Abstract

Objective

   This study aims to develop a ceRNA network associated with the
   chromosomal passenger complex (CPC) and identify a prognostic signature
   in lung cancer, the most diagnosed globally, to better understand the
   molecular mechanisms underlying tumor progression.

Methods

   The study used R packages and publicly available databases to conduct
   multi-omics In-silico analyses on CPC. These tools facilitated gene
   expression profiling, prognostic assessment, exploration of miRNA
   (microRNA), lncRNA (long non-coding RNA), transcription factor
   interactions, and pathway enrichment analysis. Molecular docking tools
   were used to evaluate binding affinities of CPC proteins with tobacco
   carcinogens, selective Aurora kinase B inhibitors, FDA-approved
   chemotherapeutics, and natural compounds. Immune landscape analysis was
   conducted using the SPRING viewer to visualize immune cell
   subpopulations in NSCLC, validated by correlation analysis using the
   GSCA database.

Results

   The study reveals that CPC genes—BIRC5, CDCA8, INCENP, and AURKB—are
   overexpressed in lung adenocarcinoma (LUAD) and are associated with
   poor overall survival, especially in smokers. A dysregulated ceRNA axis
   involving lncRNA TMPO-AS1 and miRNA-hsa-let-7b-5p was identified, along
   with transcription factor E2F1, which shows a strong correlation with
   the CPC genes. Notably, TMPO-AS1 and E2F1 are positively correlated,
   while hsa-let-7b-5p is negatively correlated with the CPCs,
   contributing to tumor progression. Downregulation of hsa-let-7b-5p is
   linked to poorer outcomes, highlighting its potential as a therapeutic
   target. Nicotine and NNK show stable binding, suggesting potential
   roles in activating the CPC pathway and contributing to LUAD
   progression. CPCs have a strong binding affinity with Hesperidin, a
   natural bioflavonoid, compared to known chemotherapeutic agents like
   docetaxel and paclitaxel. CPC genes are negatively correlated with CD4⁺
   T cells, indicating a role in promoting an immunosuppressive tumor
   microenvironment.

Conclusion

   Lung adenocarcinoma patients have poorer prognosis due to higher levels
   of CPCs, TMPO-AS1, and E2F1. A sponge complex between TMPO-AS1 and
   hsa-let-7b-5p may contribute to the tumor progression, and targeting
   CPCs with natural compounds could offer therapeutic potential.

   Highlights
    1. The overexpression of chromosomal passenger complex genes, AURKB,
       BIRC5, CDCA8, and INCENP is significantly associated with poor
       prognosis in lung adenocarcinoma (LUAD), particularly among
       smokers.
    2. The competing endogenous RNA (ceRNA) axis, which involves the long
       non-coding RNA TMPO-AS1 and the miRNA hsa-let-7b-5p, regulates the
       expression of these CPC genes. TMPO-AS1 shows a positive
       correlation with CPC genes, while hsa-let-7b-5p shows a negative
       correlation.
    3. Survival analysis indicates that the combined expression of CPC
       genes, TMPO-AS1, hsa-let-7b-5p, and E2F1 may serve as a reliable
       prognostic biomarker panel for LUAD in smokers.
    4. Hesperidin exhibits a strong binding affinity to CPC proteins,
       particularly AURKB, when compared to Barasertib, Docetaxel, and
       Paclitaxel, highlighting its potential as a therapeutic agent.
    5. The overexpression of CPC genes, E2F1, and TMPO-AS1 in LUAD is
       strongly associated with reduced infiltration of CD4⁺ T cells,
       indicating their role in promoting an immunosuppressive tumor
       microenvironment.

Graphical Abstract

   [38]graphic file with name 13008_2025_166_Figa_HTML.jpg
   [39]Open in a new tab

   Schematic representation of the regulatory axis involving TMPO-AS1,
   hsa-let-7b-5p, and CPC genes in lung tissue. A Normal lung
   microenvironment: Under physiological conditions, low expression of
   lncRNA TMPO-AS1 and high levels of miRNA hsa-let-7b-5p maintain
   controlled expression of CPC genes. The upregulated miRNA binds CPC
   transcripts, leading to their degradation or translational repression,
   thereby preserving chromosomal stability and supporting an active
   immune microenvironment with effective CD4^+ T cell infiltration. B
   Tumor lung microenvironment (LUAD associated with smoking): Chronic
   exposure to tobacco carcinogens leads to upregulation of TMPO-AS1 and
   downregulation of hsa-let-7b-5p. TMPO-AS1 sponges hsa-let-7b-5p,
   limiting its ability to suppress CPC transcripts, resulting in CPC
   overexpression. This promotes chromosomal instability, supports tumor
   growth, and contributes to the establishment of an immunosuppressive
   microenvironment characterized by reduced CD4^+ T cell infiltration

Supplementary Information

   The online version contains supplementary material available at
   10.1186/s13008-025-00166-w.

   Keywords: Lung adenocarcinoma, ceRNA network, Chromosomal passenger
   complex, Prognosis, Smokers, Carcinogens, Hesperidin, Immune evasion

Introduction

   Lung cancer remains a major global health concern, with 20 million new
   cases and 9.7 million deaths reported in 2022, according to the World
   Health Organization [[40]1]. Approximately 85% of lung cancer is
   classified as NSCLC, with lung adenocarcinoma (LUAD) and lung squamous
   cell carcinoma (LUSC) being the most common subtypes [[41]2]. Among
   these, Lung adenocarcinoma, has emerged as the most diagnosed cancer in
   younger men and women. Despite advances in treatment, the overall
   5-year survival rate for lung cancer remains low at 18%. However, early
   diagnosis can substantially improve outcomes, with survival rates
   rising to around 55% and up to 40% with effective treatment strategies
   [[42]3]. Notably, over 70% of NSCLC patients are diagnosed at an
   advanced-stage, rendering them ineligible for surgical intervention
   [[43]4]. This challenge is largely attributed to difficulties in early
   detection and lack of effective therapeutic targets for patients with
   NSCLC [[44]5]. Therefore, accurate identification of potential gene
   biomarkers and effective candidate drugs are essential for reducing
   mortality rates among the patients. Recent advances in gene expression
   profiling, driven by high-throughput sequencing technologies, have
   enabled the discovery of genetic signatures associated with early-stage
   lung cancer. These developments have laid the foundation for the
   identification of novel biomarkers that hold promises for enhancing
   diagnostic precision and guiding targeted therapies in NSCLC.

   Additionally, studies are increasingly examining the role of non-coding
   RNAs and epigenetic modifications, which may provide new paths for
   targeted therapies and improve the precision of diagnostic tools.
   Non-coding RNAs (ncRNAs), including miRNAs and lncRNAs, play crucial
   roles in gene regulation and have been implicated in the progression
   and development of NSCLC [[45]6]. They can function as oncogenes or
   tumor suppressors, influencing cell proliferation, apoptosis, and
   metastasis [[46]7]. Understanding the specific functions and mechanisms
   of these non-coding RNAs in tumor progression could lead to the
   development of novel biomarkers for early detection and new therapeutic
   targets for more personalized treatment approaches.

   Our research group has previously explored the regulatory axis
   involving hsa-let-7b-5p, TMPO-AS1, and E2F1 in the context of lung
   cancer, highlighting downstream targets such as FOXM1, MAD2L1, NCAPG,
   and NUSAP1 [[47]8–[48]10]. Building on these findings, the current
   study extends this axis to investigate its regulation of a critical
   molecular regulator of chromosomal integrity, the chromosomal passenger
   complex (CPC), which consists of Aurora B kinase, INCENP, Survivin, and
   Borealin, offering novel insights into mitotic control as it is crucial
   for genomic stability, regulating processes during both nuclear and
   cytoplasmic division [[49]4]. It ensures the accurate segregation of
   duplicated chromosomes during mitosis, prevents chromosome damage
   during cytokinesis, and manages proper cytoplasmic division to avoid
   tetraploidization [[50]11]. Aneuploidy and chromosomal instability
   (CIN) are prevalent in various human cancers, including lung cancer,
   but whether this is a result of CPC dysregulation remains uncertain.
   Several new gene expression targets are currently under investigation
   for combating cancer, particularly those associated with chromosomal
   instability, which is frequently linked to defective cell cycles and
   regulatory checkpoints, such as cyclin/cyclin-dependent kinases (CDKs).
   In the context of the cell cycle, cyclins and CDKs oversee four
   significant checkpoints: G0/G1, S phase, G2, and the M phase spindle
   assembly checkpoint (SAC). Abnormalities in these checkpoints can lead
   to improper chromosomal segregation and mitotic catastrophe,
   contributing to tumor aggressiveness [[51]12–[52]14].

   The role of CPC genes and their noncoding RNAs in cell cycle
   progression could lead to novel treatment options for various cancer
   types. Therefore, this study aims to investigate the prognostic role of
   CPC genes in LUAD patients using non- coding RNA-based therapeutics,
   including miRNAs and lncRNAs. The study employed In- silico methods to
   determine the ceRNA network associated with CPC genes and its role in
   the prognosis of LUAD patients. By utilizing In-silico methods, deeper
   insights into the molecular mechanisms underlying cancer progression
   can be gained, ultimately facilitating the development of more targeted
   and effective treatment strategies. By utilizing computational
   approaches, the study further explored the interaction of CPC proteins
   with various therapeutic agents, including known chemotherapeutics,
   natural compounds, and selective inhibitors. Additionally, the immune
   landscape of NSCLC to assess immune cell infiltration patterns and
   their potential association with CPC gene expression was investigated,
   ultimately aiding the identification of novel biomarkers and
   therapeutic targets for more effective and personalized treatment
   interventions.

Results

CPC genes exhibit prognostic significance in lung adenocarcinoma

   Given the role of CPC genes in cancer progression, we sought to
   investigate the clinical relevance of the CPC genes across TCGA
   cancers. We first analyzed their expression profiles using the UALCAN
   database and found that these genes are significantly overexpressed in
   tumor tissues compared to normal across most malignancies, as
   illustrated in Supplementary Fig. 1A–D. Consistent with this,
   pan-cancer analyses using the Firehose Broad GDAC and TIMER databases
   also revealed elevated expression of CPC genes across various cancer
   types (Supplementary Fig. 2A–D and 3A–D), suggesting their potential
   oncogenic roles. Further, OncoMX database analysis confirmed
   significantly high expression of CPC genes in lung cancer, with log2
   fold changes of 4.32 for BIRC5, 3.54 for CDCA8, 1.59 for INCENP, and
   4.03 for AURKB (Supplementary Table 1). Furthermore, to explore their
   prognostic significance, we utilized the KM Plotter database to analyze
   survival outcomes in NSCLC patients. Our results indicated that higher
   BIRC5, CDCA8, INCENP, and AURKB expression in tumor tissues correlated
   with poor patient prognosis and was significantly associated with
   reduced overall survival (OS) based on hazard ratios (HR) and
   confidence intervals (CI) as follows: BIRC5 (HR = 1.49, CI: 1.33–1.68,
   P = 2.9e−11), CDCA8 (HR = 1.59, CI: 1.41–1.79, P = 2.2e−14), INCENP
   (HR = 1.38, CI: 1.22–1.55, P = 1.4e−07), and AURKB (HR = 1.78, CI:
   1.58–2.01, P < 1e−16), as shown in Fig. [53]1A–D. As shown in
   Table [54]1, overexpression of CPC genes substantially reduces patient
   survival. For example, low BIRC5 expression was associated with a
   median survival of 86 months (~7 years), while high expression reduced
   survival to 51.27 months (~4.2 years), indicating a strong correlation
   between elevated BIRC5 expression and poor prognosis in NSCLC patients.

Fig. 1.

   [55]Fig. 1
   [56]Open in a new tab

   Prognostic role of mRNA expression of the chromosomal passenger complex
   (CPC) genes in lung cancer patients. Kaplan–Meier survival curves were
   plotted for BIRC5, CDCA8, INCENP and AURKB (STK12), respectively for
   the parameters consisting of A–D Overall Survival (OS) (n = 2166), E–H
   OS+ Lung Adenocarcinoma (LUAD) (n = 1161), I–L OS + LUAD + Males
   (n = 566), M–P OS + LUAD + Females (n = 537), and Q–T
   OS + LUAD + Smokers (n = 546)

Table 1.

   Association of chromosomal passenger gene expression with the survival
   outcome of patients
   S. No. Gene Index Patient number Hazard ratio CI Log(P) Low expression
   cohort (months) High expression cohort (months)
   1 BIRC5 (survivin) OS 2166 1.49 1.33–1.68 2.9e−11 86 51.27
   FP 1252 1.81 1.52–2.15 6.7e−12 28.7 11
   PPS 477 1.4 1.13–1.72 0.0015 19.47 10.38
   2 CDCA8 (borealin) OS 2166 1.59 1.41–1.79 2.2e−14 94.5 48
   FP 1252 1.79 1.51–2.13 1.4e−11 29 11
   PPS 477 1.42 1.16–1.76 0.00086 21 10
   3 INCENP OS 2166 1.38 1.22–1.55 1.4e−07 86 54
   FP 1252 1.78 1.5–2.11 2.8e−11 27 11
   PPS 477 1.07 0.87–1.32 0.53 15.62 12
   4 AURKB OS 2166 1.78 1.58–2.01 <1e−16 93 43.83
   FP 1252 1.85 1.56–2.2 8.3e−13 31.61 10
   PPS 477 1.6 1.3–1.98 8.3e−06 21.9 9
   5 BIRC5+CDCA8+ICENP+AURKB OS 2166 1.63 1.45–1.84 6.6e−16 93 48
   FP 1252 1.79 1.51–2.13 1.3e−11 28.7 10.3
   PPS 477 1.36 1.1–1.68 0.0037 20.11 10.38
   Histology
   1 BIRC5 (survivin) Adenocarcinoma 1161 1.96 1.65–2.34 1.5e−14 112.67 52
   Squamous cell carcinoma 780 1.01 0.83–1.22 0.92 52 62.2
   2 CDCA8 (borealin) Adenocarcinoma 1161 1.66 1.39–1.98 1e−08 106 59
   Squamous cell carcinoma 780 1.02 0.84–1.23 0.87 56 54
   3 INCENP Adenocarcinoma 1161 1.57 1.32–1.86 2.8e−07 110.27 66
   Squamous cell carcinoma 780 0.98 0.81–1.19 0.84 58 53
   4 AURKB Adenocarcinoma 1161 2.11 1.77–2.52 <1e−16 117.33 49.43
   Squamous cell carcinoma 780 1.19 0.98–1.45 0.073 62.47 49.97
   5 BIRC5+CDCA8+INCENP+AURKB Adenocarcinoma 1161 1.92 1.61–2.28 1.6e−13
   110.27 52
   Squamous cell carcinoma 780 1.07 0.88–1.3 0.49 58 52.97
   Gender
   1 BIRC5 (survivin) Male 566 1.7 1.34–2.15 8e−06 105 52
   Female 537 2.29 1.73–3.05 3.3e−09 117.33 65
   2 CDCA8 (borealin) Male 566 1.67 1.32–2.12 1.8e−05 103 48
   Female 537 1.6 1.21–2.1 0.00083 110.27 86
   3 INCENP Male 566 1.41 1.12–1.79 0.0039 99 59
   Female 537 1.86 1.41–2.44 8.2e−06 119.87 71
   4 AURKB Male 566 1.59 1.37–1.84 8.5e−10 74 41.37
   Female 537 2.08 1.67–2.6 4.2e−11 119.87 62.47
   5 BIRC5+CDCA8+INCENP+AURKB Male 566 1.73 1.36–2.19 4.9e−06 103 46
   Female 537 2.14 1.61–2.85 6.8e−08 116 68.67
   Smoking history
   1 BIRC5 (survivin) Smoker 546 1.85 1.42–2.41 4.1e−06 116 63
   Non-smoker 192 2.14 1.15–3.99 0.014 80 49
   2 CDCA8 (borealin) Smoker 546 1.69 1.29–2.2 9.5e−05 95 62
   Non-smoker 192 1.18 0.65–2.13 0.58 76 69
   3 INCENP Smoker 546 1.96 1.5–2.57 5.5e−07 116 61
   Non-smoker 192 1.35 0.75–2.44 0.32 75.43 76
   4 AURKB Smoker 546 1.78 1.44–2.19 4.3e−08 95 54.3
   Non-smoker 192 2.41 1.31–4.46 0.0037 76 49
   5 BIRC5+CDCA8+INCENP+AURKB Smoker 546 1.92 1.47–2.5 1.1e−06 96 62
   Non-smoker 192 1.55 0.85–2.82 0.15 76 52
   [57]Open in a new tab

   Similarly, high expressions of CDCA8, INCENP, and AURKB also correlated
   with decreased survival. Given the clinical relevance, we examined CPC
   gene expression in histological subtypes and other clinicopathological
   factors. In LUAD patients, elevated expression of BIRC5 (HR = 1.96, CI:
   1.65–2.34, P = 1.5e−14), CDCA8 (HR = 1.66, CI: 1.39–1.98, P = 1e−08),
   INCENP (HR = 1.57, CI: 1.32–1.86, P = 2.8e−07), and AURKB (HR = 2.11,
   CI: 1.77–2.52, P < 1e−16) was significantly associated with poor OS
   (Fig. [58]1E–H). However, no significant association was observed in
   LUSC patients: BIRC5 (P = 0.97), CDCA8 (P = 0.87), INCENP (P = 0.84),
   and AURKB (P = 0.073), as depicted in Supplementary Fig. 4A–D and
   Table [59]1. The magnitude of survival differences in LUAD patients
   further underscores the prognostic value of CPC genes. For instance,
   BIRC5 showed a 2.1-fold difference in OS (112.67 vs. 52 months), CDCA8
   had a 1.7-fold difference (106 vs. 59 months), INCENP showed a 1.6-fold
   reduction (110.27 vs. 66 months), and AURKB exhibited the most
   significant impact with a 2.3-fold difference (117.33 vs.
   49.43 months). Consequently, we found that the expression of CPC genes
   was significantly associated with both males and females LUAD patients
   (Table [60]1 and F[61]ig. [62]1I–P). Similarly, the expression of CPC
   genes was significantly associated with the poor survival outcome of
   smokers with LUAD (BIRC5: HR = 1.85, CI: 1.42–2.41, P = 4.1e−06),
   (CDCA8: HR = 1.69, CI: 1.29–2.2, P = 9.5e−05), (INCENP: HR = 1.96, CI:
   1.5–2.57, P = 5.5e−07), (AURKB: HR = 2.1, CI: 1.61–2.75, P = 3.3e−08)
   (Fig. [63]1Q–T). A decline in survival outcome of approximately 1.5- to
   2-folds from low expression cohort to high expression cohort in smoker
   LUAD patients was noted (Table [64]1).

   Notably, as shown in Supplementary Fig. 4E–P, we found that the KM
   plotter analysis with similar clinicopathological characteristics in
   LUSC did not correlate with the CPC gene expression. To further
   corroborate our results a multivariate analysis taking mean expression
   of all the four genes was carried out to assess the prognostic
   significance of the CPC complex as a whole and the survival plots
   showed significant overexpression of these associated with poor OS
   (HR = 1.63, P = 6.6e−16, low expression = 93 months, high
   expression = 48 months), OS + LUAD (HR = 1.92, P = 1.6e−13, low
   expression = 110.27 months, high expression = 52 months),
   OS + LUAD + Males (HR = 1.73, P = 4.9e−06, low expression = 103 months,
   high expression = 46 months), OS + LUAD + Females (HR = 2.14,
   P = 6.8e−08, low expression = 116 months, high
   expression = 68.67 months), and OS + LUAD + Smokers (HR = 1.92,
   P = 4.3e−08, low expression = 95 months, high
   expression = 54.3 months), whereas insignificant results were found in
   the cases of OS + LUSC (HR = 1.07, P = 0.49, low
   expression = 58 months, high expression = 52.97 months) and
   OS + LUAD + Non-Smokers (HR = 1.55, P = 0.15, low
   expression = 76 months, high expression = 52 months) as shown in
   Table [65]1 and Supplementary Fig. 5A–G.

Expression of CPC genes in human lung adenocarcinoma

   To further explore their role in cancer conditions, we next assessed
   the expression patterns of CPC genes specifically in LUAD patient
   samples. Analysis using the UALCAN database revealed that tumor from
   LUAD patients had greater levels of BIRC5, CDCA8, INCENP, and AURKB
   gene expression, with an approximately ninefold increase (P < 0.05)
   when compared to normal tissues (Fig. [66]2A–D). Further, the log fold
   change in expression levels of BIRC5, CDCA8, INCENP, and AURKB genes in
   tumor samples was analyzed using ENCORI (Log FC: 14.08, 9.16, 2.60,
   11.93, respectively) as shown in Fig. [67]2E–H. Additionally, to ensure
   an unbiased comparison, we normalized the sample size using R packages
   (Normal = 59, Tumor = 59), the box plots revealed a significant
   overexpression of CPC genes in LUAD tumor tissues compared to normal
   tissues (P < 0.05) as shown in F[68]ig. [69]2I–L. Furthermore,
   Single-cell transcriptome analysis using TCGAnalyzeR revealed elevated
   Log FC values for BIRC5 (3.50), CDCA8 (2.18), INCENP (1.15), and AURKB
   (3.06), indicating their mRNA transcript upregulation in LUAD tumor
   cells as shown in Fig. [70]2M–P. The data suggests that these genes
   could play a significant role in the progression of LUAD. Further,
   UALCAN was used to evaluate CPC gene expression in LUAD across various
   clinicopathological parameters, including smoking status as shown in
   Fig. [71]2Q–T, gender, tumor stage, and nodal metastasis (Supplementary
   Fig. 6A–L). The results showed a significant upregulation (P < 0.05) of
   all CPC genes, particularly in male smokers, as well as in patients
   with advanced stages and positive nodal metastasis, reinforcing their
   potential clinical significance.

Fig. 2.

   [72]Fig. 2
   [73]Open in a new tab

   Differential mRNA expression analysis of BIRC5, CDCA8, INCENP, and
   AURKB, respectively, in LUAD. Using the A–D UALCAN database, E–H ENCORI
   database, I–L R packages, M–P TCGAnalyzeR database and expression
   analysis of these genes based on patients’s smoking habits using the
   Q–T UALCAN database

   Notably, tobacco smoke remains a leading cause of lung carcinogenesis,
   with nicotine and its derivative NNK recognized as key carcinogens. To
   assess their potential impact on mitotic regulation, molecular docking
   was performed with CPC proteins. The binding affinities (kcal/mol)
   observed for nicotine were: BIRC5 (−5.3), CDCA8 (−4.5), INCENP (−4.1),
   and AURKB (−6.0); and for NNK: BIRC5 (−5.5), CDCA8 (−4.4), INCENP
   (−3.3), and AURKB (−6.6) (Supplementary Table 2). Further, the
   interaction profiling revealed predominant π–π stacking and van der
   Waals forces (Supplementary Fig. 7A–H), suggesting that these
   tobacco-derived compounds may contribute to chromosomal instability and
   mitotic disruption in LUAD through direct CPC targeting, possibly
   underlying the altered expression patterns observed in smokers.

CPC genes are involved in biological processes and metastasis

   Next, to understand how dysregulation of CPC genes may influence
   cellular behavior, we investigated their role in key biological
   processes and metastatic potential in LUAD. The CancerSEA database
   revealed a significant correlation between gene expression and various
   biological processes involved in LUAD. BIRC5 displayed a significant
   correlation with the cell cycle (R = 0.80), DNA damage (R = 0.68),
   proliferation (R = 0.64), DNA repair (R = 0.61), invasion (R = 0.47),
   and EMT (R = 0.30). CDCA8 was significantly associated with the cell
   cycle (R = 0.59), proliferation (R = 0.54), DNA damage (R = 0.39), DNA
   repair (R = 0.33), and invasion (R = 0.33). INCENP showed a significant
   association with the cell cycle (R = 0.32) only, while AURKB was
   involved in the cell cycle (R = 0.69), DNA damage (R = 0.56), DNA
   repair (R = 0.52), proliferation (R = 0.50), invasion (R = 0.43),EMT
   (R = 0.31), Stemness (R = 0.25), and metastasis (R = 0.23)
   (Supplementary Fig. 8A–D). These results indicate that an increased
   level of CPC genes in LUAD cells facilitates metastasis and the
   transition from epithelial to mesenchymal cells, thereby achieving
   resistance to available therapies. Moreover, the TNMplot database
   confirmed significant upregulation of BIRC5, CDCA8, INCENP, and AURKB
   in metastasis, with P values of BIRC5: 1.09e−89, CDCA8: 5.17e−135,
   INCENP: 5.37e−20, and AURKB: 8.73e−84 (Supplementary Fig. 8E–H). These
   findings suggest that CPC genes could serve as biomarkers for early
   LUAD detection and prognosis. Their upregulation positions them as
   promising targets for therapies aimed at inhibiting metastasis,
   improving patient outcomes, and enhancing the efficacy of existing
   treatments by overcoming resistance.

Regulatory network of lncRNAs, miRNAs, and CPC genes identified

   Furthermore, emerging evidence highlights the key role of competitive
   endogenous RNA networks in orchestrating gene expressions in cancer by
   enabling the crosstalk between lncRNAs, miRNAs, and mRNAs. In this
   loop, the lncRNAs and mRNAs, competitively binds to the shared miRNA,
   modulating its availability. Given the dynamic overexpression of CPC
   genes in LUAD, we hypothesize that post-transcriptional regulation via
   the ceRNA network may contribute to this dysregulation. Hence, to
   determine the miRNAs associated with CPC genes in LUAD patients, the
   miRNet database was used and a total of 41 miRNAs were found in the
   network. The UALCAN database was used to analyze their expression level
   in LUAD samples, where we aimed to identify downregulated miRNAs,
   indicating a plausible direct regulation between both. The
   downregulated miRNAs with a significant P value (<0.05) were selected
   for further analysis (Supplementary Table 3).

   Interestingly, only 3 out of 41 miRNAs: hsa-let-7b-5p, hsa-let-7g-5p,
   and hsa-mir138-5p were found to be significantly downregulated in LUAD.
   Further correlation analysis of these miRNAs (hsa-let-7b-5p,
   hsa-let-7g-5p, and hsa-mir138-5) with the CPC genes using the ENCORI
   database revealed that, only hsa-let-7b-5p negatively correlated with
   all the genes: BIRC5 (R = −0.361), CDCA8 (−0.362), INCENP: (−0.309),
   and AURKB: (−0.448), as shown in Supplementary Table 4 and
   Fig. [74]3A–D. Surprisingly, using the concentric circle analysis in
   the miRNet database, only hsa-let-7b-5p was shown to be in direct
   proximity with all the CPC genes (Supplementary Fig. 9A), strengthening
   our data. Consequently, to further assess this correlation, binding
   affinities between hsa-let-7b-5p and the CPC were analyzed using
   miRWalk database, which revealed highly favorable binding affinities
   ranging from −18.6 to −21.6 kcal/mol across BIRC5, CDCA8, INCENP, and
   AURKB transcripts. Further, complementary structural interaction
   predictions using the RNA22v2 algorithm confirmed stable folding
   energies, with the minimum free energy of the heteroduplex structures
   falling between −15.40 and −18.10 kcal/mol, as shown in Table [75]2.

Fig. 3.

   [76]Fig. 3
   [77]Open in a new tab

   Regulatory mechanism based on ceRNA network analysis consisting of
   mRNA/miRNA/lncRNA correlation and differential expression. Using the
   ENCORI database to analyze the correlation between A hsa-let-7b-5p vs.
   BIRC5, B hsa-let-7b-5p vs. CDCA8, C hsa-let-7b-5p vs. INCENP, and D
   hsa-let-7b-5p vs. AURKB. Differential miRNA expressions using E
   CancerMIRNome, F UALCAN and G R packages. H Expression of hsa-let-7b-5p
   in LUAD based on smoking status by using UALCAN. I Area under the curve
   for hsa-let-7b-5p in LUAD. Survival analysis of J hsa-let-7b-5p in
   LUAD, K TMPO-AS1 in OS + LUAD, and L TMPO-AS1 in OS + LUAD + Smokers
   using KM Plotter. lncRNA differential expression analysis using M
   ENCORI N UALCAN and O R packgaes. Correlation analysis using ENCORI
   between P TMPO-AS1 vs. BIRC5, Q TMPO-AS1 vs. CDCA8, R TMPO-AS1 vs.
   INCENP, S TMPO-AS1 vs. AURKB, and T hsa-let-7b-5p vs. TMPO-AS1. B–O was
   previously published in Saini et al. [8], and is reused here with
   appropriate citation and acknowledgment for conceptual continuity