Glycometabolic Reprogramming-Induced XRCC1 Lactylation Confers Therapeutic Resistance in ALDH1A3-Overexpressing Glioblastoma

Biochemistry Life Sciences Neurosurgery Oncology Medicine Cell Biology
Glioblastoma Metabolic reprogramming Lactylation Therapeutic resistance ALDH1A3 DNA repair

Background

Glioblastoma (GBM) is the most common and aggressive malignant primary brain tumor in adults, characterized by high recurrence and mortality rates. Despite active chemotherapy and radiotherapy post-surgery, GBM remains insensitive to existing treatment regimens, and patients usually have a poor prognosis. Recent studies indicate that the presence of glioblastoma stem cells (GSCs) significantly increases the tumor’s resistance to treatment and chances of recurrence. The lack of specific markers for these cells limits the development of targeted therapies. Therefore, investigating the mechanisms of therapeutic resistance induced by GBM metabolic reprogramming and developing related targeted therapies is expected to improve tumor sensitivity to chemotherapy and radiotherapy, thereby extending patient survival.

Research Origin

This study was led by Guanzhang Li and colleagues with affiliations including Beijing Tiantan Hospital, Beijing Neurosurgical Institute of Capital Medical University, and Otto von Guericke University Magdeburg, among others. The paper was published in the journal “Cell Metabolism” on August 6, 2024.

Detailed Research Process

1. Research Aim and Design

The study first divided GBM patients into high and low ALDH1A3 expression groups through immunofluorescence staining and found that patients in the high expression group benefitted less from postoperative chemoradiotherapy. These patients did not show significant differences in other clinicopathological prognostic factors. Further, ALDH1A3 knockout and restoration cell lines were constructed in patient-derived GSCs through CRISPR-Cas9 gene editing, showing that knockout of ALDH1A3 increased sensitivity to TMZ and radiotherapy, while the restored group regained resistance.

2. RNA Sequencing and Metabolomics Analysis

To explore the mechanisms of treatment resistance driven by ALDH1A3 overexpression, researchers conducted RNA sequencing and untargeted metabolomics analysis of ALDH1A3-edited GSCs. Results showed significant changes in metabolism and metabolites in these GSCs. The study found that ALDH1A3 expression or re-expression led to an increase in L-lactate and glycolytic metabolism, while changes in glycolytic pathway-related gene expression were not obvious.

3. ALDH1A3 and PKM2 Interaction

Using immunoprecipitation and mass spectrometry analysis, researchers discovered a protein interaction between ALDH1A3 and PKM2. Further validation showed that ALDH1A3 physically binds with PKM2 in GSCs. The GST pull-down assay further confirmed the direct protein-protein interaction between ALDH1A3 and PKM2.

4. Lactate-Induced XRCC1 Lactylation

Proteomics analysis revealed that ALDH1A3 knockout significantly altered the overall level of lactate modification of proteins in GSCs. Mass spectrometry identified four lysine sites of XRCC1 that were lactylated. Further analysis showed lactylation at the K247 site of XRCC1 altered its surface charge, thereby enhancing its affinity with nuclear import protein α, significantly increasing DNA repair activity, and contributing to enhanced resistance to radiotherapy and chemotherapy.

5. Screening and Action of Small Molecule Compound D34-919

Based on in vitro and in vivo compound screening, researchers found D34-919 could effectively block the protein interaction between ALDH1A3 and PKM2, significantly inhibiting PKM2 tetramerization in target cells, and effectively preventing the chemotherapy and radiotherapy resistance induced by ALDH1A3 overexpression.

6. In Vivo Animal Experiment Validation

In in vivo animal experiments with mice, three treatment combinations (D34-919 with TMZ, D34-919 with radiotherapy, and a combination of all three) exhibited significant tumor suppression effects and significantly extended overall survival time in mice. Tumors were almost completely eliminated in combination treatment groups without resulting in weight loss or major organ damage.

Research Results

Overall, the study revealed ALDH1A3 promotes lactate accumulation in GSCs by interacting with PKM2 to enhance its tetramerization. This further elevated DNA repair capabilities through XRCC1 lactylation modification, ultimately resulting in resistance to chemotherapy and radiotherapy. By screening the small molecule compound D34-919, the study successfully blocked this interaction, significantly enhancing the sensitivity of GBM cells with high ALDH1A3 expression to chemotherapy and radiotherapy.

Research Significance and Application Value

This study is the first to reveal the specific mechanism of ALDH1A3 through PKM2 tetramerization and XRCC1 lactylation, elucidating the direct link between GBM metabolic reprogramming and therapeutic resistance. The small molecule compound D34-919, as a new chemosensitizer and radiosensitizer, provides a new treatment option for GBM patients with high ALDH1A3 expression.

Research Highlights and Innovations

  1. Discovery of a New Metabolic Mechanism: For the first time, it was revealed that ALDH1A3 induces lactate accumulation in GSCs through interaction with PKM2 and enhances DNA repair ability through XRCC1 lactylation modification.
  2. Small Molecule Compound D34-919: The small molecule compound D34-919, obtained through screening, effectively interferes with the interaction between ALDH1A3 and PKM2, significantly increasing the sensitivity of GBM cells with high ALDH1A3 expression to chemotherapy and radiotherapy.
  3. Clinical Significance: This study provides a new treatment idea and potential drug for GBM patients with high ALDH1A3 expression.

Conclusion

This study deeply reveals the molecular mechanism of GBM metabolic reprogramming and resistance to chemotherapy and radiotherapy, offering new treatment strategies and laying a solid foundation for future clinical research.