RESUMO
The field of cancer immunotherapy has shown significant growth, and researchers are now focusing on effective strategies to enhance and prolong local immunomodulation. Injectable hydrogels (IHs) have emerged as versatile platforms for encapsulating and controlling the release of small molecules and cells, drawing significant attention for their potential to enhance antitumor immune responses while inhibiting metastasis and recurrence. IHs delivering natural killer (NK) cells, T cells, and antigen-presenting cells (APCs) offer a viable method for treating cancer. Indeed, it can bypass the extracellular matrix and gradually release small molecules or cells into the tumor microenvironment, thereby boosting immune responses against cancer cells. This review provides an overview of the recent advancements in cancer immunotherapy using IHs for delivering NK cells, T cells, APCs, chemoimmunotherapy, radio-immunotherapy, and photothermal-immunotherapy. First, we introduce IHs as a delivery matrix, then summarize their applications for the local delivery of small molecules and immune cells to elicit robust anticancer immune responses. Additionally, we discuss recent progress in IHs systems used for local combination therapy, including chemoimmunotherapy, radio-immunotherapy, photothermal-immunotherapy, photodynamic-immunotherapy, and gene-immunotherapy. By comprehensively examining the utilization of IHs in cancer immunotherapy, this review aims to highlight the potential of IHs as effective carriers for immunotherapy delivery, facilitating the development of innovative strategies for cancer treatment. In addition, we demonstrate that using hydrogel-based platforms for the targeted delivery of immune cells, such as NK cells, T cells, and dendritic cells (DCs), has remarkable potential in cancer therapy. These innovative approaches have yielded substantial reductions in tumor growth, showcasing the ability of hydrogels to enhance the efficacy of immune-based treatments. STATEMENT OF SIGNIFICANCE: As cancer immunotherapy continues to expand, the mode of therapeutic agent delivery becomes increasingly critical. This review spotlights the forward-looking progress of IHs, emphasizing their potential to revolutionize localized immunotherapy delivery. By efficiently encapsulating and controlling the release of essential immune components such as T cells, NK cells, APCs, and various therapeutic agents, IHs offer a pioneering pathway to amplify immune reactions, moderate metastasis, and reduce recurrence. Their adaptability further shines when considering their role in emerging combination therapies, including chemoimmunotherapy, radio-immunotherapy, and photothermal-immunotherapy. Understanding IHs' significance in cancer therapy is essential, suggesting a shift in cancer treatment dynamics and heralding a novel period of focused, enduring, and powerful therapeutic strategies.
Assuntos
Hidrogéis , Neoplasias , Humanos , Hidrogéis/uso terapêutico , Imunoterapia/métodos , Neoplasias/patologia , Linfócitos T , Terapia Combinada , Microambiente TumoralRESUMO
Ras plays an essential role in the development of acinar-to-ductal metaplasia (ADM) and pancreatic ductal adenocarcinoma (PDAC). However, mutant Kras is an inefficient driver for PDAC development. The mechanisms of the switching from low Ras activity to high Ras activity that are required for development and progression of pancreatic intraepithelial neoplasias (PanINs) are unclear. In this study, we found that hematopoietic progenitor kinase 1 (HPK1) was upregulated during pancreatic injury and ADM. HPK1 interacted with the SH3 domain and phosphorylated Ras GTPase-activating protein (RasGAP) and upregulated RasGAP activity. Using transgenic mouse models of HPK1 or M46, a kinase-dead mutant of HPK1, we showed that HPK1 inhibited Ras activity and its downstream signaling and regulated acinar cell plasticity. M46 promoted the development of ADM and PanINs. Expression of M46 in KrasG12D Bac mice promoted the infiltration of myeloid-derived suppressor cells and macrophages, inhibited the infiltration of T cells, and accelerated the progression of PanINs to invasive and metastatic PDAC, while HPK1 attenuated mutant Kras-driven PanIN progression. Our results showed that HPK1 plays an important role in ADM and the progression of PanINs by regulating Ras signaling. Loss of HPK1 kinase activity promotes an immunosuppressive tumor microenvironment and accelerates the progression of PanINs to PDAC.
Assuntos
Carcinoma in Situ , Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Camundongos , Animais , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Neoplasias Pancreáticas/patologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Carcinoma in Situ/genética , Carcinoma in Situ/metabolismo , Carcinoma in Situ/patologia , Camundongos Transgênicos , Microambiente Tumoral , Neoplasias PancreáticasRESUMO
Cancer cells release extracellular vesicles known as extracellular vesicles (EVs), containing tumor-derived DNA, RNA and proteins within their cargo, into the circulation. Circulating tumor-derived extracellular vesicles (TEV) can be used in the context of serial "liquid biopsies" for early detection of cancer, for monitoring disease burden in patients, and for assessing recurrence in the post-resection setting. Nonetheless, isolating sufficient TEV by ultracentrifugation-based approaches, in order to enable molecular assessment of EVs cargo, can be an arduous, time-consuming process and is inconsistent in the context of yield and purity among institutions. Herein, we describe a microfluidic platform, which we have named MITEV (Microfluidic Isolation of Tumor-derived Extracellular Vesicles) for the rapid isolation of TEV from the plasma of pancreatic cancer patients. The device, which has ~100,000 pillars placed in a zigzag pattern and is coated with antibodies against generic EV surface proteins (anti-CD63, -CD9, and -CD81 antibodies) or the TEV specific anti-Epithelial Cell Adhesion Molecule (EpCAM) antibody, is capable of high-throughput EVs isolation and yields sufficient DNA (total of ~2-14 ng from 2-ml plasma) for downstream genomic analysis. Using two independent quantitative platforms, droplet digital polymerase chain reaction (ddPCR) and molecular barcoding using nanoString nCounter® technology, we can reliably identify KRAS mutations within isolated TEV of treatment-naïve metastatic pancreatic cancer patients. Our study suggests that the MITEV device can be used for point-of-care applications, such as in the context of monitoring residual or recurrent tumor presence in pancreatic cancer patients undergoing therapy.
Assuntos
Separação Celular/instrumentação , Análise Mutacional de DNA/instrumentação , Vesículas Extracelulares/genética , Vesículas Extracelulares/patologia , Dispositivos Lab-On-A-Chip , Neoplasias Pancreáticas/patologia , Linhagem Celular Tumoral , Genômica , Humanos , Proteínas Proto-Oncogênicas p21(ras)/genéticaRESUMO
PURPOSE: The objective of this study was to bioengineer 3D patches from cardiac myocytes that have been reprogrammed from human adipogenic mesenchymal stem cells (hADMSCs). METHODS: Human adipogenic mesenchymal stem cells (hADMSCs) were reprogrammed to form cardiac myocytes using transcription factors ETS2 and MESP1. Reprogrammed cardiac myocytes were cultured in a fibrin gel to bioengineer 3D patch patches. The effect of initial plating density (1-25 million cells per patch), time (28-day culture period) and treatment with 1 µM isoproterenol and 1 µM epinephrine were evaluated. RESULTS: 3D patches were fabricated using cardiac myocytes that have been reprogrammed from hADMSCs. Based on optimization studies, it was determined that 10 million cells were needed to bioengineer a single patch, that measured 2 × 2 cm2. Furthermore, 3D patches fabricated 10 million cells were stable in culture for up to 28 days. Treatment of 3D patches with 1 µM isoproterenol and 1 µM epinephrine resulted in an increase in the electrical properties, as measured by electrical impulse amplitude and frequency. An increase in the expression of mTOR, KCNV1, GJA5, KCNJ16, CTNNT2, KCNV2, MYO3, FOXO1 and KCND2 was noted in response to treatment of 3D patches with isoproterenol and epinephrine. CONCLUSION: Based on the results of this study, there is evidence to support the successful fabrication of a highly functional 3D patches with measurable electrical activity using cardiac myocytes reprogrammed from hADMSCs. 3D patches fabricated using optimal conductions described in this study can be used to improve the functional properties of failing hearts. Predominantly, in case of the infarcted hearts with partial loss of electrical activity, the electrical properties of the 3D patches may restore the electrical activity of the heart.
Assuntos
Adipogenia , Técnicas de Reprogramação Celular , Reprogramação Celular , Insuficiência Cardíaca/cirurgia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais , Miócitos Cardíacos/transplante , Engenharia Tecidual , Agonistas Adrenérgicos/farmacologia , Células Cultivadas , Condutividade Elétrica , Epinefrina/farmacologia , Fibrina/metabolismo , Regulação da Expressão Gênica , Insuficiência Cardíaca/genética , Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/fisiopatologia , Humanos , Isoproterenol/farmacologia , Potenciais da Membrana , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Fatores de TempoRESUMO
The oncogenic receptor tyrosine kinase AXL is overexpressed in cancer and plays an important role in carcinomas of multiple organs. However, the mechanisms of AXL overexpression in cancer remain unclear. In this study, using HEK293T, Panc-1, and Panc-28 cells and samples of human pancreatic intraepithelial neoplasia (PanIN), along with several biochemical approaches and immunofluorescence microscopy analyses, we sought to investigate the mechanisms that regulate AXL over-expression in pancreatic ductal adenocarcinoma (PDAC). We found that AXL interacts with hematopoietic progenitor kinase 1 (HPK1) and demonstrate that HPK1 down-regulates AXL and decreases its half-life. The HPK1-mediated AXL degradation was inhibited by the endocytic pathway inhibitors leupeptin, bafilomycin A1, and monensin. HPK1 accelerated the movement of AXL from the plasma membrane to endosomes in pancreatic cancer cells treated with the AXL ligand growth arrest-specific 6 (GAS6). Moreover, HPK1 increased the binding of AXL to the Cbl proto-oncogene (c-Cbl); promoted AXL ubiquitination; decreased AXL-mediated signaling, including phospho-AKT and phospho-ERK signaling; and decreased the invasion capability of PDAC cells. Importantly, we show that AXL expression inversely correlates with HPK1 expression in human PanINs and that patients whose tumors have low HPK1 and high AXL expression levels have shorter survival than those with low AXL or high HPK1 expression (p < 0.001). Our results suggest that HPK1 is a tumor suppressor that targets AXL for degradation via the endocytic pathway. HPK1 loss of function may contribute to AXL overexpression and thereby enhance AXL-dependent downstream signaling and tumor invasion in PDAC.
Assuntos
Regulação para Baixo , Oncogenes , Neoplasias Pancreáticas/enzimologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Carcinoma in Situ/enzimologia , Carcinoma in Situ/patologia , Linhagem Celular Tumoral , Citoplasma/metabolismo , Endocitose , Endossomos/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Estimativa de Kaplan-Meier , Sistema de Sinalização das MAP Quinases , Invasividade Neoplásica , Neoplasias Pancreáticas/patologia , Ligação Proteica , Transporte Proteico , Proteólise , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-cbl/metabolismo , Ubiquitinação , Receptor Tirosina Quinase AxlRESUMO
Clinical trials using human adipogenic mesenchymal stem cells (hAdMSCs) for the treatment of cardiac diseases have shown improvement in cardiac function and were proven safe. However, hAdMSCs do not convert efficiently into cardiomyocytes (CMs) or vasculature. Thus, reprogramming hAdMSCs into myocyte progenitors may fare better in future investigations. To reprogramme hAdMSCs into electrically conductive cardiac progenitor cells, we pioneered a three-step reprogramming strategy that uses proven MESP1/ETS2 transcription factors, ß-adrenergic and hypoxic signalling induced in three-dimensional (3D) cardiospheres. In Stage 1, ETS2 and MESP1 activated NNKX2.5, TBX5, MEF2C, dHAND, and GATA4 during the conversion of hAdMSCs into cardiac progenitor cells. Next, in Stage 2, ß2AR activation repositioned cardiac progenitors into de novo immature conductive cardiac cells, along with the appearance of RYR2, CAV2.1, CAV3.1, NAV1.5, SERCA2, and CX45 gene transcripts and displayed action potentials. In Stage 3, electrical conduction that was fostered by 3D cardiospheres formed in a Synthecon®, Inc. rotating bioreactor induced the appearance of hypoxic genes: HIF-1α/ß, PCG 1α/ß, and NOS2, which coincided with the robust activation of adult contractile genes including MLC2v, TNNT2, and TNNI3, ion channel genes, and the appearance of hyperpolarization-activated and cyclic nucleotide-gated channels (HCN1-4). Conduction velocities doubled to ~200 mm/s after hypoxia and doubled yet again after dissociation of the 3D cell clusters to ~400 mm/s. By comparison, normal conduction velocities within working ventricular myocytes in the whole heart range from 0.5 to 1 m/s. Epinephrine stimulation of stage 3 cardiac cells in patches resulted in an increase in amplitude of the electrical wave, indicative of conductive cardiac cells. Our efficient protocol that converted hAdMSCs into highly conductive cardiac progenitors demonstrated the potential utilization of stage 3 cells for tissue engineering applications for cardiac repair.
Assuntos
Técnicas de Cultura de Células/métodos , Células-Tronco Mesenquimais/citologia , Receptores Adrenérgicos beta/metabolismo , Adipogenia , Adrenérgicos , Reatores Biológicos , Diferenciação Celular/fisiologia , Proliferação de Células , Condutividade Elétrica , Epinefrina/farmacologia , Humanos , Hipóxia , Cinética , Miócitos Cardíacos/citologia , Transdução de Sinais , Engenharia Tecidual/instrumentação , Engenharia Tecidual/métodos , Alicerces Teciduais , Fatores de Transcrição/metabolismoRESUMO
Fragile histidine triad (FHIT) serves a critical function as a tumor suppressor that inhibits p53 degradation by mouse double minute 2 (MDM2). The functional domains of FHIT involved in tumor inhibition was interpreted. In-silico screening data were employed to construct truncated forms of FHIT to assess their cytotoxic effects on the HT1080 cell line. Full FHIT expression was confirmed by western blotting and expression of two FHIT truncates were confirmed by RT-PCR. Transfection of these truncated forms into HT1080 cells showed that the N-terminal truncated form (amino acids 17-102) better inhibited proliferation than the full-length FHIT. The combined effects of these truncated forms augmented doxorubicin-induced cytotoxicity. Functional analysis demonstrated that these fragments and their combination with doxorubicin can arrest cells in the G2 phase of the cell cycle as specified by flow cytometry. The FHIT functional domains can be used as lead compounds for development of drug designs and gene transfer for cancer therapy.