Do germline genetic variants affect the efficacy of epidermal growth factor receptor tyrosine kinase inhibitors?

Guan et al. identified the germline single nucleotide polymorphism (SNP), ubiquitin-specific peptidase 36 (USP36) rs3744797 (C > A), as a predictor of resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) in advanced non-small cell lung cancer (NSCLC) patients harboring EGFR activating mutations. Because variants with an allele frequency of ≥0.01 were selected for the analysis, the term “single nucleotide polymorphism” used in this study is appropriate. However, as the term “polymorphism” implies functional neutrality, “single nucleotide variant” has been more commonly used in recent years. This variant increases the expression of USP36, stabilizing mixed-lineage leukemia translocated to chromosome 3 (MLLT3), which in turn promotes resistance and tumor aggressiveness. This study suggests that USP36 rs3744797 could serve as a biomarker for predicting the EGFR-TKI response in NSCLC (1).

This study introduced a novel approach to the methodology of genetic association studies. Traditionally, when discussing the relationship between genetic variants and the risk of disease onset or progression, evaluation has predominantly focused on two aspects: (I) differences in gene expression levels, which are assessed by the presence or absence of nonsense mutations or frameshift mutations within the cDNA region, the characteristics of variants in the promoter region, the presence of structural variants, or variations in copy number; and (II) whether the function of the gene is maintained, diminished (or in rare cases, enhanced), which is evaluated using in silico assays, such as the Combined Annotation Dependent Depletion (CADD) score, or by introducing mutant alleles into cells. In contrast, this study takes a different approach by focusing not on the protein information derived from the sequence but on mRNA stability, identifying variants that are likely to have a functional impact. This approach provides a new perspective for genetic association studies.

The rs374977-A variant, which leads to a lysine to asparagine change (p. Lys814Asn), is relatively rare across different populations and is classified as likely benign according to the American College of Medical Genetics and Genomics (ACMG) criteria (https://varsome.com/). It is located at the C-terminus of the USP36 gene, which corresponds to an m6A modification site. Notably, while this variant alone does not show any significant impact on vital functions, it reduces the addition of m6A modifications, resulting in increased epigenetic stability of USP36 mRNA, subsequently leading to the elevated expression of USP36.

Furthermore, the variant of the deubiquitinating enzyme USP36 stabilizes MLLT3 by reducing its ubiquitination levels of MLLT3 in the nucleoli. MLLT3, which is involved in chromatin remodeling, transcriptional regulation, and histone modification, has become more stable. This suggests a potential role of the variant in resistance to EGFR-TKIs. However, the implications of this factor may not be apparent unless multiple layers of this pathway are examined in detail.

No previous reports have suggested an association between SNPs involved in epigenetics and EGFR-TKIs (2). Notably, this study provides evidence for this observation using both in vitro and in vivo clinical data. For points that require clarification, the primary issue appears to be the inability to clearly elucidate the mechanism connecting the overexpression of MLLT3 with the efficacy of EGFR-TKIs.

Arguably, the most significant issue with this study is that it presents findings that do not align with the established clinical observations in lung cancer. As discussed in the manuscript, this variant is more prevalent in Asian populations, and data from the Genome Aggregation Database (gnomAD) confirmed its relatively high frequency in East Asians (Table 1). Consequently, approximately 1 in 10 individuals in this population carries the deleterious allele, suggesting that the efficacy of EGFR-TKIs might be reduced in NSCLC patients with EGFR mutations within this demographic. However, this expectation is not supported by clinical reality. In contrast to the present study, numerous previous studies have reported that Asian lung adenocarcinoma patients, even those with the same EGFR mutations, often respond better to EGFR-TKIs than patients from other ethnic groups (3-5). This discrepancy highlights a significant inconsistency with the findings of the current study and warrants further investigation.

MLLT3 has been reported as a partner gene for fusion genes in hematological malignancies (6). Additionally, it was recently identified as a regulator of self-renewal in hematopoietic stem cells (7). MLLT3 also acts as a master regulator of human erythroid and megakaryocyte fate decisions via GATA1 regulation (8). Given the role of MLLT3 as a transcriptional regulator that interacts with other transcription factors, the elucidation of its relationship with the action of EGFR-TKIs remains complex at this stage.

A study has reported correlations between germline polymorphisms and EGFR-TKI efficacy and the prognosis of EGFR-TKIs. Polymorphisms within the EGFR gene, AKT gene, and SMAD3 genes associated with the TGF-β pathway, as well as polymorphisms in cellular efflux transporters such as ABCB1 and ABCG2, and in cytochrome P450 family genes, including CYP3A4/5, CYP2D6, and CYP1A1, have been shown to correlate with the efficacy and prognosis of EGFR-TKI therapy (9).

The authors previously proposed an algorithm for germline single-nucleotide variants (SNVs) known as the Genomic Prostate Score Estimator (GPSE) risk score, which includes mutations in XRCC1, FOXM1, FOXO3, and ERCC1, as a prognostic tool for stratifying patients with EGFR mutation-positive NSCLC (2). Combining the effects of multiple risk SNPs, rather than relying on a single SNP, is important for building a prognostic prediction model, such as UGT1A1 SNPs that predict the side effects of irinotecan (10). There may be further potential for improvement by incorporating germline SNVs that are thought to be involved in resistance to other EGFR-TKIs. As demonstrated in this study, while it is well known that polymorphisms in BIM lead to resistance to EGFR-TKIs (11), it is also conceivable that information on polymorphisms in USP36 could be incorporated into this predictive model. The introduction of various risk SNPs raises the issue of how to interpret the presence of multiple risk factors and address the problem of multicollinearity in the output for the prognostic prediction of EGFR-TKI treatment.

The focus of this study was germline SNPs rather than somatic mutations. Furthermore, these SNPs were intended to examine changes in the body’s response to an artificial phenomenon that does not occur during the natural course of developing EGFR lung cancer and administering EGFR-TKIs. In other words, these changes in genes, which are not associated with germline alterations that maintain life or lead to cancer, are prone to being overlooked.

This study suggests that many germline SNPs, which have often been neglected, may be associated with the efficacy of various cancer treatments and the prognosis of cancer patients. This indicates that many SNPs should be investigated as potential research targets in future studies. By shedding light on these overlooked genetic variations, we can enhance our understanding of cancer treatment outcomes and improve therapeutic strategies.

Acknowledgments

Funding: None.

Footnote

Provenance and Peer Review: This article was commissioned by the editorial office, Translational Cancer Research. The article has undergone external peer review.

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Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tcr.amegroups.com/article/view/10.21037/tcr-24-1628/coif). The authors have no conflicts of interest to declare.

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