Exploring the Key Role of Nanotechnology on Intratumoral Microbiome Modulation for Cancer Immunotherapy

Mesenchymal stem cells (MSCs) are multipotent stem cells that reside in various parts of the body like adipose tissue, bone marrow and umbilical cord with an ability to differentiate into chondrocytes, adipocytes and osteocytes. Rigorous research has helped us understand that MSCs home to wound sites and this homing mechanism has been used in the treatment of many inflammatory diseases. It is now emerging that MSCs are an important component of the tumor microenvironment (TME) and contribute to tumor plasticity Plasticity in tumor cells refers to the ability to undergo molecular and phenotypic changes due to environmental cues or genetic alterations. . MSCs are one of the key players within the TME and can either inhibit or promote tumor cell growth by distinct types of cellular interaction. These multifunctional cells can reorganize the tumor stroma : It refers to the supportive tissue consisting of connective tissue or blood vessels around an organ or tumor. which, in turn, can trigger changes in metastatic behavior and promote dedifferentiation to develop cancer stem-like cells. On the contrary, MSCs have been proposed as ideal candidates as drug delivery agents in treatment of various cancers. The double-edged role of MSCs in tumor has made it difficult to pinpoint whether MSCs promote or inhibit tumor growth and progression. During cancer progression, tumor cells undergo molecular and phenotypic changes as a result of microenvironmental cues, genetic and epigenetic alterations and treatment-imposed selective pressures which contribute to tumor heterogeneity and therapy resistance. So, understanding the mechanisms underlying the tumor plasticity may deliver new strategies for targeting cancer metastasis and resistance to therapy. Accordingly, this review focuses on diverse mechanisms of interaction between MSCs and cancer cells with emphasis on different types of intercellular communication affecting tumor progression and metastasis.

Cancer remains one of the most significant causes of mortality across the world. Despite remarkable advancements made in early detection, therapeutic strategies, and the advent of immunotherapy in recent years, numerous challenges continue to hinder optimal outcomes. The development and progression of cancer are driven not only by genetic and epigenetic alterations within tumor cells but also by dynamic interactions occurring with the surrounding tumor microenvironment (TME). It is a highly complex milieu composed of tumor cells, non-tumor stromal cells, extracellular matrix components, immune cells, blood vessels, and diverse signaling molecules. Emerging evidence underscores the pivotal role of fungi in influencing cancer biology, including initiation, progression, immune evasion, and the modulation of TME. Fungi, which are omnipresent microorganisms, have traditionally been considered opportunistic pathogens. However, recent research highlights their broader impact on host immunity and their potential contributions to cancer pathogenesis. For instance, in patients with cancer, fungal infections not only exacerbate clinical complications but also create conditions conducive to tumor growth, metastasis, and immune escape by altering the immune microenvironment. In addition, fungal-derived metabolites and their interactions with host immune pathways can significantly modulate the efficacy of immunotherapies. These findings have spurred interest in exploring antifungal strategies as adjunctive approaches in cancer management, positioning antifungal therapy as a burgeoning area of oncological research. This review provides an in-depth exploration of the complex interplay between fungi and cancer. It examines the multifaceted role of fungal infections in tumor biology, the mechanisms through which fungi reshape the TME through immune modulation and their influence on immune-evasion strategies and therapeutic resistance. Furthermore, the potential for integrating antifungal therapies into comprehensive cancer treatment regimens has been highlighted, offering insights into novel avenues for improving patient outcomes.

Targeting the tumor microenvironment is increasingly recognized as an effective treatment of advanced lung adenocarcinoma (LUAD). However, few studies have addressed the efficacy of immunotherapy for LUAD. Here, a novel method for predicting immunotherapy efficacy has been proposed, which combines single-cell and bulk sequencing to characterize the immune microenvironment and metabolic profile of LUAD. TCGA bulk dataset was used to cluster two immune subtypes: C1 with "cold" tumor characteristics and C2 with "hot" tumor characteristics, with different prognosis. The Scissor algorithm, which is based on these two immune subtypes, identified GSE131907 single cell dataset into two groups of epithelial cells, labeled as Scissor_C1 and Scissor_C2. The enrichment revealed that Scissor_C1 was characterized by hypoxia, and a hypoxic microenvironment is a potential inducing factor for tumor invasion, metastasis, and immune therapy non-response. Furthermore, single cell analysis was performed to investigate the molecular mechanism of hypoxic microenvironment-induced invasion, metastasis, and immune therapy non-response in LUAD. Notably, Scissor_C1 cells significantly interacted with T cells and cancer-associated fibroblasts (CAF), and exhibited epithelial-mesenchymal transition and immunosuppressive features. CellChat analysis revealed that a hypoxic microenvironment in Scissor_C1elevated TGFβ signaling and induced ANGPTL4 and SEMA3C secretion. Interaction with endothelial cells with ANGPTL4, which increases vascular permeability and achieves distant metastasis across the vascular endothelium. Additionally, interaction of tumor-associated macrophages (TAM) and Scissor_C1 via the EREG/EFGR pathway induces tyrosine kinase inhibitor drug-resistance in patients with LAUD. Thereafter, a subgroup of CAF cells that exhibited same features as those of Scissor_C1 that exert immunosuppressive functions in the tumor microenvironment were identified. Moreover, the key genes (EPHB2 and COL1A1) in the Scissor_C1 gene network were explored and their expressions were verified using immunohistochemistry. Finally, the metabolism dysfunction in cells crosstalk was determined, which is characterized by glutamine secretion by TAM and uptake by Scissor_C1 via SLC38A2 transporter, which may induce glutamine addiction in LUAD cells. Overall, single-cell sequencing clarifies how the tumor microenvironment affects immunotherapy efficacy via molecular mechanisms and biological processes, whereas bulk sequencing explains immunotherapy efficacy based on clinical information.

Cancer-associated fibroblasts (CAFs) and tumor-associated macrophages (TAMs); where do they stand in tumorigenesis and how they can change the face of cancer therapy?

Innate immune cells in the tumor microenvironment.

Macrophages play a dual role in the tumor microenvironment(TME), capable of secreting pro-inflammatory factors to combat tumors while also promoting tumor growth through angiogenesis and immune suppression. This study aims to explore the characteristics of macrophages in lung adenocarcinoma (LUAD) and establish a prognostic model based on macrophage-related genes. We performed scRNA-seq analysis to investigate macrophage heterogeneity and their potential pseudotime evolutionary processes. Specifically, we used scRNA-seq data processing, intercellular communication analysis, pseudotime trajectory analysis, and transcription factor regulatory analysis to reveal the complexity of macrophage subpopulations. Data from The Cancer Genome Atlas (TCGA) was used to assess the impact of various macrophage subtypes on LUAD prognosis. Univariate Cox regression was applied to select prognostic-related genes from macrophage markers. We constructed a prognostic model using Lasso regression and multivariate Cox regression, categorizing LUAD patients into high and low-risk groups based on the median risk score. The model's performance was validated across multiple external datasets. We also examined differences between high and low-risk groups in terms of pathway enrichment, mutation information, tumor microenvironment(TME), and immunotherapy efficacy. Finally, RT-PCR confirmed the expression of model genes in LUAD, and cellular experiments explored the carcinogenic mechanism of COL5A1. We found that signals such as SPP1 and MIF were more active in tumor tissues, indicating potential oncogenic roles of macrophages. Using macrophage marker genes, we developed a robust prognostic model for LUAD that effectively predicts prognosis and immunotherapy efficacy. A nomogram was constructed to predict LUAD prognosis based on the model's risk score and other clinical features. Differences between high and low-risk groups in terms of TME, enrichment analysis, mutational landscape, and immunotherapy efficacy were systematically analyzed. RT-PCR and cellular experiments supported the oncogenic role of COL5A1. Our study identified potential oncogenic mechanisms of macrophages and their impact on LUAD prognosis. We developed a prognostic model based on macrophage marker genes, demonstrating strong performance in predicting prognosis and immunotherapy efficacy. Finally, cellular experiments suggested COL5A1 as a potential therapeutic target for LUAD.

Tumor development and metastasis are facilitated by the complex interactions between cancer cells and their microenvironment, which comprises stromal cells and extracellular matrix (ECM) components, among other factors. Stromal cells can adopt new phenotypes to promote tumor cell invasion. A deep understanding of the signaling pathways involved in cell‐to‐cell and cell‐to‐ECM interactions is needed to design effective intervention strategies that might interrupt these interactions. In this review, we describe the tumor microenvironment (TME) components and associated therapeutics. We discuss the clinical advances in the prevalent and newly discovered signaling pathways in the TME, the immune checkpoints and immunosuppressive chemokines, and currently used inhibitors targeting these pathways. These include both intrinsic and non‐autonomous tumor cell signaling pathways in the TME: protein kinase C (PKC) signaling, Notch, and transforming growth factor (TGF‐β) signaling, Endoplasmic Reticulum (ER) stress response, lactate signaling, Metabolic reprogramming, cyclic GMP–AMP synthase (cGAS)–stimulator of interferon genes (STING) and Siglec signaling pathways. We also discuss the recent advances in Programmed Cell Death Protein 1 (PD‐1), Cytotoxic T‐Lymphocyte Associated Protein 4 (CTLA4), T‐cell immunoglobulin mucin‐3 (TIM‐3) and Lymphocyte Activating Gene 3 (LAG3) immune checkpoint inhibitors along with the C‐C chemokine receptor 4 (CCR4)‐ C‐C class chemokines 22 (CCL22)/ and 17 (CCL17), C‐C chemokine receptor type 2 (CCR2)‐ chemokine (C‐C motif) ligand 2 (CCL2), C‐C chemokine receptor type 5 (CCR5)‐ chemokine (C‐C motif) ligand 3 (CCL3) chemokine signaling axis in the TME. In addition, this review provides a holistic understanding of the TME as we discuss the three‐dimensional and microfluidic models of the TME, which are believed to recapitulate the original characteristics of the patient tumor and hence may be used as a platform to study new mechanisms and screen for various anti‐cancer therapies. We further discuss the systemic influences of gut microbiota in TME reprogramming and treatment response. Overall, this review provides a comprehensive analysis of the diverse and most critical signaling pathways in the TME, highlighting the associated newest and critical preclinical and clinical studies along with their underlying biology. We highlight the importance of the most recent technologies of microfluidics and lab‐on‐chip models for TME research and also present an overview of extrinsic factors, such as the inhabitant human microbiome, which have the potential to modulate TME biology and drug responses.

Glioblastoma (GBM) is the most aggressive primary brain tumor, characterized by rapid progression, profound heterogeneity, and resistance to conventional therapies. This review provides an integrated overview of GBM's pathophysiology, highlighting key mechanisms such as neuroinflammation, genetic alterations (e.g., EGFR, PDGFRA), the tumor microenvironment, microbiome interactions, and molecular dysregulations involving gangliosides and sphingolipids. Current diagnostic strategies, including imaging, histopathology, immunohistochemistry, and emerging liquid biopsy techniques, are explored for their role in improving early detection and monitoring. Treatment remains challenging, with standard therapies-surgery, radiotherapy, and temozolomide-offering limited survival benefits. Innovative therapies are increasingly being explored and implemented, including immune checkpoint inhibitors, CAR-T cell therapy, dendritic and peptide vaccines, and oncolytic virotherapy. Advances in nanotechnology and personalized medicine, such as individualized multimodal immunotherapy and NanoTherm therapy, are also discussed as strategies to overcome the blood-brain barrier and tumor heterogeneity. Additionally, stem cell-based approaches show promise in targeted drug delivery and immune modulation. Non-conventional strategies such as ketogenic diets and palliative care are also evaluated for their adjunctive potential. While novel therapies hold promise, GBM's complexity demands continued interdisciplinary research to improve prognosis, treatment response, and patient quality of life. This review underscores the urgent need for personalized, multimodal strategies in combating this devastating malignancy.

Tumor microenvironment reprogramming by nanomedicine to enhance the effect of tumor immunotherapy

Our view of cancer biology radically shifted from a "cancer-cell-centric" vision to a view of cancer as an organ disease. The concept that genetic and/or epigenetic alterations, at the basis of cancerogenesis, are the main if not the exclusive drivers of cancer development and the principal targets of therapy, has now evolved to include the tumor microenvironment in which tumor cells can grow, proliferate, survive, and metastasize only within a favorable environment. The interplay between cancer cells and the non-cellular and cellular components of the tumor microenvironment plays a fundamental role in tumor development and evolution both at the primary site and at the level of metastasis. The shape of the tumor cells and tumor mass is the resultant of several contrasting forces either pro-tumoral or anti-tumoral which have at the level of the tumor microenvironment their battle field. This crucial role of tumor microenvironment composition in cancer progression also dictates whether immunotherapy with immune checkpoint inhibitor antibodies is going to be efficacious. Hence, tumor microenvironment deconvolution has become of great relevance in order to identify biomarkers predictive of efficacy of immunotherapy. In this short paper we will briefly review the relationship between inflammation and cancer, and will summarize in 10 short points the key concepts learned so far and the open challenges to be solved.

The tumor microenvironment (TME) plays a crucial role in tumor initiation, progression, and metastasis, and immunotherapy targeting the TME has received increasing attention. However, single-agent immunotherapy has certain limitations and often requires combination with other adjuvant strategies to enhance therapeutic efficacy. Among these, ultrasound has emerged as a promising adjunct to cancer immunotherapy. By modulating the TME, ultrasound combined with immunotherapy shows great potential in enhancing antitumor responses. This review summarizes the application of various ultrasound modalities in enhancing antitumor immunity, improving the efficacy of immunotherapy, and regulating the TME. Ultrasound can amplify the therapeutic effects of immunotherapy through multiple mechanisms, including thermal effects, mechanical effects, microbubble cavitation, and sonodynamic therapy. Thermal effects induced by high-intensity focused ultrasound (HIFU) can destroy tumor tissues, releasing tumor antigens and heat shock proteins, thereby activating systemic immune responses. Mechanical approaches such as histotripsy can liquefy tumors without thermal damage, preserving antigenic structures and enhancing immune responses within the TME. Ultrasound-mediated microbubble cavitation increases vascular permeability, facilitating the delivery of immune cells and immune checkpoint inhibitors into tumor tissues and enhancing signal transduction to convert “cold” tumors into immune-active “hot” tumors. Sonodynamic therapy generates reactive oxygen species under ultrasound stimulation, inducing immunogenic cell death and reshaping the TME. Furthermore, this review outlines the research progress of ultrasound-immunotherapy combinations in various cancers, including lung cancer, breast cancer, and melanoma, demonstrating superior efficacy compared to immunotherapy alone. Ultrasound not only enhances antitumor immune effects but also enables real-time monitoring of tumor progression and immune modulation within the TME. Finally, the review discusses current challenges and future prospects. By systematically summarizing the types of ultrasound-assisted immunotherapy, their mechanisms within the TME, and recent advances in clinical applications, this article aims to provide a theoretical foundation and technical reference for developing ultrasound-immunotherapy strategies targeting the TME.

The tumor microenvironment (TME) is a critical determinant of cancer progression and therapeutic resistance. Recent advances in nanotechnology offer promising strategies to overcome the challenges posed by the TME, particularly in enhancing drug delivery and improving treatment efficacy. This review examines the composition and dynamics of the TME, emphasizing the complex interactions between cancer cells, stromal cells, immune components, and extracellular matrix factors that contribute to tumor growth and metastasis. Nanotherapeutics, with their ability to target the TME specifically, provide a unique opportunity to bypass the barriers that hinder traditional cancer therapies, such as hypoxia, acidity, and dense extracellular matrices. By exploring both passive and active targeting mechanisms, this review highlights the role of nanomaterials, such as liposomes, polymeric nanoparticles, and metallic nanoparticles, in overcoming these barriers and improving therapeutic outcomes. It also discusses the integration of nanotechnology with immunotherapies and other treatment modalities, enhancing immune responses and modulating the TME. Despite significant progress, challenges remain in the clinical translation of these nano-therapeutics. These include issues related to heterogeneity within the TME, resistance mechanisms, and the potential toxicity of nanomaterials. The review also covers the latest clinical trials, offering a comprehensive view of the current landscape and future directions for nano-based cancer therapies. By synthesizing recent research, this review provides valuable insights into the potential of nanomedicine in revolutionizing cancer treatment strategies, addressing key challenges, and improving patient outcomes.

Non-small cell lung carcinoma (NSCLC) is the leading cause of cancer-related deaths worldwide, with adenocarcinoma being the primary histological subtype. Third-generation EGFR tyrosine kinase inhibitor (TKI) osimertinib is the first-line treatment for NSCLC patients with tumors harboring EGFR kinase domain mutations, significantly improving patient survival. However, mechanisms of acquired drug resistance inevitably emerge and must be addressed. While the acquisition of secondary mutations in EGFR, such as C797S mutation, is a known resistance mechanism, non-genetic mechanisms of drug resistance, including vasoconstriction reducing the flow of the blood and the drug to the tumors, have also been observed and are highly significant. We have developed microfluidic devices that support the independent growth of tumor spheroids and patient-derived organoids (PDOs). These platforms feature U-shaped microwells that enable the sustained growth of spheroids for several weeks. This system has proven highly effective for viability assays using small samples including PDOs, suggesting its potential application in personalized medicine assays. More recently, these devices have been employed to evaluate the impact of tumor microenvironment (TME) components, such as endothelial and aortic smooth muscle cells, on drug sensitivity. Indirect co-culture of endothelial HUVEC cells and H6080 aortic smooth muscle cells with various EGFR mutant NSCLC cells promoted EGFR TKI resistance in the epithelial cells, increasing the IC50 by the factor of 10 in H1975 and HCC827. Pharmacodynamic analyses revealed that ERK phosphorylation levels in the presence of supernatants from endothelial and vascular cells, indicating a significant paracrine interaction between TME cells and tumor cells. Notably, this enhanced drug resistance was also observed in KRASG12C H358 mutant cells in response to adagrasib, a KRASG12C inhibitor.Interestingly, this microfluidic device can be optimized to measure relevant chemokines and cytokines secreted by cell lines and PDOs. Characterizing the content and concentrations of various metabolites is particularly important. For example, we have demonstrated that elevated levels of EDN1 promote vasoconstriction leading to drug resistance both in vitro and in vivo. The HCC827, HCC4006 and H1975 EGFR-mutant cell lines show a significant increase in EDN1 mRNA expression and decrease in VEGFA mRNA after 72h of osimertinib treatment. This transcriptional regulation of EDN1 and VEGF-A result in elevated level of EDN1 peptide and depleted VEGF-A in cell lines and EGFR-mutant PDOs including RLUN007. The resulting changes in the peptides promoted vasoconstriction in tumor-feeding vessels and reduced concentration of osimertinib in tumors. These findings underscore the need to develop tools that replicate and mimic the intricate interactions between tumor cells and the TME. Such advancements will enable the use of relevant and limited patient samples to implement personalized medicine approaches and improve patients outcomes. Citation Format: Mohammad Amir Mishan, Ali Rahnama, Ines Pulido Endrino, Laura Gunder, Malek Massad, Khaled Abdelhady, Agustin Lahoz, Ian Papautsky, Takeshi Shimamura. Microfluidic devices to decipher the role of the microenvironment in drug resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr LB220.

Making the Case for DCC and UNC5C as Tumor-Suppressor Genes in the Colon

As recent decades have seen extensive resources dedicated to exploring tumor metastasis, discoveries have exposed a need for understanding the intricacies of the tumor microenvironment (TME) in an ex vivo model. Much of the heterogeneity of the disease stems from the complexity of interactions between the cells that comprise the TME, placing its investigation at the heart of developing effective treatment. A number of findings implicate chemokines and their receptors, expressed early in cancer progression, in promoting tumor metastasis. These findings identify several cell types significantly involved in secreting and responding to chemokines, such as tumor cells, fibroblasts, and endothelial cells, among others. Chemokine expression depends not only on the cell types of the TME but also on the conditions in the surrounding matrix, especially interstitial flow. A strong link has been identified between interstitial flow and tumor metastasis. This flow, a result of rapid lymphangiogenesis, correlates with the expression of a number of chemokines responsible for tumor invasion and metastasis. Current limitations for studying the human TME have stalled progress since mouse models cannot fully engage all the relevant human cell types. Thus, the emergence of complex intercellular relationships within the TME generates a need for heterotypic three-dimensional in vitro models to cover the gaps between animal models and human studies. The need for 3D in vitro models is amplified by the exposed importance of interstitial flow. To mimic and examine the breast cancer tumor, we have designed a bioreactor, machined through femtosecond laser-etching technology that allows for co-culture of the numerous cell types that comprise the TME. The design incorporates a semicircular chamber connected to a channel 200 μm in width by a thin porous filter of laser-etched glass. The device is coated with poly-L-lysine and Collagen I to improve cell adhesion. To recreate the TME, the semicircular chamber is loaded with cancer-associated fibroblasts and tumor cells (MCF-7 and MDA-MB-231 cell lines) in a ratio of 3:1 in 3D reconstituted basement membrane with 1.5 mg/mL Collagen I (rat-tail) and 10% Matrigel. This 3D cell culture is grown for 5 days to allow spheroid formation. Human microvascular endothelial cells are cultured to form a confluent monolayer in the channel that is then perfused with medium, of which a higher quantity is aspirated from the outlet to generate interstitial flow through the 3D cell culture. Cancer-associated leukocytes can be infused through the channel to better mimic the TME. This unique and innovative design will allow in vitro visualization of cell migration and expression, which can shed light on the mechanisms of metastasis. Experiments can then be performed to elucidate the specific relationships linking interstitial flow and metastasis. Citation Format: Kathryn Hockemeyer, Tammy Sobolik, Alexander Terekhov, Melody Swartz, William Hofmeister, John Wikswo, Chris Janetopoulos, Ann Richmond. A three-dimensional in vitro bioreactor for analysis of the tumor microenvironment: investigation of interstitial flow and the mechanism of metastasis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2614. doi:10.1158/1538-7445.AM2013-2614