The implications for urological malignancies of non-coding RNAs in the the tumor microenvironment

Urological malignancies are a major global health issue because of their complexity and the wide range of ways they affect patients. There's a growing need for in-depth research into these cancers, especially at the molecular level. Recent studies have highlighted the importance of non-coding RNAs (ncRNAs) – these don't code for proteins but are crucial in controlling genes – and the tumor microenvironment (TME), which is no longer seen as just a background factor but as an active player in cancer progression. Understanding how ncRNAs and the TME interact is key for finding new ways to diagnose and predict outcomes in urological cancers, and for developing new treatments. This article reviews the basic features of ncRNAs and goes into detail about their various roles in the TME, focusing specifically on how different ncRNAs function and act in urological malignancies.


Introduction
Urological malignancies constitute a substantial threat to worldwide health [1,2].These malignancies cover a wide range of diseases, including cancers of the prostate, bladder, kidney, and testicle, among others.It is imperative to underscore the severity of prostate cancer, holding the title of the second most common cancer in males across the globe.The Global Cancer Statistics 2020 reported an incredible 1.4 million fresh cases and a grim total of 375,000 fatalities in just the year 2020 [3].According to estimates, 151,297 people in Europe were estimated to have bladder cancer in 2012, with age-standardized incidence rates (per 100,000 people) of 17.7 for men and 3.5 for women [4].Age, lifestyle, including smoking and food, and genetic liability are some risk factors affecting these malignancies [5,6].Despite a wide range of available therapies, including targeted therapy, radiation therapy, chemotherapy, immunotherapy, and surgical intervention4, the incidence and mortality toll continue to rise.This pathetic reality highlights the essential urgency for treatments of higher efficacy.
The TME emerges as a critical factor in the advancement of cancer.This dynamic and complex ecosystem is composed of diverse cellular constituents, such as cancer cells, immune cells, fibroblasts, and endothelial cells, all embedded within the elaborate network of the extracellular matrix [7][8][9].The architecture and operational dynamics of the TME are not static but vary according to the type and stage of the tumor, as well as the patient's unique characteristics.The TME exerts a significant influence on every stage of tumor development, encompassing carcinogenesis, tumor growth, angiogenesis, invasion, and metastasis [7,8,10,11].
In this review, we have amalgamated the latest insights from advanced technologies, such as HTS and bioinformatics, to enhance our understanding of ncRNAs in tumor initiation, progression, and resistance to treatment.Additionally, this article delves into the complex interplay between ncRNAs and the TME specifically in urological malignancies.This exploration provides valuable insights and crucial information, which could be instrumental in developing future therapeutic strategies.

Overview of NcRNAs
NcRNAs form a diverse group of RNA molecules that play significant roles in cellular functions, despite not being involved in protein encoding [19].MiRNAs, a subset of small ncRNAs approximately 22 nucleotides in length, act as key regulators of gene expression at the post-transcriptional level.They exert their function by binding to the 3′ UTRs of their target mRNAs, leading to either mRNA degradation or translational inhibition [20].The biogenesis of miRNAs commences with the transcription of a primary miRNA (pri-miRNA), which is then processed into a precursor miRNA (pre-miRNA) by the Drosha-DGCR8 complex in the nucleus.Subsequently, the pre-miRNA is transported to the cytoplasm and further refined into its mature form by the enzyme Dicer.Upon maturation, the miRNA integrates into the RNA-induced silencing complex (RISC), targeting specific mRNA molecules for regulation [21].LncRNAs, exceeding 200 nucleotides in length, are involved in various biological processes, including chromatin remodeling, transcriptional, and post-transcriptional regulation [22,23].LncRNAs are transcribed and interact with DNA, RNA, and proteins to execute their diverse functions.Notably, some lncRNAs can also serve as precursors for small ncRNAs [21].CircRNAs, characterized by their closed-loop structure resulting from back-splicing, form another unique class of ncRNAs.Their functions in cellular processes are varied, such as acting as miRNA sponges, interacting with RBPss, and in certain instances, encoding proteins [24,25].Understanding the general biological roles of ncRNAs is fundamental before delving into their specific functions within the TME of urological malignancies.

NcRNAs: the invisible movers of genetic interactions
NcRNAs, including miRNAs, lncRNAs, and circRNAs, play crucial roles in cellular processes such as gene regulation, cell differentiation, and development.MiRNAs, for instance, regulate gene expression by destabilizing messenger RNAs (mRNAs) and inhibiting their translation, thus impacting a range of cellular functions [26].An illustrative example is the overexpression of MicroRNA-361-3p, leading to decreased SH2B1 levels, which in turn inhibits the proliferation and spread of Non-Small Cell Lung Cancer cells [27].Additionally, miRNAs are instrumental in cellular development, particularly in regulating macrophage activation.They modulate the effects of Th2 cell cytokines interleukin-4 (IL-4) and IL-13 on macrophage biology, biosynthesis, and environmental response, underscoring their importance in immune cell development and reactivity [28].
LncRNAs, another type of ncRNA, regulate gene expression at various stages, including chromatin modification, transcription, and post-transcriptional processing.A prominent example is linc-MD1 in mouse and human myoblasts, which acts as a 'sponge' for miR-133 and miR-135, thereby regulating the transcription factors MAML1 and MEF2C crucial for muscle-specific gene expression and muscle cell differentiation [29].LncRNAs such as HOTAIR function as scaffolds for protein complexes, like the combination of PRC2 and the LSD1/CoR-EST/REST complex, to orchestrate gene silencing [30].Additionally, certain lncRNAs function as molecular decoys; for instance, lncRNA GAS5 inhibits the binding of the glucocorticoid receptor to target gene promoters [31].In the realm of cellular differentiation, the lncRNA Braveheart (Bvht) is critical for cardiovascular lineage commitment in mouse embryonic stem cells, mediating epigenetic changes essential for activating cardiovascular genes through its interaction with the transcription factor SUZ12, a component of PRC2 [32].Furthermore, certain lncRNAs exhibit enhancer-like functions, activating key regulators of development and differentiation in human cell lines [33].CircRNAs, too, play significant roles in gene regulation and cell differentiation.For instance, ciRS-7 functions as a miRNA sponge, regulating gene expression by sequestering miR-7 [34].The circRNA hsa-circ-0074834 promotes osteogenic differentiation of bone marrow stem cells by regulating ZEB1 and VEGF expression via the downregulation of miR-942-5p, highlighting circRNAs' roles in gene regulation and development [35].Interestingly, some circRNAs, like circ-ZNF609, have been found to possess protein-coding potential, expanding our understanding of the functional diversity of circRNAs in cellular processes [36].
In summary, ncRNAs are integral in directing gene expression and various cellular processes, and their dysregulation is often associated with diseases, including cancer, making them potential targets for therapeutic interventions (Table 1).

The impact of genetic variations on ncRNA networks and disease development
Transcriptome-Wide Association Studies (TWAS) have played a crucial role in uncovering genetic variations that are linked with health conditions, including aspects of cancer risk, prognosis, and drug response [85].These studies draw particular attention to the impact of single nucleotide polymorphisms (SNPs) and copy number variations (CNVs), which are found to significantly influence the complex networks of ncRNAs.
SNPs within ncRNAs are known to influence disease development through several pathways.A notable mechanism involves the alteration of splicing models, where SNPs impact the splicing of lncRNA transcripts.This process results in the production of various isoforms with differing binding affinities, subsequently affecting the regulation of genes and cellular pathways.For instance, the lncRNA lnc13, implicated in inflammatory diseases, can be influenced by SNP (rs917997).This variation can lead to an increase in the levels of lnc13-regulated genes, heightening the risk of diseases like coeliac [86].Similarly, an increase in the expression of lncRNA BCLET, associated with a reduced risk of bladder cancer, can be attributed to SNP rs558814, potentially improving survival rates [87].In addition, SNPs can alter ncRNA expression levels, influencing susceptibility to various diseases.An example of this is SNP rs2839698 in lncRNA H19, linked to a decreased risk of non-muscle-invasive bladder cancer [88].Another SNP, rs67311347, enhances the expression of ENTPD3-AS1, which acts protectively in renal cell carcinoma (RCC) by inhibiting cell proliferation via the miR-155-5p/HIF-1α pathway [89].The SNP rs11672691, situated in a region that acts as a promoter for the short isoform of PCAT19 and as an enhancer for the long isoform, along with its linked SNP rs887391, reduces the binding affinity of transcription factors such as NKX3.1 and YY1.This reduced binding leads to a decrease in the expression of the tumor-suppressing PCAT19-short and an increase in the oncogenic PCAT19-long, promoting cell proliferation and tumor growth through interaction with HNRNPAB [90].Moreover, SNPs located within lncRNA genes can disrupt their normal functions,  contributing to disease by altering critical transcriptional regulatory networks.For example, SNP rs6983267, associated with the lncRNA CCAT1-L, can affect the expression of the MYC oncogene, a significant factor in many cancers [91,92].Moreover, SNPs enriched within lncRNAs can also affect their interactions with RNA-binding proteins (RBPs), essential for regulating gene expression [93,94].In bladder cancer, the C allele of SNP rs2910164 changes the expression of miR-146a, leading to an increased expression of its target genes YAP1 and COX2, potentially promoting cancer stem cell characteristics and increasing the risk of recurrence [95].Similarly, the A allele of SNP rs2073859 disrupts the binding of miR-135a, reducing post-transcriptional regulation of LIMK2.This deregulation promotes enhanced cellular processes such as proliferation, migration, and invasion, and decreases apoptosis [96].The lncRNA PCAT1 is influenced by SNP rs7463708 within its enhancer region.This SNP has been shown to enhance the binding of the androgen receptor transcription factor ONECUT2, increasing PCAT1 expression in the presence of androgens, which influence prostate cancer progression [97].
CNVs, structural variations in the genome resulting in abnormal numbers of DNA sections, can overlap with regions encoding ncRNAs, leading to changes in their expression and function [98].In lung adenocarcinoma, for example, CNVs can amplify lncRNA ALAL-1, contributing to cancer cell proliferation and immune evasion [99].Similarly, CNV also contributes to the oncogenic potential of PLANE by upregulating its expression, impacting cancer development and progression through its interaction with NCOR2 pre-mRNA splicing and hnRNPMM [100].Furthermore, ncRNAs can influence DNA methylation targets, either activating or inhibiting them, and this regulation is also modulated by CNVs [101].Specifically, lncRNAs can interact with DNA methyltransferases (DNMTs), recruiting or inhibiting these enzymes at chromatin loci, thereby affecting gene expression [102].

Elucidating ncRNA functions through single-cell 2.3.1. Discovery and characterization of cell types
Single-cell RNA sequencing (scRNA-seq) is revolutionizing our understanding of cellular complexity.This advanced technique has uncovered a multitude of previously unrecognized cell types and states.In the field of neuroscience, for instance, Zeisel et al. utilized scRNA-seq to classify cell types in the mouse somatosensory cortex and hippocampus.Their study identified 47 distinct subclasses, including a variety of neurons and glial cells, each characterized by unique gene expression profiles [103].This pivotal research expanded our catalog of neural cell types and shed light on their functional and developmental interconnections.
In immunology, Villani et al. applied scRNA-seq to dissect the human immune system, revealing an extensive array of immune cell types [104].The team characterized new subtypes of dendritic cells, monocytes, and progenitors, each distinguished by distinct gene expression patterns.These discoveries have significant implications for understanding immune responses and the development of targeted immunotherapies.
In the oncology domain, Patel et al. employed scRNA-seq to explore the cellular makeup of glioblastoma, an aggressive form of brain cancer [105].Their research identified a diverse range of cell types within tumors, including various malignant subpopulations and non-malignant cells, each contributing differently to tumor progression and therapy resistance.Single-cell analysis has been crucial in understanding the role of CD4 + T cells in antitumor immunity.Notably, patients responding to immunotherapy showed a marked enrichment of Th1-like CD4 + T cells, essential in mediating therapeutic outcomes [106].Furthermore, single-cell studies have identified specific subsets of TAMss (TAMs) with distinct roles in cancer development.For example, TAMs expressing C1Q have been implicated in shaping the tumor's immune microenvironment, influencing T cell recruitment and regulation [107].Such research provides a more detailed perspective of tumor heterogeneity, vital for developing effective personalized treatments.
In a related study, Gouin et al. combined scRNA-seq with spatial transcriptomics and proteomics to investigate the cellular details of bladder cancer [108].They identified a novel subpopulation of epithelial cells characterized by CDH12 expression, which exhibits stem-like properties.This subgroup significantly affects the tumor's response to treatments, including surgery, chemotherapy, and immunotherapy.This study not only deepened our understanding of cellular heterogeneity in bladder cancer but also opened new avenues for targeted treatment strategies and personalized medicine.More findings on cellular heterogeneity are shown in Table 2.

Multifaceted Roles of ncRNAs Revealed by scRNA-seq
scRNA-seq has been instrumental in uncovering the diverse and complex roles of ncRNAs in cellular processes.This technique offers a high-resolution analysis of gene expression at the individual cell level, enabling detailed observation of the nuanced regulatory functions of ncRNAs across development, health, and disease.
For example, scRNA-seq facilitated the identification of a lncRNA, lncMMPA, within exosomes from tumor-associated macrophages (TAMs), which plays a crucial role in the malignancy of hepatocellular carcinoma (HCC).The study revealed that lncMMPA promotes tumor cell glycolysis and proliferation by sequestering miR-548 s, leading to scRNA-seq and ST Nine distinct macrophage clusters were identified, each showing unique enrichment patterns at the tumor core and periphery [110] scRNA-seq and TCR-seq An enrichment of CD8A+ tissue-resident T cells has been associated with positive responses to ICB therapy.Conversely, an increased presence of TAMs has been observed in patients resistant to ICB [111] scRNA-seq and scATAC-seq Tumor cell-specific regulatory programs were identified, mediated by key transcription factors (TFs): HOXC5, VENTX, ISL1, and OTP [112] scRNA-seq and scATAC-seq Identification of specific long non-coding RNAs, RP11-661C8.2 and CTB-164N12.1, potentially contributing to the invasion and migration abilities of ccRCC [113] scRNA-Seq The expression of complement serine protease C1S in tumors correlated positively with levels of macrophage infiltration [114] scRNA-Seq Identification of SERPINE2 as a key gene for metastasis and EMT [115] BCa scRNA-seq and TCR sequencing Two cytotoxic phenotypes of CD4 + T cells, CD4GZMB and CD4GZMK, have been identified, capable of eliminating autologous tumor cells through an MHC class II-dependent pathway [116] scRNA-seq A new subpopulation of macrophages was identified in UCB, named Macro-C3.The Macro-C3 subset score is predictive of the response to immunotherapy in these patients [117] scRNA-seq and ATAC-seq Increased expression of EZH2 enhances stemness in cancer stem cells and diminishes cell adhesion in bladder cancer stem cells, thereby contributing to the recurrence of bladder cancer [118] BCa scRNA-seq and TCR-seq CD57 + CD8 T cells in the peripheral blood of mUC patients can serve as a potential biomarker for predicting patient response to atezolizumab [119] scRNA-seq Recruitment and activation of TAM by iCAF through CXCL12-CXCR4 as well as THY1, CSF signaling pathways plays a key role in orchestrating TME, which leads to T-cell depletion and establishes an immunosuppressive environment conducive to tumor progression [120] PCa scRNA-seq Four tumor endothelial cell subtypes have been identified: arterial, venous plexus, immature vascular, and tip cells.The CXCR4/CXCL12 axis, mainly expressed in arterial cells and associated with angiogenesis, presents a potential target for anti-angiogenic therapy in prostate cancer [121] scRNA-seq EEF2 + and FOLH1 + luminal subpopulations are exclusive to LNM in PCa.MYC facilitates PCa progression and TME immunosuppression by modulating PDL1 and CD47 [122] scRNA-seq Primary prostate cancer typically shows a low presence of tumor-infiltrating immune cells, and mCSPC exhibits a robust immune infiltrate [123] (continued on next page) the upregulation of ALDH1A3.This discovery sheds light on the contribution of TAM-derived exosomes to HCC progression, positioning lncMMPA as a potential prognostic marker [126].
In the context of cancer research, scRNA-seq has been vital in exploring the role of ncRNAs in intratumoral heterogeneity and drug resistance.Zhao et al. found that the lncRNA NEAT1 was upregulated in specific metabolically advantaged subpopulations within hepatobiliary tumor organoids.Notably, in the CD44 positive cells of the HCC272 organoid, NEAT1 expression correlated with drug resistance mediated through the Jak-STAT pathway [127].Additionally, scRNA-seq uncovered ZNFX1 antisense RNA 1 (ZFAS1) in sorafenib-resistant HCC cells.The increased expression of ZFAS1 is associated with enhanced stemness and EMT, contributing to a drug-resistant phenotype [128].
In cutaneous melanoma (SKCM), PRRT3-AS1, another lncRNA identified through scRNA-seq, was found to be highly expressed in advanced cases and associated with poor prognosis.This lncRNA promotes cancer cell migration, potentially through the EMT signaling pathway, and may influence immune cell infiltration, thereby affecting responses to immunotherapy [129].
Furthermore, scRNA-seq has been utilized to dissect the complex regulatory landscape of miRNAs during epithelial-to-mesenchymal transition (EMT).This research emphasized the role of the miR200 family in maintaining epithelial characteristics, highlighting their interaction with ZEB transcription factors.MiR101 was also noted for its role in preserving the epithelial phenotype by targeting EZH2, suggesting its potential as a therapeutic target 130].
These studies underscore the power of scRNA-seq in revealing the intricate functions of ncRNAs in regulating cell identity and their pivotal roles in disease processes, thereby opening new avenues for therapeutic intervention.

Deciphering lncRNA Functions: Computational Insights and Structural Revelations
The rapidly expanding field of lncRNAs is gaining significant attention for its potential in deciphering the regulatory architecture of the genome.LncRNAs, as critical modulators of gene expression and epigenetic regulation, are not only central to scientific exploration but also pivotal in identifying new therapeutic approaches.
LncRNAs deep involvement in vital cellular operations and disease pathways underscores the urgency of enhancing our comprehension of their functions.This complex landscape of lncRNA functionality calls for an interdisciplinary approach, combining traditional laboratory experimentation with computational innovation.Recent strides in computational biology have yielded advanced methods for identifying lncRNAs [217].For example, deep learning-based tools like 'lncRNA-Mdeep' have shown exceptional efficacy in distinguishing lncRNAs from coding sequences, extracting intricate features from sequence data [131].These tools have demonstrated superior performance in discriminating between lncRNAs and coding sequences by extracting complex features from sequence data [132,133].Similarly, 'lncRScan-SVM', employing SVM-based algorithms, has demonstrated notable accuracy in lncRNA identification, integrating both sequence and structural features and underscoring the importance of secondary structures in lncRNAs [134].
In terms of structural understanding, approaches like cross-linking and immunoprecipitation (CLIP) combined with HTS have illuminated lncRNA binding sites and interaction domains, revealing their spatial configurations and protein interactions [135,136].Additionally, the use of cryo-electron microscopy has been instrumental in revealing the three-dimensional structures of lncRNA complexes.A notable example is the elucidation of the compaction domain of the lncRNA XIST, which plays a crucial role in X-chromosome inactivation [137].
Computational predictions have also significantly enhanced our understanding of lncRNA functions.Tools like 'lncPath' use novel algorithms to predict lncRNA functions based on gene ontology term enrichment in co-expression networks, providing more precise insights into the roles of lncRNAs in biological pathways [138,139].'LncADeep', integrating deep learning, has been crucial in revealing the regulatory roles of numerous uncharacterized lncRNAs, especially in cancer contexts [140].DeepLncLoc, with its innovative approach to predicting lncRNA subcellular localization, has provided essential functional insights [141].Lastly, the updated LncBook 2.0 enhances lncRNA annotations with multi-omics data, furthering research into their biological roles and associations with diseases [142].
These developments underscore the dynamic nature of computational and structural studies in elucidating lncRNA functions.As we continue to improve these technologies, our understanding of lncRNA roles in health and disease is set to expand significantly, bridging current knowledge gaps and opening new avenues for therapeutic interventions.

The complex roles of NcRNAs in bladder cancer pathogenesis
NcRNAs have emerged as crucial elements in our understanding of bladder cancer, playing vital roles from the initial stages of tumor formation to the disease's metastatic spread.Their significant impact on the progression of bladder cancer is detailed in Table 3.
The progression of bladder cancer is influenced by a myriad of molecular pathways, with diverse RNA types playing critical roles.A notable instance is the role of microRNA-148b-3p, derived from exosomes of cancer-associated fibroblasts (CAFS), which significantly inhibits tumor growth.This inhibition is achieved by suppressing the Wnt/ β-catenin pathway and enhancing PTEN, both of which are key in tumor suppression [143].Conversely, the lncRNA SNHG14 exacerbates bladder cancer progression by interacting with the microRNA-211-3p/ESM1 axis [144].By sequestering microRNA-211-3p, SNHG14 induces an increase in ESM1, a protein that facilitates cell proliferation, migration, and invasion-essential processes in cancer development.Furthermore, the lncRNA PSMA3-AS1 acts as a competing endogenous RNA (ceRNA) for microRNA-214-5p, enhancing cell viability while reducing apoptosis, thereby contributing to tumor growth [145].CircRNAs also play a role in the progression of bladder cancer.For example, circRNA-0000326 promotes disease progression by activating the phosphoinositide-3 kinase/Akt pathway through the sequestration of microRNA-338-3p, thus supporting cell survival and growth [146].In contrast, the inhibition of circCEP128 and circRNA\_0000629 has been shown to impede bladder cancer progression, suggesting their potential as therapeutic targets [147,148].MicroRNA-381's involvement in bladder cancer progression is marked by its suppression of tumor growth and metastasis via downregulation of BMI1 and inactivation of the Rho/ROCK axis, critical pathways in cancer progression [149].
Bladder cancer invasion and metastasis, key determinants of prognosis and patient survival, are influenced by various ncRNAs.The lncRNA SNHG20, for instance, plays a notable role in advancing bladder cancer by activating the Wnt/β-catenin signaling pathway, a crucial driver of cell proliferation and tissue invasion [150].This underlines the  Facilitates prostate cancer progression and metastasis. [163] LOXL1-AS1 miR-let-7a-5p/EGFR Upregulation of lncRNA LOXL1-AS1 promotes cell proliferation and migration while inhibiting apoptosis.
provides a potential therapeutic strategy for patients with drug-resistant prostate cancer.
[164] PCAT6 IGF2BP2 and IGF1R Forms a complex with IGF2BP2 and IGF1R, stabilizing IGF1R mRNA and activating downstream pathways promotes bone metastasis and tumor growth in prostate cancer [165] (continued on next page) direct impact of ncRNAs on significant signaling pathways.Beyond influencing signaling pathways directly, ncRNAs contribute to bladder cancer progression through complex regulatory networks.For example, lncRNA SNHG1 promotes bladder cancer progression via interactions with miR-143-3p and the epigenetic modifier EZH2.This suggests a multi-faceted regulatory mechanism at play [151].Additionally, the overexpression of lncRNA plasmacytoma variant translocation 1 (PVT1) in multidrug-resistant urothelial bladder carcinoma tissues and cell lines is linked to increased cell proliferation, invasion, and chemoresistance [152].CircLPAR1, proposed as a biomarker for muscle-invasive bladder cancer, seems to regulate invasion and metastasis by interacting with miR-762, demonstrating the intricate interplay between ncRNAs and miRNAs in cancer progression [153].The lncRNA small nucleolar RNA host gene 3 (SNHG3) further exemplifies this complexity, promoting bladder cancer proliferation and metastasis through the miR-515-5p/-GINS2 axis [154].LncRNA-urothelial cancer-associated 1 (lncRNA-UCA1), prevalent in hypoxic tumor-derived exosomes, has been implicated in promoting bladder tumor growth and development [155].Lastly, lncRNA-ZEB2NAT, induced by TGF-$\beta$1 in CAFs, is involved in the epithelial-mesenchymal transition of bladder cancer cells, a crucial step in cancer metastasis [156].
The diverse functions of ncRNAs in bladder cancer highlight their potential as diagnostic biomarkers, prognostic indicators, and therapeutic targets.

Deciphering the multidimensional impact of NcRNAs in prostate cancer
NcRNA play an integral role in the complex process of carcinogenesis, steering the course from initiation to metastasis in prostate cancer.A plethora of evidence has identified numerous ncRNAs that are key players in directing these intricate processes within this disease [180][181][182].
For instance, microRNA-99b-5p in bladder cancer assumes multiple roles.It suppresses the expression of mTOR, a critical regulator of cellular growth and proliferation, thus inducing apoptosis and slowing cell proliferation.Additionally, this microRNA enhances the susceptibility of prostate cancer cell lines to the anti-cancer drug docetaxel, indicating its potential role in combating drug resistance.It also promotes autophagy, potentially leading to cell death through mTOR inactivation [157].Similarly, miR-145-5p acts as a significant player by inhibiting Phospholipase D5 (PLD5), a protein known for promoting cell proliferation and metastasis.This leads to reduced cell proliferation, invasion, and migration, and it also hinders the expression of metastasis-associated proteins like N-cadherin, E-cadherin, Vimentin, and Snail, while blocking the AKT pathway, essential for cell survival and growth [160].miR-124-3p specifically targets and reduces the expression of EZH2, a gene promoting cell proliferation and metastasis, thereby decreasing cellular proliferation, invasion, and migration [158].Similarly, overexpression of miR-221-5p is associated with reduced cell proliferation and cancer cell migration [161].Focusing on lncRNAs, PCAT6 has been identified as a key promoter of prostate cancer tumor growth, invasion, and metastasis.Its overexpression, particularly in prostate cancer with bone metastasis, correlates with worse prognosis.PCAT6 stabilizes the mRNA of IGF1R through its interaction with IGF2BP2 and IGF1R, activating pathways that encourage tumor growth and metastasis [165].In a similar mechanism, CASC11 enhances prostate cancer development by interacting with YBX1 and repressing the tumor suppressor gene p53 [167].Overexpression of lncRNAs PlncRNA-1 and SNHG11 in prostate cancer is also critical for cell proliferation and metastasis.Notably, SNHG11 boosts IGF-1R expression and recruits miR-184, leading to increased prostate cancer cell proliferation, invasion, and migration [163,166].The role of circRNAs is equally complex.CircHIPK3, for instance, functions as a sponge for miR-193a-3p, a tumor suppressor microRNA.Its upregulation in prostate cancer cells hinders miR-193a-3p's ability to suppress the oncogene MCL1, thereby promoting cell survival and disease progression [168].
In conclusion, the diverse roles of ncRNAs in prostate cancer, from tumor growth to chemoresistance, underscore their importance in disease progression and highlight their potential as targets for novel therapeutic strategies.Further research is necessary to fully unravel the complex interplay of these ncRNAs in prostate cancer.

NcRNAs: unraveling their intricate roles in RCC
The approach of ncRNAs as significant participants in the molecular schema of genitourinary cancers has necessitated re-considering their roles in cancer progression [183].The lncRNA PANDAR, for instance, is linked with poor prognosis in clear cell RCC (ccRCC), the most common RCC subtype.PANDAR interacts with transcription factors and regulatory proteins, altering the transcriptional landscape to promote tumorigenesis [173].In contrast, the lncRNA HEIRCC is associated with metastatic behavior in RCC, facilitating the spread of RCC cells through its role in EMT, a crucial process in metastasis [178].
Recent studies have identified a hypoxia-associated lncRNA signature in ccRCC, including lncRNAs like SLC16A1-AS1, which are thought to modulate the cellular response to hypoxia, a hallmark of RCC, thereby influencing tumor growth and progression [184,185].The lncRNA PCAT1 is known to fuel ccRCC by enhancing the transcriptional coactivator YAP.It acts as a molecular sponge for miR-656 and miR-539, miRNAs that usually suppress YAP expression, illustrating lncRNAs' role as competing endogenous RNAs in RCC [172].Similarly, lncRNA PCED1B-AS1 advances ccRCC progression through the miR-484/ZEB1 axis, hypothesized to sponge miR-484 and thereby alleviate the suppression of ZEB1, a key EMT inducer [171].It is hypothesized to sponge miR-484, thus relieving the suppression of ZEB1, a prominent EMT inducer.
Chemoresistance in RCC adds further complexity to treatment, as it allows cancer cells to persist and resist elimination.The lncRNA lncARSR, for instance, induces resistance to sunitinib, a standard therapeutic agent for RCC.It acts as a ceRNA, sequestering miR-34 and miR-449, which would typically inhibit the AXL and c-MET genes associated with drug resistance and cancer progression [175].Another lncRNA, SNHG12, promotes RCC progression and sunitinib resistance by upregulating CDCA3, a protein involved in cell cycle regulation, thus enhancing cancer cell proliferation and survival [170].HOTAIR contributes to sunitinib resistance in RCC by acting as a ceRNA for miR-17-5p, thereby preventing the microRNA from suppressing Beclin1, a gene linked to autophagy, a process associated with drug resistance.This leads to increased autophagy, fortifying the cancer cells' resistance to sunitinib [177].Furthermore, the lncRNA IGFL2-AS1 enhances TP53INP2 expression by binding competitively to hnRNPC, a protein that typically represses TP53INP2.The consequent upregulation of TP53INP2 promotes autophagy, leading to sunitinib resistance [176].lncRNA MALAT1, overexpressed in sunitinib-resistant RCC tissues and cells, acts as a ceRNA for miR-362-3p, regulating the expression of G3BP1, a gene associated with chemotherapy resistance.Knockdown of MALAT1 in sunitinib-resistant RCC reduces chemotherapy resistance, cell proliferation, and invasiveness, while enhancing apoptosis [174].Finally, the circRNA Eps15 hinders the action of miR-4731-5p, which usually suppresses ABCF2, a gene linked to drug resistance.By sponging miR-4731-5p, Eps15 increases ABCF2 expression, augmenting sunitinib resistance [179].
In conclusion, ncRNAs, particularly lncRNAs and circRNAs, are key to the development of chemoresistance in RCC.They offer potential targets for novel therapeutic strategies to overcome drug resistance and improve patient outcomes.

A comprehensive overview of the TME
The initiation and progression of cancer are significantly influenced by the TME, a complex and dynamic ecosystem comprising various cellular and non-cellular components.In a simplified view, the TME encompasses neoplastic cells, the extracellular matrix (ECM), immune cells, stromal cells, endothelial cells, and a range of signaling molecules (Fig. 1).These elements interact synergistically, establishing the TME as a crucial factor in cancer development [186].Central to the TME are angiogenic vascular cells, essential in new blood vessel formation (angiogenesis), facilitating tumor nourishment and expansion.The role of immune cells within the TME is dualistic; while some attack the tumor, others may unintentionally aid its growth.CAFs often accelerate tumor progression by remodeling the ECM and releasing growth factors.Moreover, acellular components like the ECM and tumor vasculature are integral to the TME.The ECM provides structural support and acts as a growth factor reservoir, while architectural changes in the ECM can stimulate tumor progression [187,188].The tumor vasculature, crucial for nutrient and oxygen supply, can create conditions like hypoxia and acidosis, promoting tumor growth and therapeutic resistance [189,190].Furthermore, extracellular physicochemical conditions such as altered pH, hypoxia, and fibrosis significantly impact the TME, influencing tumor dynamics and interactions with the surrounding environment.This leads to metastasis, immunosuppression, and drug resistance [191].
Within the TME, the Tumor Immune Microenvironment (TIME) comprises various immune cells that significantly affect tumor behavior through their interactions with cancer cells and other TME constituents [192,193].T cells within the TIME can exert anti-tumor effects, whereas cancer cells may manipulate the TIME to evade immune surveillance by upregulating immune checkpoint proteins [193,194].TAMs within the TIME can promote tumor growth and metastasis by secreting growth factors and pro-angiogenic factors and producing immunosuppressive cytokines, thus impeding anti-tumor immune responses [195].
The interplay of each component within the TME is instrumental in dictating cancer progression.Understanding these interactions is vital for developing effective cancer therapies.
Cancer cells, as central players in the TME, profoundly impact cancer progression.They orchestrate complex interactions within the TME, aiding their survival, proliferation, and spread.These cells secrete substances such as growth factors, cytokines, and chemokines, stimulating angiogenesis to ensure a steady supply of nutrients and oxygen for tumor growth [196].A prime example is the production of vascular endothelial growth factor (VEGF) by cancer cells, promoting endothelial cell proliferation and migration, leading to new blood vessel formation [197].Additionally, cancer cells manipulate the immune landscape of the TME.They attract and activate immune cells like TAMs, MDSCs, and Tregs, fostering an immunosuppressive environment conducive to tumor growth and facilitating immune evasion [198].For instance, cancer cells release Chemokine (C-C Motif) Ligand 2 (CCL2) to draw TAMs and MDSCs into the TME.These cells then produce anti-inflammatory cytokines such as IL-10 and TGF-β, suppressing the body's natural anti-tumor immune response [199].Cancer cells also contribute to the formation of a fibrotic and hypoxic TME.They produce ECM components and activate Hypoxia-Inducible Factors (HIFs), creating a protective barrier against immune detection while triggering metabolic changes that promote tumor growth and treatment resistance [200].Furthermore, cancer cells alter the function of stromal cells, such as CAFs and endothelial cells, through signaling molecule secretion.TGF-$ \beta$, secreted by cancer cells, activates CAFs, leading to ECM component production and growth factor release, enhancing tumor growth and metastasis [201].
In conclusion, cancer cells play a pivotal role in modulating the TME, significantly influencing cancer progression.

Exploring the TME's Influence on Urological Malignancies
The intricate relationship between the TME and urological cancers, including prostate, bladder, and kidney cancers, is a critical focus of contemporary cancer research.
Within the TME, fibroblasts and endothelial cells play pivotal roles.They secrete various growth factors and cytokines that are instrumental in fostering the growth, survival, and metastasis of cancer cells.In bladder cancer, for instance, CAFs significantly contribute to disease progression by releasing factors such as the Hepatocyte Growth Factor (HGF) and VEGF, which aid in tumor development and spread [202].
Immune cells within the TME also significantly influence the progression of urological malignancies.Tumor-infiltrating lymphocytes (TILs), comprising both CD8 + and CD4 + T cells, possess the capacity to target and eliminate cancer cells.However, in many cases of urological cancers, the effectiveness of these immune cells is diminished due to the immunosuppressive nature of the TME.For example, RCC typically harbors immunosuppressive cells like regulatory T cells (Tregs) and Myeloid-Derived Suppressor Cells (MDSCs), which hinder the activity of cytotoxic T cells and promote tumor progression [203].
Moreover, alterations in the ECM) within the TME, such as increased collagen deposition and crosslinking, play a significant role in cancer progression.In prostate cancer, these ECM modifications lead to a stiffer tumor structure [204].As suggested by Chevrier et al. [203], such changes may promote cancer cell invasion and metastasis by supporting cancer cell survival, proliferation, and immune evasion.Therefore, a comprehensive understanding of the TME's multifaceted influence on urological cancer progression is crucial.
The TME of urological cancers is a complex ecosystem comprising tumor cells, extracellular matrix, a variety of immune cells, stromal cells, and endothelial cells.These components interact through signaling molecules such as HIF-1α, Vascular Endothelial Growth Factor, IL-6, and IL-10, influencing tumor progression, immune evasion, and response to treatment in urological cancers.The figure was created with BioRender.com.

Elucidating NcRNA and the TME interactions in urological malignancies
Recent advances in cancer research have highlighted the significant role of ncRNAs in urological cancers such as prostate, bladder, and kidney cancers, particularly in the context of their interaction with the TME.The interplay between ncRNAs and the TME is crucial in cancer onset, development, and treatment resistance (Figure2, Table 4).

NcRNA-TME interactions in bladder cancer
The TME of bladder cancer showcases dynamic interactions between cancer cells and immune cells, notably involving circRNAs and lncRNAs.circFAM13B, for instance, overexpressed in bladder cancer, stimulates CD8 + T cells to secrete IFN-γ, enhancing immunotherapy effectiveness.It also interferes with IGF2BP1's binding to PKM2, reducing glycolysis in cancer cells [205].Another key molecule, LINC00702, inhibits bladder cancer cell growth and inflammatory TME development by recruiting JUND to promote DUSP1 gene transcription, essential for halting cancer progression.This lncRNA also reduces the release of inflammatory cytokines from M2-TAMs, which are known for aiding bladder cancer progression [206].Additionally, LINC01140, associated with muscle-invasive bladder cancer, correlates with FGF9 and promotes M2 macrophage polarization, facilitating cancer cell invasiveness and progression [207,208].In addition, ncRNAs also influence macrophage polarization and T cell exhaustion in the TME.For example, miR-21, found in exosomes from bladder cancer cells, impacts macrophage polarization by downregulating PTEN and activating the PI3K/AKT pathway, leading to enhanced M2 polarization and cancer progression [209].circTRPS1 in exosomes modulates T cell exhaustion by affecting PD-1 and Tim-3 expression on CD8 + T cells, contributing to immune evasion and tumor advancement [210].

NcRNA-TME interactions in RCC
In RCC, the TME presents a unique landscape where ncRNAs significantly affect cancer progression.miRNA-21-5p, encapsulated in exosomes released by M2 macrophages, promotes RCC metastasis by modulating the PTEN/AKT signaling pathway, essential for cancer cell survival and spread [211].In addition to miRNA-21-5p, miR-155 transported within extracellular vesicles (EVs) in the TME enhances RCC cell viability and contributes to tumor progression.Hypoxic conditions in the TME increase the release of these EVs, with miR-155 targeting FOXO3, a protein crucial for cell cycle regulation and apoptosis, thus accelerating RCC cell proliferation, invasion, and metastasis [212].

NcRNA-TME interactions in prostate cancer
In prostate cancer, the interplay between ncRNAs and the TME is critical for understanding disease progression and identifying potential therapeutic targets.CircRNA CircSMARCC1 influences the TME by modulating chemokine secretion.It binds to miR-1322, enhancing CCL20 secretion, which interacts with CCR6 on TAMs, facilitating tumor cell and TAM communication that promotes tumor progression [213].Another significant player is miR-149-3p, which directly impacts cancer cells and the surrounding TME.By downregulating the tumor suppressor DAB2IP, it activates NF-κB, a key effector in cancer cell invasion.This modification not only escalates the aggressiveness of cancer cells but also influences the cells' response to inflammatory cytokines present in the TME, fostering a pro-invasive environment [214].Furthermore, miR-95 plays a crucial role in the cross-talk between TAMs and cancer cells.Transferred via exosomes from TAMs to cancer cells, miR-95 downregulates JunB, a transcription factor essential for cell proliferation and apoptosis.This transfer promotes cancer cell proliferation, migration, and invasion, underlining the complex interplay within the prostate cancer TME [215].Lastly, the role of miR-423-5p, secreted by CAFs in exosomes, is noteworthy.Upon transfer to prostate cancer cells, it induces chemoresistance by targeting and downregulating GREM2 via the TGF-β signaling pathway.This interaction significantly affects the efficacy of chemotherapy drugs like docetaxel, taxane, and bicalutamide, presenting a challenging obstacle in treatment [216].
The complex interactions of ncRNAs within the TME of urological  Promotes PCa progression via exosome-mediated transfer [215] miR-423-5p CAFs Promotes chemotherapy resistance by targeting GREM2 through the TGF-β signaling pathway [216] cancers underscore their potential as therapeutic targets.While research has illuminated their role in tumor progression, metastasis, and drug resistance, a deeper understanding of these intricate relationships is necessary for developing novel treatments.This figure illustrates the complex interaction between ncRNAs and the TME in urological malignancies.It presents how specific ncRNAs affect cellular behavior and tumor progression by influencing immune responses, cell viability, migration and invasiveness within the TME.It highlights the pivotal role of ncRNAs in tumor development and their potential as therapeutic targets.The figure was created with BioRender.com.

Conclusion
This review article provides a comprehensive of the biogenesis and regulatory mechanisms of ncRNAs, with a particular focus on their varied roles within the TME.We delve into the complex interactions between ncRNAs and urological malignancies, unveiling their significant potential in cancer diagnosis and treatment.
Recent advances in HTS technologies and bioinformatics have substantially enriched our understanding of ncRNAs, elucidating their functions and intricate interactions within the TME.There is a growing body of evidence highlighting the crucial involvement of ncRNAs in the etiology of urological cancers.The application of ncRNAs in diagnostic and therapeutic strategies offers promising prospects for enhancing clinical outcomes.However, research into the complex relationship between ncRNAs and the TME in urological malignancies is still in its initial stages.A more expansive understanding of ncRNA's role in cancer progression and tumor development is needed.Currently, the majority of research focuses on single or limited numbers of ncRNAs, which restricts a complete grasp of their roles in cancer processes and the tumor environment in urological systems.
For ncRNAs to be effectively used in clinical settings for the diagnosis and treatment of urological malignancies, further experimental validation and methodological development are essential.

Fig. 1 .
Fig. 1.A Comprehensive Overview of the TME in urological malignancies.

Fig. 2 .
Fig. 2. The interaction of ncRNA with the TME in Urological Malignancies.

Table 1
The Timeline of ncRNA Studies in Cancer Research.

Table 2
Single-cell sequencing reveals the heterogeneity of urological malignancies.

Table 3
Influence of NcRNA on Proliferation, Metastasis, and Chemotherapy Resistance in urological malignancies.

Table 4
Impact of ncRNA-TME Interactions on Malignancy in Urological Malignancies.