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BACKGROUND: Interleukin-6 (IL-6) is a multifunctional cytokine that controls the immune response, and its role has been described in the development of autoimmune diseases. Signaling via its cognate IL-6 receptor (IL-6R) complex is critical in tumor progression and, therefore, IL-6R represents an important therapeutic target. METHODS: An albumin-binding domain-derived highly complex combinatorial library was used to select IL-6R alpha (IL-6Rα)-targeted small protein binders using ribosome display. Large-scale screening of bacterial lysates of individual clones was performed using ELISA, and their IL-6Rα blocking potential was verified by competition ELISA. The binding of proteins to cells was monitored by flow cytometry and confocal microscopy on HEK293T-transfected cells, and inhibition of signaling function was examined using HEK-Blue IL-6 reporter cells. Protein binding kinetics to living cells was measured by LigandTracer, cell proliferation and toxicity by iCELLigence and Incucyte, cell migration by the scratch wound healing assay, and prediction of binding poses using molecular modeling by docking. RESULTS: We demonstrated a collection of protein variants called NEF ligands, selected from an albumin-binding domain scaffold-derived combinatorial library, and showed their binding specificity to human IL-6Rα and antagonistic effect in HEK-Blue IL-6 reporter cells. The three most promising NEF108, NEF163, and NEF172 variants inhibited cell proliferation of malignant melanoma (G361 and A2058) and pancreatic (PaTu and MiaPaCa) cancer cells, and suppressed migration of malignant melanoma (A2058), pancreatic carcinoma (PaTu), and glioblastoma (GAMG) cells in vitro. The NEF binders also recognized maturation-induced IL-6Rα expression and interfered with IL-6-induced differentiation in primary human B cells. CONCLUSION: We report on the generation of small protein blockers of human IL-6Rα using directed evolution. NEF proteins represent a promising class of non-toxic anti-tumor agents with migrastatic potential.
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Movimiento Celular , Proliferación Celular , Receptores de Interleucina-6 , Humanos , Proliferación Celular/efectos de los fármacos , Receptores de Interleucina-6/metabolismo , Movimiento Celular/efectos de los fármacos , Células HEK293 , Línea Celular Tumoral , Unión Proteica/efectos de los fármacosRESUMEN
Breast cancer is the most frequently diagnosed cancer in women worldwide. Although dramatically increased survival rates of early diagnosed cases have been observed, late diagnosed patients and metastatic cancer may still be considered fatal. The present study's main focus was on cancerassociated fibroblasts (CAFs) which is an active component of the tumor microenvironment (TME) regulating the breast cancer ecosystem. Transcriptomic profiling and analysis of CAFs isolated from breast cancer skin metastasis, cutaneous basal cell carcinoma, and squamous cell carcinoma unravelled major gene candidates such as IL6, VEGFA and MFGE8 that induced coexpression of keratins8/14 in the EMG3 cell line derived from infiltrating ductal breast carcinoma. Western blot analysis of selected keratins (keratin8, 14, 18, 19) and epithelialmesenchymal transitionassociated markers (SLUG, SNAIL, ZEB1, E/Ncadherin, vimentin) revealed specific responses pointing to certain heterogeneity of the studied CAF populations. Experimental in vitro treatment using neutralizing antibodies against IL-6, VEGFA and MFGE8 attenuated the modulatory effect of CAFs on EMG3 cells. The present study provided novel data in characterizing and understanding the interactions between CAFs and EMG3 cells in vitro. CAFs of different origins support the proinflammatory microenvironment and influence the biology of breast cancer cells. This observation potentially holds significant interest for the development of novel, clinically relevant approaches targeting the TME in breast cancer. Furthermore, its implications extend beyond breast cancer and have the potential to impact a wide range of other cancer types.
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Neoplasias de la Mama , Fibroblastos Asociados al Cáncer , Femenino , Humanos , Antígenos de Superficie , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Fibroblastos Asociados al Cáncer/metabolismo , Línea Celular Tumoral , Fibroblastos/metabolismo , Queratinas/genética , Queratinas/metabolismo , Células MCF-7 , Proteínas de la Leche/genética , Proteínas de la Leche/metabolismo , Pronóstico , Transcriptoma , Microambiente Tumoral/genética , Melanoma Cutáneo MalignoRESUMEN
BACKGROUND: The aim of this prospective study was to evaluate the role of serum IL-6 as a potential predictive biomarker of postoperative complications (POC) in elective colorectal surgery. METHOD: A total of 115 patients underwent colorectal surgery for malignancy. IL-6 was measured on the first and third postoperative days (POD1, POD3), and C-reactive protein (CRP) was measured on the POD3. POC was analysed in subgroups according to ClavienâDindo (CD), antibiotic (ATB) treatment, intensive care unit (ICU) and hospital length of stay. The predictive power of variables for evaluated endpoints was analysed using receiver-operating characteristic (ROC) analysis and described by area under the curve (AUC). ROC analysis was adopted for the identification of optimal cut-offs. Histological analysis was performed to verify IL-6 production by the tumour. RESULTS: Out of 115 patients who were analysed, 42% had POC. Patients with POC had significantly higher serum levels of IL-6 on POD1 (p < 0.001) and POD3 (p < 0.001). IL-6 early on POD1 as a predictor of antibiotic treatment, ICU stay and hospital stay (AUC 0.818; 0.811; 0.771) did not significantly differ from the AUC of CRP late on POD3 (0.879; 0.838, 0.752). A cut-off IL-6 value of 113 pg/ml on POD1 and 180.5 pg/ml on POD3 in severe complications (CD > 3a) resulted in 75% and 72% sensitivity, 78.6% and 99% specificity, negative predictive value 96.4% and 97% and positive predictive value 29% and 88.9%. CONCLUSION: The serum level of interleukin-6 can predict severe (CD > 3a) POC early on POD1. On POD3, IL-6 is superior to CRP in terms of high positive predictive power of severe POC. Interestingly, the advantage of IL-6 on POD1 is early prediction of the need for antibiotic treatment, ICU stay and hospital stay, which is comparable to the CRP serum level late on the third POD.
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Cirugía Colorrectal , Interleucina-6 , Humanos , Antibacterianos , Biomarcadores , Proteína C-Reactiva/análisis , Complicaciones Posoperatorias/etiología , Estudios Prospectivos , Curva ROCRESUMEN
Soft tissue sarcomas are aggressive mesenchymal-origin malignancies. Undifferentiated pleomorphic sarcoma (UPS) belongs to the aggressive, high-grade, and least characterized sarcoma subtype, affecting multiple tissues and metastasizing to many organs. The treatment of localized UPS includes surgery in combination with radiation therapy. Metastatic forms are treated with chemotherapy. Immunotherapy is a promising treatment modality for many cancers. However, the development of immunotherapy for UPS is limited due to its heterogeneity, antigenic landscape variation, lower infiltration with immune cells, and a limited number of established patient-derived UPS cell lines for preclinical research. In this study, we established and characterized a novel patient-derived UPS cell line, JBT19. The JBT19 cells express PD-L1 and collagen, a ligand of the immune checkpoint molecule LAIR-1. JBT19 cells can form spheroids in vitro and solid tumors in immunodeficient nude mice. We found JBT19 cells induce expansion of JBT19-reactive autologous and allogeneic NK, T, and NKT-like cells, and the reactivity of the expanded cells was associated with cytotoxic impact on JBT19 cells. The PD-1 and LAIR-1 ligand-expressing JBT19 cells show ex vivo immunogenicity and effective in vivo xenoengraftment properties that can offer a unique resource in the preclinical research developing novel immunotherapeutic interventions in the treatment of UPS.
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Histiocitoma Fibroso Maligno , Sarcoma , Ratones , Animales , Humanos , Antígeno B7-H1/metabolismo , Ratones Desnudos , Ligandos , Sarcoma/patología , Inmunoterapia , Línea CelularRESUMEN
Identification of therapeutic targets for treating fibrotic diseases and cancer remains challenging. Our study aimed to investigate the effects of TGF-ß1 and TGF-ß3 on myofibroblast differentiation and extracellular matrix deposition in different types of fibroblasts, including normal/dermal, cancer-associated, and scar-derived fibroblasts. When comparing the phenotype and signaling pathways activation we observed extreme heterogeneity of studied markers across different fibroblast populations, even within those isolated from the same tissue. Specifically, the presence of myofibroblast and deposition of extracellular matrix were dependent on the origin of the fibroblasts and the type of treatment they received (TGF-ß1 vs. TGF-ß3). In parallel, we detected activation of canonical signaling (pSMAD2/3) across all studied fibroblasts, albeit to various extents. Treatment with TGF-ß1 and TGF-ß3 resulted in the activation of canonical and several non-canonical pathways, including AKT, ERK, and ROCK. Among studied cells, cancer-associated fibroblasts displayed the most heterogenic response to TGF-ß1/3 treatments. In general, TGF-ß1 demonstrated a more potent activation of signaling pathways compared to TGF-ß3, whereas TGF-ß3 exhibited rather an inhibitory effect in keloid- and hypertrophic scar-derived fibroblasts suggesting its clinical potential for scar treatment. In summary, our study has implications for comprehending the role of TGF-ß signaling in fibroblast biology, fibrotic diseases, and cancer. Future research should focus on unraveling the mechanisms beyond differential fibroblast responses to TGF-ß isomers considering inherent fibroblast heterogeneity.
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Cicatriz Hipertrófica , Factor de Crecimiento Transformador beta1 , Humanos , Factor de Crecimiento Transformador beta1/farmacología , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta3/metabolismo , Factor de Crecimiento Transformador beta3/farmacología , Fibroblastos/metabolismo , Cicatrización de Heridas , Cicatriz Hipertrófica/metabolismo , Cicatriz Hipertrófica/patología , Factor de Crecimiento Transformador beta/metabolismo , Carcinogénesis/metabolismo , Carcinogénesis/patología , Transformación Celular Neoplásica/metabolismo , Isoformas de Proteínas/metabolismo , Células CultivadasRESUMEN
Pentamethinium indolium salts are promising fluorescence probes and anticancer agents with high mitochondrial selectivity. We synthesized two indolium pentamethinium salts: a cyclic form with quinoxaline directly incorporated in the pentamethinium chain (cPMS) and an open form with quinoxaline substitution in the γ-position (oPMS). To better understand their properties, we studied their interaction with mitochondrial phospholipids (cardiolipin and phosphatidylcholine) by spectroscopic methods (UV-Vis, fluorescence, and NMR spectroscopy). Both compounds displayed significant affinity for cardiolipin and phosphatidylcholine, which was associated with a strong change in their UV-Vis spectra. Nevertheless, we surprisingly observed that fluorescence properties of cPMS changed in complex with both cardiolipin and phosphatidylcholine, whereas those of oPMS only changed in complex with cardiolipin. Both salts, especially cPMS, display high usability in mitochondrial imaging and are cytotoxic for cancer cells. The above clearly indicates that conjugates of pentamethinium and quinoxaline group, especially cPMS, represent promising structural motifs for designing mitochondrial-specific agents.
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Antineoplásicos , Cardiolipinas , Quinoxalinas/farmacología , Sales (Química) , Antineoplásicos/farmacología , Antineoplásicos/química , FosfatidilcolinasRESUMEN
Therapy for pancreatic ductal adenocarcinoma remains challenging, and the chances of a complete cure are very limited. As in other types of cancer, the expression and role of miRNAs in controlling the biological properties of this type of tumor have been extensively studied. A better insight into miRNA biology seems critical to refining diagnostics and improving their therapeutic potential. In this study, we focused on the expression of miR-21, -96, -196a, -210, and -217 in normal fibroblasts, cancer-associated fibroblasts prepared from a ductal adenocarcinoma of the pancreas, and pancreatic carcinoma cell lines. We compared these data with miRNAs in homogenates of paraffin-embedded sections from normal pancreatic tissues. In cancer-associated fibroblasts and cancer cell lines, miRNAs differed significantly from the normal tissue. In detail, miR-21 and -210 were significantly upregulated, while miR-217 was downregulated. Similar transcription profiles were earlier reported in cancer-associated fibroblasts exposed to hypoxia. However, the cells in our study were cultured under normoxic conditions. We also noted a relation to IL-6 production. In conclusion, cultured cancer-associated fibroblasts and carcinoma cells reflect miR-21 and -210 expression similarly to the cancer tissue samples harvested from the patients.
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Fibroblastos Asociados al Cáncer , Carcinoma Ductal Pancreático , MicroARNs , Neoplasias Pancreáticas , Humanos , MicroARNs/genética , Fibroblastos Asociados al Cáncer/metabolismo , Neoplasias Pancreáticas/patología , Carcinoma Ductal Pancreático/patología , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Neoplasias PancreáticasRESUMEN
Interleukin 6 (IL-6) belongs to a broad class of cytokines involved in the regulation of various homeostatic and pathological processes. These activities range from regulating embryonic development, wound healing and ageing, inflammation, and immunity, including COVID-19. In this review, we summarise the role of IL-6 signalling pathways in cancer biology, with particular emphasis on cancer cell invasiveness and metastasis formation. Targeting principal components of IL-6 signalling (e.g., IL-6Rs, gp130, STAT3, NF-κB) is an intensively studied approach in preclinical cancer research. It is of significant translational potential; numerous studies strongly imply the remarkable potential of IL-6 signalling inhibitors, especially in metastasis suppression.
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Antineoplásicos , Neoplasias , Humanos , Antineoplásicos/uso terapéutico , Interleucina-6/metabolismo , Neoplasias/tratamiento farmacológico , Transducción de SeñalRESUMEN
IL-6 signaling is involved in the pathogenesis of a number of serious diseases, including chronic inflammation and cancer. Targeting of IL-6 receptor (IL-6R) by small molecules is therefore an intensively studied strategy in cancer treatment. We describe the design, synthesis, and characteristics of two new bis-pentamethinium salts 5 and 6 (meta and para) bearing indole moieties. Molecular docking studies showed that both compounds have the potential to bind IL-6R (free energy of binding -9.5 and -8.1 kcal/mol). The interaction with IL-6R was confirmed using microscale thermophoresis analyses, which revealed that both compounds had strong affinity for the IL-6R (experimentally determined dissociation constants 26.5 ± 2.5 nM and 304 ± 27.6 nM, respectively). In addition, both compounds were cytotoxic for a broad spectrum of cancer cell lines in micromolar concentrations, most likely due to their accumulation in mitochondria and inhibition of mitochondrial respiration. In summary, the structure motif of bis-pentamethinium salts represents a promising starting point for the design of novel multitargeting compounds with the potential to inhibit IL-6 signaling and simultaneously target mitochondrial metabolism in cancer cells.
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Recent evidence indicates that targeting IL-6 provides broad therapeutic approaches to several diseases. In patients with cancer, autoimmune diseases, severe respiratory infections [e.g. coronavirus disease 2019 (COVID-19)] and wound healing, IL-6 plays a critical role in modulating the systemic and local microenvironment. Elevated serum levels of IL-6 interfere with the systemic immune response and are associated with disease progression and prognosis. As already noted, monoclonal antibodies blocking either IL-6 or binding of IL-6 to receptors have been used/tested successfully in the treatment of rheumatoid arthritis, many cancer types, and COVID-19. Therefore, in the present review, we compare the impact of IL-6 and anti-IL-6 therapy to demonstrate common (pathological) features of the studied diseases such as formation of granulation tissue with the presence of myofibroblasts and deposition of new extracellular matrix. We also discuss abnormal activation of other wound-healing-related pathways that have been implicated in autoimmune disorders, cancer or COVID-19.
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Enfermedades Autoinmunes , COVID-19 , Neoplasias , Humanos , Autoinmunidad , Inflamación , Enfermedades Autoinmunes/tratamiento farmacológico , Neoplasias/tratamiento farmacológico , Cicatrización de Heridas , Microambiente TumoralRESUMEN
Cancer-associated fibroblasts (CAFs) are an essential component of the tumour microenvironment. They represent a heterogeneous group of cells that are under the control of cancer cells and can reversely influence the cancer cell population. They affect the cancer cell differentiation status, and the migration and formation of metastases. This is achieved through the production of the extracellular matrix and numerous bioactive factors. IL-6 seems to play the central role in the communication of noncancerous and cancer cells in the tumour. This review outlines the role of exosomes in cancer cells and cancer-associated fibroblasts. Available data on the exosomal cargo, which can significantly intensify interactions in the tumour, are summarised. The role of exosomes as mediators of the dialogue between cancer cells and cancer-associated fibroblasts is discussed together with their therapeutic relevance. The functional unity of the paracrine- and exosome-mediated communication of cancer cells with the tumour microenvironment represented by CAFs is worthy of attention.
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Fibroblastos Asociados al Cáncer/metabolismo , Exosomas/metabolismo , Interleucina-6/metabolismo , Neoplasias/metabolismo , Movimiento Celular , Proliferación Celular , Regulación Neoplásica de la Expresión Génica , Humanos , Comunicación Paracrina , Microambiente TumoralRESUMEN
Vestibular schwannoma is the most common benign neoplasm of the cerebellopontine angle. It arises from Schwann cells of the vestibular nerve. The first symptoms of vestibular schwannoma include hearing loss, tinnitus, and vestibular symptoms. In the event of further growth, cerebellar and brainstem symptoms, along with palsy of the adjacent cranial nerves, may be present. Although hearing impairment is present in 95% of patients diagnosed with vestibular schwannoma, most tumors do not progress in size or have low growth rates. However, the clinical picture has unpredictable dynamics, and there are currently no reliable predictors of the tumor's behavior. The etiology of the hearing loss in patients with vestibular schwannoma is unclear. Given the presence of hearing loss in patients with non-growing tumors, a purely mechanistic approach is insufficient. A possible explanation for this may be that the function of the auditory system may be affected by the paracrine activity of the tumor. Moreover, initiation of the development and growth progression of vestibular schwannomas is not yet clearly understood. Biallelic loss of the NF2 gene does not explain the occurrence in all patients; therefore, detection of gene expression abnormalities in cases of progressive growth is required. As in other areas of cancer research, the tumor microenvironment is coming to the forefront, also in vestibular schwannomas. In the paradigm of the tumor microenvironment, the stroma of the tumor actively influences the tumor's behavior. However, research in the area of vestibular schwannomas is at an early stage. Thus, knowledge of the molecular mechanisms of tumorigenesis and interactions between cells present within the tumor is crucial for the diagnosis, prediction of tumor behavior, and targeted therapeutic interventions. In this review, we provide an overview of the current knowledge in the field of molecular biology and tumor microenvironment of vestibular schwannomas, as well as their relationship to tumor growth and hearing loss.
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The incidence of cutaneous malignant melanoma is increasing worldwide. While the treatment of initial stages of the disease is simple, the advanced disease frequently remains fatal despite novel therapeutic options . This requires identification of novel therapeutic targets in melanoma. Similarly to other types of tumours, the cancer microenvironment plays a prominent role and determines the biological properties of melanoma. Importantly, melanoma cell-produced exosomes represent an important tool of intercellular communication within this cancer ecosystem. We have focused on potential differences in the activity of exosomes produced by melanoma cells towards melanoma-associated fibroblasts and normal dermal fibroblasts. Cancer-associated fibroblasts were activated by the melanoma cell-produced exosomes significantly more than their normal counterparts, as assessed by increased transcription of genes for inflammation-supporting cytokines and chemokines, namely IL-6 or IL-8. We have observed that the response is dependent on the duration of the stimulus via exosomes and also on the quantity of exosomes. Our study demonstrates that melanoma-produced exosomes significantly stimulate the tumour-promoting proinflammatory activity of cancer-associated fibroblasts. This may represent a potential new target of oncologic therapy .
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Exosomas/metabolismo , Fibroblastos/metabolismo , Melanoma Experimental/metabolismo , Fibroblastos/patología , Humanos , Melanoma Experimental/patología , Células Tumorales CultivadasRESUMEN
Interleukin-6 (IL-6) is a highly potent cytokine involved in multiple biological processes. It was previously reported to play a distinct role in inflammation, autoimmune and psychiatric disorders, ageing and various types of cancer. Furthermore, it is understood that IL-6 and its signaling pathways are substantial players in orchestrating the cancer microenvironment. Thus, they appear to be potential targets in anti-tumor therapy. The aim of this article is to elucidate the role of IL-6 in the tumor ecosystem and to review the possible therapeutic approaches in head and neck cancer.
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Neoplasias de Cabeza y Cuello/inmunología , Interleucina-6/inmunología , Microambiente Tumoral , Neoplasias de Cabeza y Cuello/terapia , Humanos , Inflamación , Interleucina-6/metabolismo , Transducción de SeñalRESUMEN
Therapy targeting immune checkpoints represents an integral part of the treatment for patients suffering from advanced melanoma. However, the mechanisms of resistance are responsible for a lower therapeutic outcome than expected. Concerning melanoma, insufficient stimulation of the immune system by tumour neoantigens is a likely explanation. As shown previously, radiotherapy is a known option for increasing the production of tumour neoantigens and their release into the microenvironment. Consequently, neoantigens could be recognized by antigen presenting cells (APCs) and subjected to effector T lymphocytes. Enhancing the immune reaction can trigger the therapeutic response also at distant metastases, a phenomenon known as an abscopal effect (from "ab scopus", that is, away from the target). To illustrate this, we present the case of a 78-year old male treated by anti-CTLA-4/ipilimumab for metastatic melanoma. The patient received the standard four doses of ipilimumab administered every three weeks. However, the control CT scans detected disease progression in the form of axillary lymph nodes metastasis and liver metastasis two months after ipilimumab. At this stage, palliative cryotherapy of the skin metastases was initiated to alleviate the tumour burden. Surprisingly, the effect of cryotherapy was also observed in untreated metastases and deep subcutaneous metastases on the back. Moreover, we observed the disease remission of axillary lymph nodes and liver metastasis two months after the cryotherapy. The rarity of the abscopal effect suggests that even primed anti-tumour CD8+ T cells cannot overcome the tumour microenvironment's suppressive effect and execute immune clearance. However, the biological mechanism underlying this phenomenon is yet to be elucidated. The elicitation of a systemic response by cryotherapy with documented abscopal effect was rarely reported, although the immune response induction is presumably similar to a radiotherapy-induced one. The report is a combination case study and review of the abscopal effect in melanoma treated with checkpoint inhibitors.
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Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Ipilimumab/uso terapéutico , Melanoma/secundario , Melanoma/terapia , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/terapia , Anciano , Células Presentadoras de Antígenos/inmunología , Antígenos de Neoplasias/metabolismo , Crioterapia , Humanos , Masculino , Melanoma/inmunología , Modelos Inmunológicos , Cuidados Paliativos , Neoplasias Cutáneas/inmunología , Resultado del Tratamiento , Microambiente Tumoral/inmunologíaRESUMEN
COVID-19 is a pandemic respiratory disease caused by the SARS-CoV-2 coronavirus. The worldwide epidemiologic data showed higher mortality in males compared to females, suggesting a hypothesis about the protective effect of estrogens against severe disease progression with the ultimate end being patient's death. This article summarizes the current knowledge regarding the potential effect of estrogens and other modulators of estrogen receptors on COVID-19. While estrogen receptor activation shows complex effects on the patient's organism, such as an influence on the cardiovascular/pulmonary/immune system which includes lower production of cytokines responsible for the cytokine storm, the receptor-independent effects directly inhibits viral replication. Furthermore, it inhibits the interaction of IL-6 with its receptor complex. Interestingly, in addition to natural hormones, phytestrogens and even synthetic molecules are able to interact with the estrogen receptor and exhibit some anti-COVID-19 activity. From this point of view, estrogen receptor modulators have the potential to be included in the anti-COVID-19 therapeutic arsenal.
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COVID-19/patología , Moduladores de los Receptores de Estrógeno/farmacología , SARS-CoV-2/efectos de los fármacos , Neoplasias de la Mama/complicaciones , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , COVID-19/complicaciones , COVID-19/virología , Moduladores de los Receptores de Estrógeno/metabolismo , Moduladores de los Receptores de Estrógeno/uso terapéutico , Femenino , Humanos , Receptores de Estrógenos/química , Receptores de Estrógenos/metabolismo , SARS-CoV-2/aislamiento & purificación , SARS-CoV-2/fisiología , Proteínas de la Matriz Viral/antagonistas & inhibidores , Proteínas de la Matriz Viral/metabolismo , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
BACKGROUND/AIM: Pancreatic ductal adenocarcinoma (PDAC) still represents one of the most aggressive cancers. Understanding of the epithelial-mesenchymal crosstalk as a crucial part of the tumor microenvironment should pave the way for therapies to improve patient survival rates. Well-established cell lines present a useful and reproducible model to study PDAC biology. However, the tumor-stromal interactions between cancer cells and cancer-associated fibroblasts (CAFs) are still poorly understood. MATERIALS AND METHODS: We studied interactions between four PDAC cell lines (Panc-1, CAPAN-2, MIAPaCa-2, and PaTu-8902) and conditioned media derived from primary cultures of normal fibroblasts/PDAC-derived CAFs (PANFs). RESULTS: When the tested PDAC cell lines were stimulated by PANF-derived conditioned media, the most aggressive behavior was acquired by the Panc-1 cell line (increased number and size of colonies, remaining expression of vimentin and keratin 8 as well as increase of epithelial-to-mesenchymal polarization markers), whereas PaTu-8902 cells were rather inhibited. Of note, administration of the conditioned media to MIAPaCa-2 cells resulted in an inverse effect on the size and number of colonies, whereas CAPAN-2 cells were rather stimulated. To explain the heterogeneous pattern of the observed PDAC crosstalk at the in vitro level, we further compared the phenotype of primary cultures of cells derived from ascitic fluid with that of the tested PDAC cell lines, analyzed tumor samples of PDAC patients, and performed gene expression profiling of PANFs. Immuno-cyto/histo-chemical analysis found specific phenotype differences within the group of examined patients and tested PDAC cell lines, whereas the genomic approach in PANFs found the key molecules (IL6, IL8, MFGE8 and periostin) that may contribute to the cancer aggressive behavior. CONCLUSION: The desmoplastic patient-specific regulation of cancer cells by CAFs (also demonstrated by the heterogeneous response of PDAC cell lines to fibroblasts) precludes simple targeting and development of an effective treatment strategy and rather requires establishment of an individualized tumor-specific treatment protocol.
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Fibroblastos Asociados al Cáncer/patología , Carcinoma Ductal Pancreático/patología , Fibroblastos/patología , Neoplasias Pancreáticas/patología , Fibroblastos Asociados al Cáncer/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Línea Celular Tumoral , Transición Epitelial-Mesenquimal , Fibroblastos/metabolismo , Humanos , Neoplasias Pancreáticas/metabolismo , Microambiente TumoralRESUMEN
Heterogeneous spheroids have recently acquired a prominent position in melanoma research because they incorporate microenvironmental cues relevant for melanoma. In this study, we focused on the analysis of microenvironmental factors introduced in melanoma heterogeneous spheroids by different dermal fibroblasts. We aimed to map the fibroblast diversity resulting from previously acquired damage caused by exposure to extrinsic and intrinsic stimuli. To construct heterogeneous melanoma spheroids, we used normal dermal fibroblasts from the sun-protected skin of a juvenile donor. We compared them to the fibroblasts from the sun-exposed photodamaged skin of an adult donor. Further, we analysed the spheroids by single-cell RNA sequencing. To validate transcriptional data, we also compared the immunohistochemical analysis of heterogeneous spheroids to melanoma biopsies. We have distinguished three functional clusters in primary human fibroblasts from melanoma spheroids. These clusters differed in the expression of (a) extracellular matrix-related genes, (b) pro-inflammatory factors, and (c) TGFß signalling superfamily. We observed a broader deregulation of gene transcription in previously photodamaged cells. We have confirmed that pro-inflammatory cytokine IL-6 significantly enhances melanoma invasion to the extracellular matrix in our model. This supports the opinion that the aspects of ageing are essential for reliable melanoma 3D modelling in vitro.
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Interleukin-6 (IL-6) is a cytokine with multifaceted effects playing a remarkable role in the initiation of the immune response. The increased level of this cytokine in the elderly seems to be associated with the chronic inflammatory setting of the microenvironment in aged individuals. IL-6 also represents one of the main signals in communication between cancer cells and their non-malignant neighbours within the tumour niche. IL-6 also participates in the development of a premetastatic niche and in the adjustment of the metabolism in terminal-stage patients suffering from a malignant disease. IL-6 is a fundamental factor of the cytokine storm in patients with severe COVID-19, where it is responsible for the fatal outcome of the disease. A better understanding of the role of IL-6 under physiological as well as pathological conditions and the preparation of new strategies for the therapeutic control of the IL-6 axis may help to manage the problems associated with the elderly, cancer, and serious viral infections.
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Envejecimiento/metabolismo , Infecciones por Coronavirus/metabolismo , Interleucina-6/metabolismo , Neoplasias/metabolismo , Neumonía Viral/metabolismo , Envejecimiento/patología , Animales , COVID-19 , Infecciones por Coronavirus/patología , Humanos , Interleucina-6/genética , Neoplasias/patología , Pandemias , Neumonía Viral/patología , Transducción de SeñalRESUMEN
The incidence of cutaneous malignant melanoma (CMM) is significantly increasing worldwide.[...].