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LncRNA SPINT1-AS1 enhances the Warburg effect and promotes the progression of osteosarcoma via the miR-135b-5p/PGAM1 axis

Cancer Cell International volume 25, Article number: 124 (2025) Cite this article

Abstract

Osteosarcoma (OS) is a malignant bone tumor that originates from interstitial tissues and affects the health of children and adolescents. Long noncoding RNAs (lncRNAs) are an intriguing category of widely distributed endogenous RNAs involved in OS progression, many of which remain functionally uncharacterized. Herein, we observed an increased expression of lncRNA SPINT1-AS1 in OS tissues and cell lines. Further analysis confirmed that SPINT1-AS1 promotes the proliferation and metastasis of OS cells. Moreover, miR-135b-5p was identified as a downstream target of SPINT1-AS1. Through bioinformatics analysis, PGAM1 mRNA was validated as a target of miR-135b-5p via RIP and luciferase reporter assays. SPINT1-AS1 could enhance OS cell proliferation and metastasis by promoting aerobic glycolysis, acting as a ceRNA by binding to miR-135b-5p, thereby increasing PGAM1 expression. Taken together, these results indicate that SPINT1-AS1 functions as a tumor promoter and regulates OS cell progression through the miR-135b-5p/PGAM1 axis.

Introduction

Osteosarcoma (OS) is one of the most common malignant bone tumors originating from interstitial tissues. It has a high incidence in children and adolescents, often affecting the long bones, including the legs and arms [1]. Treatment options for osteosarcoma include chemotherapy, radiation therapy, surgery, and immunotherapy [2, 3]. Although early detection and standard treatments have improved the survival rate, drug resistance and distant recurrence result in a poor prognosis for some patients [4]. Therefore, understanding the molecular mechanisms underlying OS genesis and progression, as well as developing new techniques for its detection and treatment, is crucial.

Long noncoding RNAs (lncRNAs) have been shown to play a vital role in cancer proliferation and development [5]. These lncRNAs are transcripts longer than 200 bp with limited or no protein-coding potential. They are widely distributed throughout the human genome and exhibit diverse structures and functions [6, 7]. Recent research has demonstrated that lncRNAs play a significant role in the progression of various human cancers, including OS. For instance, lncRNA HCG18 promotes osteosarcoma growth by acting as a competing endogenous RNA (ceRNA) for miR‑365a‑3p [8], while LINC01123 enhances OS cell proliferation by downregulating miR-516b-5p expression [9]. Recent studies have also identified lncRNA SPINT1-AS1 as a potential oncogene in colorectal cancer [10,11,12]. However, its role and expression in OS remain unknown.

Metabolic reprogramming is a hallmark of many cancers [13]. A well-known metabolic feature of cancer cells is the Warburg effect, which refers to the preference of cancer cells to metabolize large amounts of glucose into lactate, even in the presence of sufficient oxygen, a process known as aerobic glycolysis [14]. Several metabolic enzymes and signaling molecules involved in glucose metabolism have been reported to play critical roles in carcinogenesis and invasion [15]. Phosphoglycerate mutase 1 (PGAM1) is a key enzyme involved in cancer metabolism and has been identified as a cancer promoter with significant impacts on various cancers, including breast cancer, uveal melanoma, prostate cancer, and ovarian cancer [16,17,18,19,20]. Previous studies have shown that microRNA-324-3p inhibits PGAM1-mediated aerobic glycolysis in OS cells [21]. Nonetheless, further research on the role of PGAM1 in osteosarcoma progression has been limited. Herein, we uncovered that SPINT1-AS1 is upregulated in OS tissues and cell lines, promoting OS proliferation and metastasis by enhancing aerobic glycolysis. Mechanistically, SPINT1-AS1 functions as a competitor for miR-135b-5p with PGAM1, alleviating miR-135b-5p's repressive effect on PGAM1, thereby increasing PGAM1 expression. This SPINT1-AS1/miR-135b-5p/PGAM1 axis may provide a theoretical foundation for identifying new therapeutic targets in the treatment of osteosarcoma.

Materials and methods

Cells lines and culture condition

Three human OS cell lines, 143B(CRL-8303), MNNG/HOS(CRL-1547), SAOS-2(HTB-85) and U2OS(HTB-96), were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). The human osteoblast hFOB1.19 cell line and two additional human OS cell lines, MG63 and SAOS-2 were supplied by the Cell Bank of the Chinese Academy of Sciences (Shanghai, China). Briefly, hFOB1.19 cells were maintained at 34.5 °C under a 5% CO2 environment, while the human OS cell lines were maintained at 37 °C under similar conditions with 5% CO2.

Quantitative RT-PCR assay

Following the instructions provided in the user manual, the TRIzol (Invitrogen) was applied to isolate the total RNA from OS cells and tissues. The procedures were conducted as previously described [9]. Relative mRNA expression levels were determined using the 2−ΔΔCt method, with β-actin as the internal control. The primers used were as follows: SPINT1-AS1 sense: 5′-GCCCTGGAGGATGAGAG-3ʹ and anti-sense: 5′-CAGATGCTGTTGGCTAAAGA-3′; β-actin sense: 5′-UUCUCCGAACGU GUCACGUTT-3ʹ and anti-sense: 5′-ACG UGACACGUUCGGAGAAT-3′; miR-135b-5p sense: 5′-CGCGTATGGCTTTTCATTCCT-3′ and anti-sense: 5′-AGTGCAGG GTCCGAGGTATT-3′; PGAM1 sense: 5′-TTGAATACAGCGACCCAGTGG-3′ and anti-sense: 5′-CTATCGATGTACAGCCGAATGGTG-3′.

Cells transfection

Transfection was conducted using Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA) following the manufacturer's instructions. The miR-135b-5p mimic, control inhibitor, miR-135b-5p inhibitor, inhibitor NC, pcDNA-PGAM1, and pcDNA-NC were all obtained from Gene-Pharma (Shanghai, China). To generate stably transfected osteosarcoma cell lines, lentiviral infection was utilized. The plasmid for SPINT1-AS1 knockdown (sh-SPINT1-AS1) and the empty plasmid (sh-CON) were also sourced from Gene-Pharma. Stable SPINT1-AS1-knockdown cell lines were established by co-infecting target cells with 1 × 108 lentiviral transducers and polybrene (Sigma-Aldrich, St. Louis, MO, USA). Seventy-two hours post-infection, infected cells were selected with puromycin at a concentration of 2.5 μg/mL.

Biotin RNA pull-down assay

The SPINT1-AS1 biotin probe was obtained from Genepharma (Shanghai, China) and transfected into U-2OS and 143B cells for 48 h. The cell lysates were then incubated with Dynabeads M-280 Streptavidin (Sigma, MO, USA). Finally, RT-qPCR was performed to measure the expression levels.

RNA immunoprecipitation (RIP) assay

The interactions among SPINT1-AS1, miR-135b-5p, and PGAM1 were detected using RIP assays. The steps of experiment were carried out in accordance with the user’s manual.

Fluorescence in situ hybridization (FISH)

A microarray containing tissue from 40 OS patients was provided by Alena Biotechnology Co., Ltd. (Xi'an, China). The OS tissue samples were primary tumors. OS tissue sections were hybridized with SPINT1-AS1 probes sourced from Servicebio (Wuhan, China). FISH was carried out as performed previously [22].

Cell proliferation, colony formation and migration assays

Cell proliferation was evaluated using a Cell Counting Kit-8 (CCK-8) assay (Solarbio, China) as previously described [9]. For colony formation, 500 cells were plated in a 6-well plate and stained after a period of 14 days. Colonies containing more than 30 cells were scored. We used the transwell and wound healing assays to inspect cell migration as mentioned previously [8].

Dual‑luciferase reporter assay

The dual-luciferase reporter assay was utilized to measure the relationships among SPINT1-AS1, miR-135b-5p, and PGAM1 according to the user’s manual as described previously [8].

Western blotting

Western blotting was conducted following the procedure previously showed in the article [9]. First, total protein was extracted using RIPA buffer (Sigma, USA) containing phenylmethylsulfonyl fluoride. The protein samples underwent electrophoresis using 12% SDS-PAGE gels before being transferred onto polyvinylidene difluoride (PVDF) membranes (Millipore, Bedford, MA, USA). The membrane was incubated with primary antibodies on the shaker at 4 °C all night. The PVDF membrane was then washed with by TBST and kept incubating with secondary antibody at room temperature for 1 h. Ultimately, enhanced chemiluminescence((ECL) (Proteinbio, Nanjing, China) was used for detection. The primary antibodies used in this experiment included anti-beta actin (Abcam, #ab6276, 1:5000) and anti-PGAM1 (Abcam, #ab129191, 1:1000).

In vivo xenograft assay

We employed an in vivo xenograft model to study the growth of OS cells. Five- to seven-week-old male nude BALB/c mice were randomly divided into two groups (n = 3). A total of 1.5 × 106 SPINT1-AS1 silenced or control cells were injected subcutaneously into the back of the mice. Tumor growth was monitored every 5 days. After 20 days, the mice were sacrificed, and the subcutaneous tumors were collected and measured. The experiments were authorized by the ethics committee of the Affiliated Hospital of Jiangsu University.

Measurement of oxidative phosphorylation and glycolysis

Following manufacturer instructions for an XF96 metabolic flux analyzer (Seahorse Biosciences; Billerica MA), the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) of different transfected OS cells were detected.

TUNEL assay and immunohistochemical staining

The method applied to detect the apoptosis of transfected OS cells in the xenograft tumors followed the procedure previously showed in the article [9]. Ki67 was detected by using a primary antibody (1:200, GB13030; Servicebio, Wuhan, China).

Measurement of glucose consumption and lactate production

The cellular glucose consumption and lactate production were carried out as previously described [23]. A lactate assay kit (BioVision, USA) was utilized to measure extracellular lactate levels.

Statistical analysis

Statistical analyses were conducted using GraphPad Prism 9.0 software and SPSS 26.0. Data are presented as mean ± standard deviation. Differences between paired groups were analyzed using a two-tailed Student’s t-test after confirming the homogeneity of variances. All experiments were performed at least three times, and a P value of 0.05 was considered statistically significant.

LncRNA SPINT1-AS1 was upregulated in OS tissues and cell lines

Initially, we measured the differences in SPINT1-AS1 expression in OS tissue samples (n = 40) using fluorescence in situ hybridization. The results indicated that elevated SPINT1-AS1 expression was associated with more advanced pathological stages of OS (Fig. 1A, B). Furthermore, as shown in Fig. 1C, the expression of SPINT1-AS1 was significantly higher in OS cell lines compared to human osteoblast cells. Collectively, these data demonstrate that SPINT1-AS1 is upregulated in OS tissues and cell lines.

Fig. 1
figure 1

LncRNA SPINT1-AS1 exhibited elevated expression levels in OS tissues and cell lines. A, B the expression of SPINT1-AS1 in OS (n = 40) tissue. *p < 0.05 (chi-square test). C Comparison of SPINT1-AS1 expression between OS cell lines and hFOB1.19 cells. **p < 0.01; ***p < 0.001 (Student's t-test)

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SPINT1-AS1 silencing suppressed the cell proliferation and metastasis of OS

To investigate the biological function of SPINT1-AS1 in OS, we first silenced SPINT1-AS1 in 143B and U-2OS cell lines. The knockdown efficacy was assessed by qRT-PCR (Fig. 2A). CCK8 assays (Fig. 2B, C) and colony formation assays (Fig. 2D) revealed that SPINT1-AS1 knockdown partially restrained the proliferation of OS cells. Subsequently, silencing SPINT1-AS1 significantly suppressed the migration of OS cells (Fig. 2E–G). The results of Transwell assays confirmed that SPINT1-AS1 silencing inhibited cell invasion (Figure S2A, B) in OS cells. Moreover, SPINT1-AS1 downregulation markedly reduced xenografted tumor volume and weight in nude mice (Fig. 2H–J). Tumor section staining indicated a decline in Ki67 expression and an increase in the apoptosis rate in the SPINT1-AS1-knockdown group compared to the control group (Fig. 2K). The knockdown efficiency of SPINT1-AS1 was also confirmed by qRT-PCR (Fig. 2L). Collectively, these findings suggest that silencing SPINT1-AS1 inhibits cell proliferation and metastasis in OS.

Fig. 2
figure 2

SPINT1-AS1 depletion suppressed the progression of OS. A Efficiency of SPINT1-AS1 knockdown in OS cells. B, C The proliferation of OS cells after silencing SPINT1-AS1. D Colony formation assay of OS cells after silencing SPINT1-AS1. E, F Wound healing assay of OS cells after silencing SPINT1-AS1 (scale bar, 100 μm). G Transwell assay of OS cells after silencing SPINT1-AS1 (scale bar, 50 μm). H–J Representative images of xenograft tumors (n = 3) resulting from SPINT1-AS1 silencing and control cells. Tumor growth curves and tumor weight are shown. Scale bars = 1 cm. K Representative images of Ki67 and TUNEL staining in different groups. A TUNEL positive cell is indicated (arrow). Scale bars = 50 μm. L Expression of SPINT1-AS1 was showed in xenograft tumors from sh-SPINT1-AS1 group and sh-NC group. *p < 0.05; **p < 0.01; ***p < 0.001 (Student's t-test)

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SPINT1-AS1 contributed to the Warburg effect in OS

It has been widely reported that cancer cells favor aerobic glycolysis over oxidative phosphorylation, a phenomenon known as the Warburg effect [24]. Some evidences have confirmed that the Warburg effect play a critical part in tumor progression [25]. We then explored whether SPINT1-AS1 contributes to glycometabolism in OS. The results indicated that SPINT1-AS1 knockdown reduced glucose consumption, lactate, and pyruvate levels in OS cells (Fig. 3A–C). We also measured the extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) in OS cells and found that ECAR was lower in the SPINT1-AS1 knockdown group, while OCR was higher compared to the control group (Fig. 3D–G). These findings demonstrate that SPINT1-AS1 regulates glucose metabolism and contributes to the Warburg effect in OS cells.

Fig. 3
figure 3

SPINT1-AS1 contributed to the Warburg effect in OS. A SPINT1-AS1 depletion inhibits glucose consumption. B SPINT1-AS1 depletion inhibits extracellular lactate production. C SPINT1-AS1 depletion inhibits pyruvate production. D, E ECAR of Vector and SPINT1-AS1-KD in OS cells. Glc: glucose, Oligo: oligomycin, 2-DG: 2-deoxy-d-glucose. F, G OCR of Vector and SPINT1-AS1-KD in OS cells. O: Oligomycin, F: FCCP, A&R: antimycin A/rotenone. Glucose consumption. *p < 0.05; **p < 0.01; ***p < 0.001 (Student's t-test)

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SPINT1-AS1 functions as a sponge for miR-135b-5p

It has been widely documented that certain long non-coding RNAs (lncRNAs) function as sponges for microRNAs, thereby regulating gene expression [26]. We utilized the StarBase website to identify multiple microRNAs that interact with SPINT1-AS1. Based on the 'clip ExpNum' levels, we selected the top ten microRNAs as potential targets (Table S1). Among these candidates, miR-135b-5p was notably downregulated in OS tissues (Figure S1), leading us to select it as a potential target of SPINT1-AS1. As shown in Fig. 4A, the potential target site of SPINT1-AS1 on miR-135b-5p was predicted via the StarBase website. The silencing of SPINT1-AS1 significantly increased miR-135b-5p expression in OS cells (Fig. 4B). Next, the RNA immunoprecipitation assay confirmed that endogenous SPINT1-AS1 enrichment was enhanced by overexpressing miR-135b-5p in OS cells (Fig. 4C, D). In addition, we mutated the predicted binding site of miR-135b-5p in SPINT1-AS1. The dual-luciferase reporter assays demonstrated that miR-135b-5p mimics reduced the luciferase activity of SPINT1-AS1-WT but had no effect on SPINT1-AS1-MUT in OS cells (Fig. 4E, F). These data collectively demonstrate that SPINT1-AS1 acts as a sponge for miR-135b-5p in OS cells.

Fig. 4
figure 4

SPINT1-AS1 functions as a sponge of miR-135b-5p. A Putative binding sites between SPINT1-AS1 and miR-135b-5p. B The miR-135b-5p expression increased in OS cells transfected with SPINT1-AS1 shRNA. C, D AGO2-RIP followed by qPCR to detect SPINT1-AS1 level after miR-135b-5p overexpression. E, F Luciferase activity of OS cells in luciferase reporter plasmid containing wild-type SPINT1-AS1 3ʹ-UTR and mutant SPINT1-AS1 3ʹ-UTR co-transfected with miR-135b-5p mimics or negative control was detected. *p < 0.05; **p < 0.01; ***p < 0.001 (Student's t-test)

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Upregulation of miR-135b-5p inhibited the cancer progression and aerobic glycolysis in OS cells

We investigated the relationship between miR-135b-5p and OS growth by transfecting miR-135b-5p mimics into OS cells. The efficiency of transfection was assessed using qRT-PCR (Fig. 5A). The CCK8 assay (Fig. 5B, C) and colony formation assay (Fig. 5D) demonstrated that the overexpression of miR-135b-5p partially suppressed OS cell growth. Furthermore, increased levels of miR-135b-5p significantly inhibited the migration of OS cells (Fig. 5E–G). Additionally, the data indicated that elevated expression of miR-135b-5p led to reduced glucose consumption and decreased lactate and pyruvate levels in OS cells (Fig. 5H–J). These findings collectively reveal that the upregulation of miR-135b-5p inhibits tumor progression and aerobic glycolysis in OS cells.

Fig. 5
figure 5

Upregulation of miR-135b-5p inhibited the tumor progression and aerobic glycolysis in OS cells. A Expression of miR-135b-5p was measured in miR-135b-5p-transfected or control OS cells. B, C The proliferation of OS cells after overexpressing miR-135b-5p. D Colony formation assay of OS cells after overexpressing miR-135b-5p. E, F Wound healing assay of OS cells after overexpressing miR-135b-5p (scale bar, 100 μm). G Transwell assay of OS cells after overexpressing miR-135b-5p (scale bar, 50 μm). H–J Glucose consumption, lactate production and pyruvate production of OS cells transfected with miR-135b-5p or control was presented. *p < 0.05; **p < 0.01; ***p < 0.001 (Student's t-test)

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PGAM1 was a target of miR-135b-5p

We predicted multiple mRNAs interacting with miR-135b-5p using the TargetScan website, selecting 12 genes related to the glycolysis pathway due to the regulatory role of miR-135b-5p in aerobic glycolysis within OS cells (Table S2). Among these, PGAM1 was identified as a gene closely associated with osteosarcoma progression. The potential target site of miR-135b-5p on PGAM1 is illustrated in Fig. 6A. We mutated the predicted binding site of miR-135b-5p in PGAM1. The dual-luciferase reporter assay demonstrated that miR-135b-5p mimics reduced the luciferase activity of PGAM1-WT but not PGAM1-MUT in OS cells (Fig. 6B, C). Furthermore, the RIP assay revealed a higher enrichment level of PGAM1 in the AGO2 group following the increase of miR-135b-5p (Fig. 6D, E). Besides, Western blotting and qRT-PCR confirmed that the transfection of miR-135b-5p mimics led to a reduction in PGAM1 expression (Fig. 6F, G). Together, these data establish that miR-135b-5p directly targets PGAM1 in OS cells.

Fig. 6
figure 6

PGAM1 was a Target of miR-135b-5p. A Putative binding sites between PGAM1 and miR-135b-5p. B, C Dual-luciferase reporter assay in miR-135b-5p-transfected or control OS cells. D, E RIP assays using antibodies against AGO2 or IgG were measured in cellular lysates from OS cells. F, G PGAM1 expression was detected in miR-135b-5p-transfected or control OS cells by western blotting and qRT-PCR. *p < 0.05; **p < 0.01; ***p < 0.001 (Student's t-test)

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SPINT1-AS1 promoted OS progression and enhanced aerobic glycolysis via modulating the miR-135b-5p/PGAM1 axis

As depicted in Fig. 7A, PGAM1 expression was elevated in OS cell lines compared to the human osteoblast cell line. Next, we conducted a rescue experiment to further investigate the role of the miR-135b-5p/PGAM1 axis in the SPINT1-AS1-mediated cellular processes of OS. We transfected either miR-135b-5p inhibitors or pcDNA-PGAM1 into SPINT1-AS1 knockdown OS cells. The results demonstrated that the growth suppression of OS cells caused by sh-SPINT1-AS1 was reversed with the overexpression of PGAM1 or knockdown of miR-135b-5p (Fig. 7B–D). In addition, PGAM1 upregulation or miR-135b-5p downregulation mitigated the inhibition of OS cell migration induced by SPINT1-AS1 knockdown (Fig. 7E). Similarly, we observed that inhibiting miR-135b-5p and upregulating PGAM1 partially restored the Warburg effect in OS cells (Fig. 7F–H). These findings indicate that SPINT1-AS1 facilitates OS progression and enhances aerobic glycolysis by modulating the miR-135b-5p/PGAM1 axis.

Fig. 7
figure 7

SPINT1-AS1 promoted OS progression and enhanced aerobic glycolysis via modulating the miR-135b-5p/PGAM1 axis. A PGAM1 expression in OS cell lines and in hFOB1.19. B, C CCK8 assay measured the proliferation in various groups (sh-Control, sh-SPINT1-AS1, sh-SPINT1-AS1 + anti-miR-135b-5p and sh-SPINT1-AS1 + PGAM1). D Colony formation by OS cells in various groups (sh-Control, sh-SPINT1-AS1, sh-SPINT1-AS1 + anti-miR-135b-5p and sh-SPINT1-AS1 + PGAM1). E Wound healing assay of OS cells within various groups (sh-Control, sh-SPINT1-AS1, sh-SPINT1-AS1 + anti-miR-135b-5p and sh-SPINT1-AS1 + PGAM1). F–H Glucose consumption, lactate production, and pyruvate production were measured within various groups (sh-Control, sh-SPINT1-AS1, sh-SPINT1-AS1 + anti-miR-135b-5p and sh-SPINT1-AS1 + PGAM1). *p < 0.05; **p < 0.01; ***p < 0.001 (Student's t-test)

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Discussion

Of note, lncRNA SPINT1-AS1 has been implicated in tumor growth and metastasis across various cancer types. In cervical cancer, SPINT1-AS1 enhances cell proliferation and migration by inhibiting the biogenesis of miR-214 and activating the Wnt/β-catenin signaling pathway [10]. Zhou et al. reported that SPINT1-AS1 regulates miR-let-7a/b/i-5p, facilitating breast cancer progression [27]. In colorectal cancer, SPINT1-AS1 has been confirmed to be upregulated in CRC tissues, playing a crucial role in disease advancement [12]. Furthermore, Sui et al. demonstrated that the SPINT1-AS1/miR-433-3p/E2F3 axis establishes a positive feedback loop that promotes cell proliferation, migration, and invasion, specifically in KRAS-mutant colorectal cancer [11]. However, the role of SPINT1-AS1 in osteosarcoma remains poorly understood. Our research findings indicate that SPINT1-AS1 expression is elevated in OS tissues and cell lines, and that it promotes the growth and migration of OS cells in vitro.

A substantial number of investigations have confirmed that lncRNAs serve as crucial mediators in intracellular signaling pathways, playing significant roles in regulating mRNA transport and microRNA expression [28]. Our findings, utilizing an online prediction tool for SPINT1-AS1 target genes, suggest that SPINT1-AS1 may contribute to cancer promotion by interacting with miR-135b-5p. Members of the miR-135b family have been associated with the progression of various tumors, including colorectal cancer, renal cell carcinoma, pancreatic cancer, triple-negative breast cancer, and ovarian cancer. For instance, aberrant levels of miR-135b-5p have been reported, with its knockdown leading to increased FOXN3 expression and promoting resistance to cetuximab in colorectal cancer [29]. Wang et al. reported that lncRNA GAPLINC influences tumorigenesis through the miR-135b-5p/CSF1 axis in renal cell carcinoma [30]. Moreover, dysregulation of miR-135b-5p has been noted in TNBC tissues, suggesting its involvement in TNBC tumorigenesis [31]. Nevertheless, the function of miR-135b-5p in osteosarcoma remains unclear. Our study indicated that miR-135b-5p was significantly downregulated in OS cell lines and plays a critical role in inhibiting the proliferation and metastasis of OS cells.

One of the characteristics of cancer is uncontrollable proliferation and increased aerobic glycolysis, which meets the energy and biosynthesis needs of tumors and adapts cells to hypoxic conditions [32, 33]. Furthermore, cancer cells exhibit a significant reliance on glucose, leading to lactate production that facilitates tumor proliferation and creates a heterogeneous tumor microenvironment (TME). This environment is characterized by hypoxia, nutrient deprivation, and extracellular acidosis [34,35,36]. The abnormal TME can inhibit immune responses to tumors and activate signaling pathways in OS in unconventional ways [37, 38]. Consequently, targeting the disruption of aerobic glycolysis has emerged as a promising therapeutic strategy. In the present study, we elucidated that SPINT1-AS1 functions as a novel promoter of aerobic glycolysis in OS by sponging miR-135b-5p, which subsequently increases the expression of PGAM1, a pivotal enzyme involved in glycolytic processes. These findings offered the potential of using SPINT1-AS1 or miR-135b-5p as diagnostic or prognostic markers in osteosarcoma.

Conclusion

In summary, the outcomes demonstrate that SPINT1-AS1 functions as an oncogene, with its expression levels significantly elevated in OS. Specifically, our data reveal that SPINT1-AS1 is critical in regulating aerobic glycolysis by sponging miR-135b-5p to increase PGAM1 expression in OS cells. Based on these collective findings, SPINT1-AS1 may serve as a promising therapeutic target for treating OS. However, specific therapeutic strategies need in-depth study.

Availability of data and materials

No datasets were generated or analysed during the current study.

Abbreviations

OS:

Osteosarcoma

lncRNAs:

Long noncoding RNAs

ceRNA:

Competing endogenous RNA

PGAM1:

Phosphoglycerate mutase 1

OCR:

Oxygen consumption rate

ECAR:

Extracellular acidification rate

TNBC:

Triple-negative breast cancer

TME:

Tumor microenvironment

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Acknowledgements

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Funding

The project of Zhenjiang First People's Hospital (YQ2023009) provided funding for this study.

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Author notes

  1. Jinyou Wang, Ning Zhang, Yan Wang, and Fanghong Chen are contributed equally to this work and shared the first authorship.

Authors and Affiliations

  1. Department of Orthopedics, The Affiliated People’s Hospital of Jiangsu University, Zhenjiang, 212002, Jiangsu, China

    Jinyou Wang, Yan Wang, Lei Wang, Xiao Yao & Xiaohui Pan

  2. Department of Orthopedics, Yixing People’s Hospital, Yixing, Jiangsu, China

    Ning Zhang

  3. The Affiliated Lianyungang Municipal Oriental Hospital of Xuzhou Medical University, Lianyungang, 222042, China

    Fanghong Chen

  4. Department of Orthopedics, Zhenjiang First People’s Hospital Branch, Zhenjiang, People’s Republic of China

    Kailun Wang

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  1. Jinyou Wang
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Contributions

J.Y.W. was drafted original manuscript. N.Z., Y.W. and F.H.C. helped us revised the grammar and sentence structure of this manuscript. L.W., X.Y. and K.L.W. handled the statistical analysis. X.H.P. is responsible for reviewing and editing.

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Correspondence to Kailun Wang, Xiao Yao or Xiaohui Pan.

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Wang, J., Zhang, N., Wang, Y. et al. LncRNA SPINT1-AS1 enhances the Warburg effect and promotes the progression of osteosarcoma via the miR-135b-5p/PGAM1 axis. Cancer Cell Int 25, 124 (2025). https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12935-025-03761-7

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Cancer Cell International

ISSN: 1475-2867

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