Discussion
In this study, we evaluated the prevalence of HLA-I LOH in Chinese cancer patients using the 1021-gene panel and characterized the genomic features of patients harboring HLA-I LOH.
Analytical results indicated that HLA-I LOH was widespread in Chinese cancer patients while varied greatly across cancer types, with an average incidence of 45.6%. When compared with a western population cohort containing 83644 samples, Chinese cancer patients shown a higher incidence of HLA-I LOH (45.6% vs. 17.0%) (Montesion et al., 2020). Since HLA-I LOH is a common immune evasion mechanism later in tumor evolution, we speculate that this difference may be due to the different tumor progression stages of the patients in the study cohort (McGranahan et al., 2017). The percentage of LOH co-occurrence at all three HLA-I loci in HLA-I LOH patients is 43.3% (293/676) in Chinese cohort and 85% in the western population cohort, respectively. (Montesion et al., 2020). Although there is a difference in the frequency of HLA-I LOH, the proportion of patients with LOH at all three sites concurrently is roughly equivalent (19.7% in our cohort vs. 14.5% in western cohort) (Montesion et al., 2020). As confirmed by previous studies, HLA-I LOH is enriched in squamous cell carcinoma (Montesion et al., 2020). In our cohort, the highest incidence of HLA-I LOH occurs in cervical squamous cell carcinoma and lung squamous cell carcinoma. Consistent with prior researches, the occurrence of HLA-I LOH was accompanied by elevated tumor mutation burden (TMB), suggesting the genomic instability in patients harboring HLA-I LOH (Anagnostou et al., 2020; McGranahan et al., 2017). When focusing on patients harboring HLA-I LOH, the TMB level of patients with LOH at a single site was lower than with LOH at all three sites. As one of the important indicators of immunotherapy, high TMB level indicated better immunotherapy prognosis (Hellmann et al., 2018; Samstein et al., 2019; Chan et al., 2019; Rizvi et al., 2018). Considering that the occurrence of multi-loci HLA-I LOH may impair antigen binding and presentation, the survival benefits of high TMB levels could be diminished, in which circumstances patients might have a poor response to immunotherapy. Thus, in clinical practice, the TMB level should be evaluated along with the occurrence of HLA-I LOH and number of LOH loci when predicting the response to immunotherapy.
However, when dividing TMB more finely, the relationship between HLA-I LOH and TMB is more in line with the “Goldilocks” pattern that tumors with low or high TMB levels show a low incidence of HLA-I LOH, while higher incidence is presented in between (Supplementary Figure 3A) (Montesion et al., 2020). Further analysis find that MSI-H patients have significantly higher TMB levels but lower incidence of HLA-I LOH compared with MSS patients, which led to the “Goldilocks” phenomenon in our cohort (Supplementary Figure 3C). Montesion et al. (2020) hypothesized “HLA-I LOH is selected for in tumors with enough neoantigens to elicit an immune response but few enough that HLA-I LOH abrogates immune recognition without eliciting an NK cell response”. We believe that the reasons behind this phenomenon need to be further explored by analyzing the genomic differences of those patients. In addition, the higher incidence of HLA-I LOH in MSS patients suggests that HLA-I LOH may be one of the reasons for the poor response to immunotherapy in some MSS patients, prompting the underlying significance of HLA-I LOH testing in predicting potential benefit population in MSS patients. Since MSI-H group mainly consist of patients with gastrointestinal tumors (59.5% [22/37]) in our cohort, conclusions above might be specifically reflected in these cancer types.
Another important discovery is that patients harboring HLA-I LOH have unique genomic features, such as increased alteration frequencies of several signaling pathways and oncogenes. Since the occurrence of HLA-I LOH is closely related to genomic instability, we first analyzed the differences in the alterations in DDR pathways (McGranahan et al., 2017). Results revealed that alteration frequencies of CPF pathway and FA pathway in the HLA-I LOH group was significantly higher than that of the HLA-I stable group. In addition, the occurrence of HLA-I LOH was accompanied by alterations in several important oncogenic signaling pathways, including p53 pathway, RTK/RAS pathway, Hippo pathway, Notch pathway and Nrf2 pathway, indicating that HLA-I LOH might involve in tumor progression by influencing the normal signaling transduction. As a signaling cascade pathway initiated by activation of RTKs, RTK/RAS pathway regulates cell proliferation and survival and is frequently altered in many cancer types (Malumbres & Barbacid, 2003; Sanchez-Vega et al., 2018). For several oncogenic genes, such as TP53 andLRP1B , the occurrence of HLA-I LOH is related to higher mutation frequencies. As the central gene of p53 pathway and CPF pathway,TP53 regulates apoptosis, cell cycle arrest, senescence, and DNA repair (Wade et al., 2013; Sanchez-Vega et al., 2018). The somatic mutations of this important tumor suppressor may cause genomic instability and replication stress, which is closely related to tumorigenesis in various cancer types (Nigro et al., 1989; Baker et al., 1989; Stracquadanio et al., 2016; Malekzadeh et al., 2019). LRP1Bmay play an essential role in tumorigenesis of NSCLCs, and mutations of this gene are associated with a higher TMB level (Lan et al., 2019; Wang et al., 2021). These factors may be involved in positive selection during tumor evolution and promote the occurrence of HLA-I LOH. Therefore, the mechanism underlying these observations and the causal relationship between HLA-I LOH and mutations of certain genes need further investigation.
One of the major limitations of this study is the insufficient collection of clinical information, such as smoking history and clinical outcome, which precluded us to set a cohort to analyze the clinical significance of our findings. Another shortcoming is that the number of samples used for analysis is relatively small compared with other large population studies. In our follow-up study, after enrolling a sufficient number of patients and collecting adequate clinical data, these limitations will be overcome. Therefore, deeper mechanism mining and clinical verification will be achieved.
In general, we confirmed the validity of 1021-gene panel for HLA-I LOH detection through comparison with whole-exome sequencing (WES). The 1021-gene panel could be applied for HLA-I LOH analysis, provided that the relevant regions are well captured. We further depicted the landscape of HLA-I LOH events in Chinese cancer patients and revealed the genomic features of patients harboring HLA-I LOH. These insights may provide valuable information for clinical practice and follow-up immunotherapy research.