting tumor growth.

Abstract IA016: Vascular control of metastasis

Abstract The molecular analysis of tumor vessel interactions during tumor progression and metastasis has primarily focused on the study of tumor cell-derived angiogenic and lymphangiogenic signals with the purpose to exploit such factors as therapeutic targets. Angiogenic factors activate endothelial cells in nearby blood and lymphatic capillaries to sprout towards the tumor. Tumor angiogenesis thereby not only nourishes the growing tumor, but access to the blood and lymphatic vasculatures enables cells from the primary tumor to enter the circulation to eventually form metastases at distant sites. The complex cellular interactions between tumor cells and endothelial cells have in this context mostly been studied from a tumor cell-centric perspective, i.e., the tumor cells send signals to which endothelial cells merely respond. Yet, the past decade has witnessed a fundamental change of paradigm with the discovery that the vascular endothelium does not just respond to exogenous cytokines but exerts active ’angiocrine’ gatekeeper roles controlling their microenvironment in an instructive manner. We have applied the concepts of angiocrine signaling towards the study of tumor progression and metastasis. Employing novel surgical preclinical metastasis models that better mimic tumor progression and the response to therapy as it occurs in humans, we have molecularly dissected vascular endothelial cells in progressing primary tumors as well as in the pre-metastatic and metastatic niches. These experiments were on the one hand aimed at establishing the systems map of endothelial transcriptomic changes during tumor progression and metastasis and on the other hand to identify and validate novel therapeutic targets. This presentation will review recent advances in the field of tumor microenvironment research and present novel angiocrine signaling mechanisms as promising targets of future mechanism-driven anti-metastatic therapy. Citation Format: Hellmut G. Augustin. Vascular control of metastasis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr IA016.

Tumor-derived EBV-miR-BART2-5p promotes nasopharyngeal carcinoma metastasis by inducing pre-metastatic endothelial cell pyroptosis.

Extravasation is a key step in tumor metastasis. Epstein‒Barr virus (EBV) plays a crucial role in nasopharyngeal carcinoma (NPC) metastasis. However, the functions and molecular mechanisms of EBV during tumor cell extravasation remains unclear. Here, we showed that the expression of pyroptosis-associated proteins is greater in the endothelial cells of metastatic NPC tissues than in those of nontumor tissues Exosomes derived from NPC cells promoted endothelial cell pyroptosis, vascular permeability, and tumor cell extravasation. Moreover, we found that BART2-5p is abundant in serum exosomes from NPC patients with metastasis and NPC cells, and that it regulates endothelial cell pyroptosis in pre-metastatic organs via MRE11A. Exosomes containing a BART2-5p inhibitor and AAV-MRE11A attenuated endothelial cell pyroptosis and tumor metastasis. Moreover, in the endothelial cells of metastatic tissues from NPC patients, the BART2-5p level was positively associated with pyroptosis-related protein expression. Collectively, our findings suggest that exosomal BART2-5p is involved in pre-metastatic niche formation, identifying secreted BART2-5p as a potential therapeutic target for NPC metastasis. Implications: The finding that secreted BART2-5p is involved in pre-metastatic niche formation may aid the development of potential therapeutic target for NPC metastasis.

A Multicellular Mechanochemical Model to Investigate Tumor Microenvironment Remodeling and Pre-Metastatic Niche Formation

A Multicellular Mechanochemical Model to Investigate Tumor Microenvironment Remodeling and Pre-Metastatic Niche Formation

LSD1 deficiency in breast cancer cells promotes the formation of pre-metastatic niches

Lysine-specific demethylase 1 (LSD1), a histone demethylating enzyme, plays a crucial role in cancer metastasis. Studies show LSD1 knockout promotes breast cancer lung metastasis, but it’s unknown if it alters the lung microenvironment for metastasis. In this study, we investigated the effects of exosomes from LSD1-knockdown (LSD1 KD) breast cancer cells on pre-metastatic niche formation. Injecting exosomes from LSD1 KD cells in mice resulted in a substantial increase in lung colonization by breast cancer cells, while treatment with exosomes derived from LSD1 KD cells decreased the expression of the ZO-1 and occludin, leading to increased vascular permeability. The LSD1 KD reduced the expression of circDOCK1, which augmented the levels of miR-1270 in exosomes. And miR-1270 inhibited ZO-1 expression in human endothelial cells, which enhanced their permeability. Our study uncovered a novel mechanism in which the LSD1 promotes the formation of pre-metastatic niches via the regulation of exosomal miRNA.

Targeting SPP1-orchestrated neutrophil extracellular traps-dominant pre-metastatic niche reduced HCC lung metastasis

BackgroundThe mechanisms by which tumor-derived factors remodel the microenvironment of target organs to facilitate cancer metastasis, especially organ-specific metastasis, remains obscure. Our previous studies have demonstrated that SPP1 plays a key role in promoting metastasis of hepatocellular carcinoma (HCC). However, the functional roles and mechanisms of tumor-derived SPP1 in shaping the pre-metastatic niche (PMN) and promoting lung-specific metastasis are unclear.MethodsOrthotopic metastasis models, experimental metastasis models, CyTOF and flow cytometry were conducted to explore the function of SPP1 in shaping neutrophil-dominant PMN and promoting HCC lung metastasis. The main source of CXCL1 in lung tissues was investigated via fluorescence activated cell sorting and immunofluorescence staining. The expression of neutrophils and neutrophil extracellular traps (NETs) markers was detected in the lung metastatic lesions of HCC patients and mouse lung specimens. The therapeutic significance was explored via in vivo DNase I and CXCR2 inhibitor assays.ResultsSPP1 promoted HCC lung colonization and metastasis by modifying pulmonary PMN in various murine models, and plasma SPP1 levels were closely associated with lung metastasis in HCC patients. Mechanistically, SPP1 binded to CD44 on lung alveolar epithelial cells to produce CXCL1, thereby attracting and forming neutrophil-abundant PMN in the lung. The recruited neutrophils were activated by SPP1 and then formed NETs-dominant PMN to trap the disseminated tumor cells and promote metastatic colonization. Moreover, early intervention of SPP1-orchestrated PMN by co-targeting the CXCL1-CXCR2 axis and NETs formation could efficiently inhibit the lung metastasis of HCC.ConclusionsOur study illustrates that HCC-lung host cell-neutrophil interactions play important roles in PMN formation and SPP1-induced HCC lung metastasis. Early intervention in SPP1-orchestrated PMN via CXCR2 inhibitor and DNase I is a potential therapeutic strategy to combat HCC lung metastasis.

Immunological Pre-Metastatic Niche in Dogs With Naturally Occurring Osteosarcoma.

Pre-metastatic niche (PMN) formation is essential for metastatic development and drives organotropism. Tumour-derived extracellular vesicles and soluble factors remodel the microenvironment of distant metastatic organs before subsequent metastasis. Dogs with osteosarcoma (OS) have proven to be excellent disease models for their human companions. Here, we show evidence of PMN formation in dogs with OS before metastasis. We necropsied and sampled lung tissues from dogs with naturally occurring treatment-naïve OS (n = 15) and control dogs without cancer (n = 10). We further divided dogs with OS into those having lung metastases (n = 5) and those without (n = 10). We stained formalin-fixed paraffin-embedded tissues using multiplex immunofluorescence to quantify the number of bone marrow-derived cells, monocytes and macrophages in the lung samples from each dog. The numbers of CD204+ macrophages, CD206+ macrophages and monocytes and CD11d+ bone marrow-derived cells (BMDCs) were significantly higher in the pre-metastatic lung of dogs with OS (n = 10) than in control dogs without cancer (n = 10). Furthermore, the total nucleated cell (DAPI+) density was higher before metastasis than in healthy lungs. In dogs with established metastases, the number of CD11d+ BMDCs was significantly lower than in the pre-metastatic lung, suggesting this recruitment is transient. Our study provides evidence of PMN existence in a naturally occurring cancer model similar to those observed in pre-clinical murine models. BMDCs are recruited to the lungs before metastases have developed. Dogs with OS may represent ideal candidates for assessing new PMN-targeting therapies.

Granulocyte-like myeloid-derived suppressor cells: The culprits of neutrophil extracellular traps formation in the pre-metastatic niche

Neutrophil extracellular traps (NETs) have been shown to exhibit chemotactic effects on circulating tumor cells at metastatic sites, promoting the progression of colorectal cancer liver metastasis (CRLM). However, the origin and factors contributing to the formation of NETs (NETosis) in the pre-metastatic niche (PMN) of target organs remain unclear. In this study, we investigated the relationship between phosphatase of regenerating liver-3 (PRL-3), myeloid-derived suppressor cells (MDSCs), neutrophils, and NETs through a retrospective clinical cohort study and a mouse model of CRLM. Our clinical findings revealed associations between PRL-3 expression and the infiltration of neutrophils and MDSCs in CRLM patients. Moreover, NETosis emerged as a robust indicator of poor prognosis for overall survival in these patients. In CRLM models, PRL-3 overexpression enhanced both primary tumor growth and liver metastasis. We found that the first week after tumor cell implantation appeared to be a crucial period for PMN formation, with notable infiltration of neutrophils, MDSCs, and NETosis during this time. Notably, co-culturing neutrophils with MDSCs induced NETosis in vitro, particularly with granulocyte-like MDSCs (Gr-MDSCs), the predominant subtype of infiltrating MDSCs. In summary, our findings elucidate the likely origin of NETs in the PMN during CRLM development and underscore the significant influence of PRL-3. These findings may offer potential immunotherapeutic targets for patients at risk of developing CRLM, warranting further investigation in clinical settings.

Ephrin A1 Stimulates CCL2 Secretion to Facilitate Pre-metastatic Niche Formation and Promote Gastric Cancer Liver Metastasis.

The liver is a primary target for distal metastasis of gastric cancer (GC). The hepatic pre-metastatic niche (PMN) facilitates crucial communications between primary tumor and liver, thereby playing an essential role in hepatic metastasis. Identification of the molecular mechanisms driving PMN formation in GC could facilitate development of strategies to prevent and treat liver metastasis. Here, we uncovered a role for ephrin A1 (EFNA1) signaling in development of the PMN. EFNA1 overexpression in GC cells significantly increased CCL2 secretion through the Hippo-YAP pathway. Secreted CCL2 activated hepatic stellate cells (HStCs) within the hepatic PMN via the WNT/β-catenin pathway. Inhibition of CCL2 significantly suppressed HStC activation and reduced liver metastasis triggered by EFNA1 signaling in GC cells. Moreover, high CCL2 expression correlated with poor survival in GC patients. Overall, these findings reveal that EFNA1 signaling in GC cells upregulates CCL2, which activates HStCs to engender establishment of a hepatic pre-metastatic niche that supports liver metastasis.

Lung bronchiectasisas a paradigm of the interplay between infection and colonization on plastic modulation of the pre-metastatic niche.

The lungs are most often a preferential target organ for malignant spreading and growth. It is well known that chronic parenchymal inflammation and prolonged injuries represents an independent risk factor for cancer onset. Growing evidence supports the implication of lung microbiota in the pathogenesis of lung cancer. However, the full interplay between chronic inflammation, bacterial colonization, pathologic condition as bronchiectasis and malignant growth deserves better clarification. We here aim at presenting and analyzing original data and discussing the state-of-the-art on the knowledge regarding how this complex milieu acts on the plasticity of the lung pre-metastatic niche to point out the rationale for early diagnosis and therapeutic targeting.

C-Reactive Protein Induces Immunosuppression by Activating FcγR2B in Pulmonary Macrophages to Promote Lung Metastasis.

C-reactive protein (CRP) is a liver-derived acute phase reactant that is a clinical marker of inflammation associated with poor cancer prognosis. Elevated CRP levels are observed in many types of cancer and are associated with significantly increased risk of metastasis, suggesting that CRP could have pro-metastatic actions. Here, we reported that CRP promotes lung metastasis by dampening the anti-cancer capacity of pulmonary macrophages in breast cancer and melanoma. Deletion of CRP in mice inhibited lung metastasis of breast cancer and melanoma cells without significantly impacting tumor growth compared to wildtype mice. In addition, the lungs of CRP deficient mice were enriched for activated pulmonary macrophages, which could be reduced to the level of wildtype mice by systemic administration of human CRP. Mechanistically, CRP blocked the activation of pulmonary macrophages induced by commensal bacteria in a FcγR2B-dependent manner, thereby impairing macrophage-mediated immune surveillance to promote the formation of a pre-metastatic niche in the lungs of tumor-bearing mice. Accordingly, treatment with specific CRP inhibitors activated pulmonary macrophages and attenuated lung metastasis in vivo. These findings highlight the importance of CRP in lung metastasis, which may represent an effective therapeutic target for patients with advanced solid cancers in clinics.

Tumour cell-released autophagosomes promote lung metastasis by upregulating PD-L1 expression in pulmonary vascular endothelial cells in breast cancer.

Establishing an immunosuppressive premetastatic niche (PMN) in distant organs is crucial for breast cancer metastasis. Vascular endothelial cells (VECs) act as barriers to transendothelial cell migration. However, the immune functions of PMNs remain unclear. Tumour cell-released autophagosomes (TRAPs) are critical modulators of antitumour immune responses. Herein, we investigated the mechanism through which TRAPs modulate the immune function of pulmonary VECs in lung PMN in breast cancer. Immortalised mouse pulmonary microvascular endothelial cells were incubated with TRAPs in vitro. RNA sequencing, flow cytometry, and western blotting were employed to assess immunosuppressive function and mechanism. In vivo, TRAP-trained and autophagy-deficient tumour mice were used to detect immunosuppression, and high-mobility group box 1 (HMGB1)-deficient TRAP-trained and TLR4 knockout mice were utilised to investigate the underlying mechanisms of pulmonary VECs. Additionally, the efficacy of anti-programmed cell death ligand-1 (PD-L1) immunotherapy was evaluated in early tumour-bearing mice. HMGB1 on TRAPs surfaces stimulated VECs to upregulate PD-L1 via a TLR4-MyD88-p38/STAT3 signalling cascade that depended on the cytoskeletal movement of VECs. Importantly, PD-L1 on TRAP-induced VECs can inhibit T cell function, promote lung PMN immunosuppression, and result in more pronounced lung metastasis. Treatment with anti-PD-L1 reduces lung metastasis in early stage tumour-bearing mice. These findings revealed a novel role and mechanism of TRAP-induced immunosuppression of pulmonary VECs in lung PMN. TRAPs and their surface HMGB1 are important therapeutic targets for reversing immunosuppression, providing a new theoretical basis for the treatment of early stage breast cancer using an anti-PD-L1 antibody.

Jiedu recipe, a compound Chinese herbal medicine, suppresses hepatocellular carcinoma metastasis by inhibiting the release of tumor-derived exosomes in a hypoxic microenvironment

ObjectiveTumor-derived exosomes (TDEs) play crucial roles in intercellular communication. Hypoxia in the tumor microenvironment enhances secretion of TDEs and accelerates tumor metastasis. Jiedu recipe (JR), a traditional Chinese medicinal formula, has demonstrated efficacy in preventing the metastasis of hepatocellular carcinoma (HCC). However, the underlying mechanism remains largely unknown. MethodsAnimal experiments were performed to investigate the metastasis-preventing effects of JR. Bioinformatics analysis and in vitro assays were conducted to explore the potential targets and active components of JR. TDEs were assessed using nanoparticle tracking analysis (NTA) and Western blotting (WB). Exosomes derived from normoxic or hypoxic HCC cells (H-TDEs) were collected to establish premetastatic mouse models. JR was intragastrically administered to evaluate its metastasis-preventive effects. WB and lysosomal staining were performed to investigate the effects of JR on lysosomal function and autophagy. Bioinformatics analysis, WB, NTA, and immunofluorescence staining were used to identify the active components and potential targets of JR. ResultsJR effectively inhibited subcutaneous-tumor-promoted lung premetastatic niche development and tumor metastasis. It inhibited the release of exosomes from tumor cells under hypoxic condition. JR treatment promoted both lysosomal acidification and suppressed secretory autophagy, which were dysregulated in hypoxic tumor cells. Quercetin was identified as the active component in JR, and the epidermal growth factor receptor (EGFR) was identified as a potential target. Quercetin inhibited EGFR phosphorylation and promoted the nuclear translocation of transcription factor EB (TFEB). Hypoxia-impaired lysosomal function was restored, and secretory autophagy was alleviated by quercetin treatment. ConclusionJR suppressed HCC metastasis by inhibiting hypoxia-stimulated exosome release, restoring lysosomal function, and suppressing secretory autophagy. Quercetin acted as a key component of JR and regulated TDE release through EGFR-TFEB signaling. Our study provides a potential strategy for retarding tumor metastasis by targeting H-TDE secretion.Please cite this article as: Jia WT, Xiang S, Zhang JB, Yuan JY, Wang YQ, Liang SF, Lin WF, Zhai XF, Shang Y, Ling CQ, Cheng BB. Jiedu recipe, a compound Chinese herbal medicine, suppresses hepatocellular carcinoma metastasis by inhibiting the release of tumor-derived exosomes in a hypoxic microenvironment through the EGFR-TFEB signaling pathway. J Integr Med. 2024; Epub ahead of print.

Integrin α6-containing extracellular vesicles promote lymphatic remodelling for pre-metastatic niche formation in lymph nodes via interplay with CD151.

Heterogeneous extracellular vesicles (EVs) from various types of tumours are acknowledged for inducing the formation of pre-metastatic "niches" in draining lymph nodes (LNs) to promote lymphatic metastasis. In order to identify the specific subpopulations of EVs involved, we performed high-resolution proteomic analysis combined with nanoflow cytometry of bladder cancer (BCa) tissue-derived EVs to identify a novel subset of tumour-derived EVs that contain integrin α6 (ITGA6+EVs) and revealed the positive correlation of ITGA6+EVs with the formation of pre-metastatic niche in draining LNs and lymphatic metastasis in multicentre clinical analysis of 820-case BCa patients. BCa-derived ITGA6+EVs induced E-selectin (SELE)-marked lymphatic remodelling pre-metastatic niche and promoted metastasis in draining LNs through delivering cargo circRNA-LIPAR to lymphatic endothelial cells in vivo and in vitro. Mechanistically, LIPAR linked ITGA6 to the switch II domain of RAB5A and sustained RAB5A GTP-bound activated state, thus maintaining the production of ITGA6+EVs loaded with LIPAR through endosomal trafficking. ITGA6+EVs targeted lymphatic vessels through ITGA6-CD151 interplay and released LIPAR to induce SELE overexpression-marked lymphatic remodelling pre-metastatic niche. Importantly, we constructed engineered-ITGA6 EVs to inhibit lymphatic pre-metastatic niche, which suppressed lymphatic metastasis and prolonged survival in preclinical models. Collectively, our study uncovers the mechanism of BCa-derived ITGA6+EVs mediating pre-metastatic niche and provides an engineered-EV-based strategy against BCa lymphatic metastasis.

Bioresponsive Nanoparticles Boost Starvation Therapy and Prevent Premetastatic Niche Formation for Pulmonary Metastasis Treatment.

In the process of tumor metastasis, tumor cells can acquire invasion by excessive uptake of nutrients and energy and interact with the host microenvironment to shape a premetastatic niche (PMN) that facilitates their colonization and progression in the distal sites. Pyruvate is an essential nutrient that engages in both energy metabolism and remodeling of the extracellular matrix (ECM) in the lungs for PMN formation, thus providing a target for tumor metastasis treatment. There is a paucity of strategies focusing on PMN prevention, which is key to metastasis inhibition. Here, we design a bioresponsive nanoparticle (HP/GU) based on a disulfide-cross-linked hyperbranched polyethylenimine (D-PEI) core and a hyaluronic acid (HA) shell with a reactive oxygen species (ROS)-sensitive cross-linker between them to encapsulate glucose oxidase (GOX) and a mitochondrial pyruvate carrier (MPC) inhibitor via electrostatic interaction, which reinforces starvation therapy and reduces PMN formation in the lungs via inhibiting pyruvate metabolism. In tumor cells, GOX and MPC inhibitors can be rapidly released and synergistically reduce the energy supply of tumor cells by consuming glucose and inhibiting pyruvate uptake to decrease tumor cell invasion. MPC inhibitors can also reduce ECM remodeling by blocking cellular pyruvate metabolism to prevent PMN formation. Consequently, HP/GU achieves an efficient inhibition of both primary and metastatic tumors and provides an innovative strategy for the treatment of tumor metastases.

Abstract A063: The role of JAK/STAT3 signaling in lung premetastatic niche formation and progression

Abstract Pancreatic ductal adenocarcinoma (PDA) is a highly deadly disease with a five-year survival rate of only 13%. Oncogenic KRAS (Kras) is a near-universal driver of PDA, and its activity is required for the formation of PDA precursor lesions, pancreatic intraepithelial neoplasia (PanIN). PanIN formation is accompanied by fibrosis and immune cell infiltration, creating a precursor tumor microenvironment that is maintained and evolves through disease progression. Our previous work has demonstrated that Kras in epithelial cells drives fibroblast activation during very early stages of pancreatic tumorigenesis and that fibroblast reprogramming occurs in a JAK/STAT3 signaling-dependent manner. Other groups have described that pancreatic cancer cells secrete specific factors to prime the liver and lung through immunomodulation and extracellular matrix remodeling, forming the premetastatic niche (PMN). Additionally, recent papers in the field have suggested a role for JAK/STAT3-reprogramming in the formation of the PMN during pancreatic tumorigenesis. Presently, it remains to be elucidated at what stage of PDA tumorigenesis the PMN is initiated, as well as the mechanisms underlying formation. In a pilot experiment, we induced pancreas specific Kras until mice had PanIN lesions but no overt malignancy. We then harvest the pancreas and lung for single cell RNA sequencing. Here, results showed that when Kras is expressed within the pancreas of a mouse, fibroblasts are reprogrammed within the lung, prior to pancreatic tumor formation. Additionally, data indicates JAK/STAT3 signaling activation present within these fibroblasts. We also used multiple lung metastasis models, both spontaneous and tail vein injection, to show persistence of this reprogramming. This suggests that pancreatic Kras expression can systemically reprogram fibroblasts to form a PMN more capable of seeding cancer cells. Our in vitro studies have showed similar results as lung fibroblasts treated with Kras ON conditioned media show increased pSTAT3. Furthermore, using multiplex ELISA on iKras conditioned media, we have identified multiple promising secreted factor hits that may play a role in lung fibroblast reprogramming. Ongoing studies aim to define the mechanism of pulmonary fibroblast reprogramming in the context of the PMN and to assess the role of pSTAT3+ fibroblast reprogramming on establishment of pulmonary metastases. Citation Format: Emily Lasse Opsahl, Carlos E Espinoza, Katelyn L Donahue, Ahmed M Elhossiny, Padma Kadiyala, Allison C Bischoff, Mary Poggi, Yaqing Zhang, Jiaqi Shi, Timothy Frankel, Marina Pasca di Magliano. The role of JAK/STAT3 signaling in lung premetastatic niche formation and progression [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A063.

Role of CD44-Positive Extracellular Vesicles Derived from Highly Metastatic Mouse Mammary Carcinoma Cells in Pre-Metastatic Niche Formation.

Malignant breast cancers pose a notable challenge when it comes to treatment options. Recently, research has implicated extracellular vesicles (EVs) secreted by cancer cells in the formation of a pre-metastatic niche. Small clumps of CD44-positive breast cancer cells are efficiently transferred through CD44-CD44 protein homophilic interaction. This study aims to examine the function of CD44-positive EVs in pre-metastatic niche formation in vitro and to suggest a more efficacious EV formulation. We used mouse mammary carcinoma cells, BJMC3879 Luc2 (Luc2 cells) as the source of CD44-positive EVs and mouse endothelial cells (UV2 cells) as the recipient cells in the niche. Luc2 cells exhibited an enhanced secretion of EVs expressing CD44 and endothelial growth factors (VEGF-A, -C) under 20% O2 (representative of the early stage of tumorigenesis) compared to its expression under 1% O2 (in solid tumor), indicating that pre-metastatic niche formation occurs in the early stage. Furthermore, UV2 endothelial cells expressing CD44 demonstrated a high level of engulfment of EVs that had been supplemented with hyaluronan, and the proliferation of UV2 cells occurred following the engulfment of EVs. These results suggest that anti-VEGF-A and -C encapsulated, CD44-expressing, and hyaluronan-coated EVs are more effective for tumor metastasis.

New Insights into the Exosome-Induced Migration of Uveal Melanoma Cells and the Pre-Metastatic Niche Formation in the Liver.

UM is an aggressive intraocular tumor characterized by high plasticity and a propensity to metastasize in the liver. However, the underlying mechanisms governing liver tropism remain poorly understood. Given the emerging significance of exosomes, we sought to investigate the contribution of UM-derived exosomes to specific steps of the metastatic process. Firstly, we isolated exosomes from UM cells sharing a common genetic background and different metastatic properties. A comparison of protein cargo reveals an overrepresentation of proteins related to cytoskeleton remodeling and actin filament-based movement in exosomes derived from the parental cells that may favor the detachment of cells from the primary site. Secondly, we assessed the role of macrophages in reprogramming the HHSCs by exosomes. The activation of HHSCs triggered a pro-inflammatory and pro-fibrotic environment through cytokine production, upregulation of extracellular matrix molecules, and the activation of signaling pathways. Finally, we found that activated HHSCs promote increased adhesion and migration of UM cells. Our findings shed light on the pivotal role of exosomes in pre-metastatic niche construction in the liver.

CircRNAs expression profile and potential roles of circRERE-PMN in pre-metastatic lungs.

The successful pulmonary metastasis of malignant cancer cells depends on the survival of circulating tumor cells in a distant and hostile microenvironment. The formation of a pre-metastatic niche (PMN) creates a supportive environment for subsequent metastasis. Circular RNAs (circRNAs) are increasingly acknowledged as crucial elements in the mechanisms of metastasis due to their stable structures and functions, making them promising early metastasis detection markers. However, the specific expression patterns and roles of circRNAs in the lungs before metastasis remain largely unexplored. Our research aims to chart the circRNA expression profile and assess their impact on the lung PMN. We developed a lung PMN model and employed comprehensive RNA sequencing to analyze the differences in circRNA expression between normal and pre-metastatic lungs. We identified 38 significantly different circRNAs, primarily involved in metabolism, apoptosis, and inflammation pathways. We then focused on one specific circRNA, circ:chr4:150406196 - 150406664 (circRERE-PMN), which exhibited a significant change in expression and was prevalent in myeloid-derived suppressor cells (MDSCs), alveolar epithelial cells, and macrophages within the pre-metastatic lung environment. CircRERE-PMN was found to potentially regulate apoptosis and the expression of cytokines and chemokines through its interaction with the downstream target HUR in alveolar epithelial cells. Overall, our study highlights the crucial role of circRNAs in the formation of lung PMNs, supporting their potential as diagnostic or therapeutic targets for lung metastasis.