DOI: https://doi.org/10.21203/rs.3.rs-1752120/v1
Purpose: A substantial need for effective and safe treatment options is still unmet for patients with heavily pre-treated human epidermal growth factor receptor 2 (HER2)-positive metastatic breast cancer (MBC). Herein, we assessed the efficacy and safety of pyrotinib plus trastuzumab and chemotherapy in patients with heavily treated HER2-positive MBC.
Methods: In this single-arm exploratory phase II trial, patients with HER2-positive MBC previously treated with trastuzumab plus lapatinib or pertuzumab, received pyrotinib plus trastuzumab and chemotherapy. The primary end point was progression-free survival (PFS) in the total population (TP). Secondary end points included PFS in the subgroup with brain metastases (Sub-BrM), confirmed objective response rate (ORR), clinical benefit rate (CBR), disease control rate (DCR), exploration of predictive factors of PFS, and safety.
Results: Between November 1, 2018, and March 31, 2021, 40 patients were eligible for this study. The median PFS reached 7.5 months (95% confidence interval [CI], 4.7 to 9.9 months) and 9.4 months (95% CI, 6.6 to 12.1 months) in the TP and Sub-BrM, respectively. ORR was 50.5% (20/40). CBR was 75.5% (30/40) and DCR reached 97.5% (39/40). Cox univariate and multivariate analyses demonstrated that liver or/and lung metastases was the significant adverse prognostic factor for PFS (p = 0.018; p = 0.026; respectively). The most frequent grade 3 or 4 treatment-related adverse events were diarrhea, neutropenia and leukopenia. No new safety signals were observed.
Conclusion: Pyrotinib plus trastuzumab and chemotherapy offered a promising option with manageable safety profile for heavily pre-treated HER2-positive MBC, especially for those without liver or/and lung metastases.
Breast cancer (BC) has surpassed lung cancer as the most commonly diagnosed malignancy in women according to Global Cancer statistics 2020 [1]. Approximately 15 to 20% of BC cases exhibit overexpression of the human epidermal growth factor receptor 2 (HER2) or/and amplification of the coding gene ERBB2, which is defined as HER2-positive subtype [2]. HER2-positive BC is more aggressive and prone to recurrence than HER2-negative tumors [3]. In the last decades, the clinical outcome of HER2-positive BC has been significantly improved since the introduction of HER2-targeted drugs mainly including monoclonal antibodies, tyrosine kinase inhibitors (TKIs) and antibody drug conjugates (ADCs). Currently, the standard of care for HER2-positive metastatic breast cancer (MBC) involves trastuzumab plus pertuzumab and a taxane as the first-line regimen, followed by second-line trastuzumab emtansine (T-DM1) or fam-trastuzumab deruxtecan-nxki (ds8201) for patients whose disease has progressed on prior dual HER2 blockade [4]. However, a large proportion of patients with HER2-positive MBC fail to be treated with standard second-line regimens due to economic and drug accessibility factors. Moreover, most patients inevitably experience disease progression on anti-HER2 targeted therapy due to de novo or acquired resistance. Therefore, it is necessary to explore novel approaches to overcome drug resistance. Key strategies include efficient suppression of the HER2 signaling pathway by dual blockade and development of more effective anti-HER2 therapies like antibody–drug conjugates, new anti-HER2 antibodies, bispecific antibodies and novel TKIs [5].
Brain metastases (BrM) is a common complication of advanced malignant disease and occurs in 1/3 of HER2-positive metastatic breast cancer [6]. Central nervous system-directed local therapies, including surgical resection and radiotherapy, are the foundations of BrM management [7]. Besides, TKIs and several chemotherapeutic drugs with blood-brain barrier penetrability have demonstrated efficacy in HER2-positive MBC patients with BrM. Despite sequential local and systemic treatment, resistance inevitably develops and options are limited for the control of BrM.
Studies have confirmed the activity of continued trastuzumab treatment beyond progression (TBP). The subgroup analysis of observational HERMINE study suggested that trastuzumab TBP offered a survival benefit to MBC patients treated with first-line trastuzumab [8]. Furthermore, a phase III study in HER2-positive BC showed that patients receiving continued trastuzumab TBP had a better post-progression survival than those not receiving (18.8m vs 13.3m, p = 0.02) [9]. Meanwhile, the potent anti-tumor efficacy of TKIs including lapatinib, neratinib, tucatinib and pyrotinib has been demonstrated in several pivotal studies [10–14]. Pyrotinib is an orally administered irreversible pan-ErbB TKI which shows anti-tumor activity and acceptable safety profile in HER2-positive advanced and metastatic breast cancer [14]. In a phase II study, pyrotinib plus capecitabine yielded statistically significant better overall response rate (ORR) and progression-free survival (PFS) than lapatinib plus capecitabine in patients with HER2-positive MBC previously treated with taxanes, anthracyclines, and/or trastuzumab [15]. Based on this phase II study, pyrotinib in combination with capecitabine received its first conditional approval in China for the treatment of HER2-positive, advanced or metastatic breast cancer previously exposed to anthracycline or taxane chemotherapy. Afterwards, a multi-center, open-label, randomized, controlled, phase III trial (PHOEBE) confirmed the efficacy and safety of pyrotinib plus capecitabine in patients with disease progression on previous trastuzumab [16]. Another phase III trial PHENIX further verified that pyrotinib plus capecitabine significantly improved PFS and ORR compared with capecitabine monotherapy in trastuzumab-treated patients with HER2-postive advanced or metastatic breast cancer, including those with brain metastases [17].
Dual HER2 blockade by trastuzumab plus TKI, simultaneously targeting the extra- and intra-cellular domains of HER2, showed encouraging antitumor activity in BC, including early breast cancer and MBC with intracranial metastases [18–21]. Our study aimed to explore the activity and safety of pyrotinib combined with trastuzumab and chemotherapy of physician’s choice in heavily pre-treated patients with HER2-positive MBC with or without BrM.
From November 1, 2018, to March 31, 2021, female patients aged ≥ 18 years old with histologically confirmed HER2-positive MBC at National Clinical Research Center for Cancer/Cancer Hospital & Shenzhen Hospital (Shenzhen, Guangdong, China) were incorporated in this study. HER2 status was determined by central review based on immunohistochemistry (IHC) or fluorescence in situ hybridization (FISH) examination using primary or metastatic lesion samples. Patients had been previously treated with trastuzumab, pertuzumab and/or lapatinib in the (neo)adjuvant/metastatic setting. Additional requirements included an Eastern Cooperative Oncology Group (ECOG) performance status score of 0 or 1 (on a 5-point scale by which greater scores reflect higher degree of disability); measurable lesions; and adequate organ function. Patients were excluded if they had previously received treatment with pyrotinib for metastatic disease; had symptomatic brain metastasis which necessitates immediate local intervention; or had leptomeningeal disease. All procedures performed involving human participants were in accordance with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. This study was also approved by the institutional review board and ethics committee. All patients provided written informed consent for use of the medical information for research purpose.
All eligible patients were treated with pyrotinib (240–400 mg orally once daily), trastuzumab (6 mg per kilogram of body weight intravenously per 21 days, with an initial loading dose of 8 mg per kilogram) and single chemotherapeutic agent (nab-paclitaxel, 260 mg per square meter of body-surface area intravenously on day 1 of each 21-day cycle; or capecitabine, 1000 mg per square meter of body-surface area orally twice daily on days 1 to 14 of each 21-day cycle; or gemcitabine, 1000 mg per square meter of body-surface area intravenously on days 1 and 8 of each 21-day cycle; or vinorelbine, 60 mg per square meter of body-surface area orally on days 1 and 8 of each 21-day cycle and dose elevation to 80 mg per square meter of body-surface area was allowed if well tolerated).
All patients were followed up until January 20, 2022. Disease response was evaluated according to imaging reports from serial clinical assessments. Patient/disease response assessments were performed at baseline, every 6 weeks for 24 weeks, and every 9 weeks thereafter, including performance status, history, laboratory examinations, electrocardiogram, contrast-enhanced spiral computed tomography (CT) or positron emission tomography/CT (PET/CT), and contrast-enhanced magnetic resonance imaging (MRI). MRI of the head was obtained for all patients at baseline and head scans were repeated at the frequency described above if BrM had been detected at baseline. MRI of the breast was not required. Disease was evaluated in accordance with Response Evaluation Criteria in Solid Tumors (RECIST) criteria, version 1.1 [22]. Safety was assessed mainly via the incidence of treatment-related adverse events (TRAEs) using the National Cancer Institute Common Terminology Criteria for Adverse Events version 5.0.
The primary end point was PFS (calculated from the date of first treatment with double targeted therapy to the date of documented disease progression or death from any cause or the last follow-up visit) in TP. The secondary end points included PFS in the subgroup with BrM (Sub-BrM) at baseline; objective response rate (ORR, defined as the percentage of patients who had a confirmed complete response or partial response); clinical benefit rate (CBR, defined as the percentage of patients who had a confirmed complete response or partial response or stable disease for at least 24 weeks); disease control rate (DCR, defined as the percentage of patients who had a confirmed complete response or partial response or stable disease for at least 4 weeks); exploration of predictive factors of PFS; and safety.
All data were analyzed using SPSS 23.0 statistical software (SPSS, Inc., Chicago, IL, USA) and GraphPad Prism 5 software (GraphPad Software, Inc., La Jolla, CA, USA). Descriptive analysis was utilized to display clinicopathological features. The Kaplan–Meier method was used to estimate PFS and 95% confidence intervals (CIs) for the total population and the subgroups. Cox univariate and multivariate models were used to determine the predictive value of variables for PFS. In the analysis of progression-free survival, data from patients who did not have any documented event, were lost to follow-up or died from any cause were censored at the last date when the patient was known to be event-free. All reported p values were two-sided, with p༜0.05 being regarded as statistically significant.
Between November 1, 2018, and March 31, 2021, a total of 40 patients were eligible for this study and included in the final analysis. In TP, the median age at diagnosis was 46 years (range, 31–62 years). 33 patients (82.5%) were aged over 35 years old at diagnosis. ECOG performance status was 0 in 21 patients (52.5%) and 1 in 19 patients (47.5%). For HER2 status, HER2 3 + by IHC was recorded in 32 patients (80.0%), and HER2 2 + by IHC and amplification by FISH in 8 patients (20.0%). 21 patients (52.5%) were estrogen receptor (ER) and/or progesterone receptor (PR) positive and 19 patients (47.5%) were ER and PR negative. 39 patients (97.5%) had been treated with trastuzumab in the (neo)adjuvant or metastatic setting; 6 patients had received pertuzumab and trastuzumab as first-line therapeutic regimen; 21 patients (52.5%) received anti-HER2 therapy with TKI (lapatinib) for their metastatic disease; none of them was previously treated with TDM-1. 15 patients (37.5%) had brain metastases and 27 patients (67.5%) had lung or/and liver metastases at baseline. Of those patients with brain metastases, 7 patients had received central nervous system (CNS) local therapies including whole-brain radiotherapy (WBRT), stereotactic radiotherapy (SRT) and surgery plus SRT before study treatment; 3 patients whose intra-cranial disease progressed while remission of extra-cranial lesions lasted, received SRT with unchanged systemic therapy. 13 patients (32.5%) received at least 3 lines of therapy in the metastatic setting. Combined chemotherapeutic agents included vinorelbine (25, 62.5%), capecitabine (6, 15.0%), nab-paclitaxel (5, 12.5%) and gemcitabine (4, 10.0%). Patient clinicopathological characteristics at baseline were shown in Table 1.
| Characteristics | cases (%) |
|---|---|
| ECOG | |
| 0 | 21 (52.5) |
| 1 | 19 (47.5) |
| Age at diagnosis | |
| ≤ 35 years | 7 (17.5) |
| > 35 years | 33 (82.5) |
| Location | |
| Left | 27 (67.5) |
| Right | 13 (32.5) |
| HER2 status | |
| IHC 3+ | 32 (80.0) |
| IHC 2 + and FISH + | 8 (20.0) |
| Hormone-receptor status | |
| ER and/or PR positive | 21 (52.5) |
| ER and PR negative | 19 (47.5) |
| Previous anti-HER2 antibody treatment in metastatic setting | |
| Yes | 37 (92.5) |
| No | 3 (7.5) |
| Previous anti-HER2 TKI treatment | |
| Yes | 21 (52.5) |
| No | 19 (47.5) |
| Brain metastases | |
| Yes | 15 (37.5) |
| No | 25 (62.5) |
| Liver or/and lung metastases | |
| Yes | 27 (67.5) |
| No | 13 (32.5) |
| Number of previous treatment lines | |
| ≤ 2 | 27 (67.5) |
| > 2 | 13 (32.5) |
| Combined chemotherapeutic drug | |
| Vinorelbine | 25 (62.5) |
| Capecitabine | 6 (15.0) |
| Nab-paclitaxel | 5 (12.5) |
| Gemcitabine | 4 (10.0) |
| Abbreviations: | |
| ECOG Eastern Cooperative Oncology Group; HER2 human epidermal growth factor receptor 2; IHC immunohistochemistry; ER estrogen receptor; PR progesterone receptor. | |
By the end of follow-up, 32 patients had experienced disease progression and 3 patients had died. The median duration of PFS was 7.5 months in TP (95% CI, 4.7 to 9.9 months, Fig. 1) and was 9.4 months in Sub-BrM (95% CI, 6.6 to 12.1 months, Fig. 2). Objective response was seen in 20 of 40 patients (including 1 patient with complete response and 19 patients with partial response, 50.0%, Table 2). All patients with objective response achieved disease remission within four cycles (12 weeks) after treatment administered. Clinical benefit was observed in 30 of 40 patients (75.5%) and disease control in 39 of 40 patients (97.5%, Table 2).
| End points | |
|---|---|
| Median progression-free survival, months (95% CI) | 7.5 (4.4 to 7.9) |
| Median progression-free survival in subgroup with brain metastases, months (95% CI) | 9.4 (6.6 to 12.1) |
| Type of response, No. (%) | |
| Complete | 1 (2.5) |
| Partial | 19 (45.0) |
| Stable disease | 19 (50.0) |
| Disease progression | 1 (2.5) |
| Objective response rate, No. (%)* | 20 (50.0) |
| Clinical benefit rate, No. (%)** | 30 (75.5) |
| Disease control rate, No. (%)*** | 39 (97.5) |
| *Include complete and partial response. **Include complete and partial response and stable disease lasting for at least 24 weeks. ***Include complete and partial response and stable disease lasting for at least 4 weeks. Abbreviations: CI confidence interval. | |
Univariate and multivariate analyses were performed to identify predictive factors of PFS for the 40 patients. By means of Cox univariate and multivariate analyses, we found that liver or/and lung metastases at baseline was the significant adverse prognostic factor for PFS (hazard ratio [HR], 0.322; 95% CI, 0.132 to 0.831; p = 0.018; HR, 0.281; 95% CI, 0.093 to 0.876; p = 0.026; respectively; Table 3). The median PFS was 6.0 months (95% CI, 2.8 to 9.2 months) in the subgroup with liver or/and lung metastases and was not reached in the subgroup without (Fig. 3).
| Variables | Univariate analysis | Multivariate analysis | |||||
|---|---|---|---|---|---|---|---|
| HR | 95% CI | P value | HR | 95% CI | P value | ||
| Age at diagnosis, years (≤ 60 vs > 60) | 1.277 | 0.484–3.368 | 0.622 | ||||
| Location (left vs right) | 1.219 | 0.547–2.716 | 0.629 | ||||
| HER2 status (IHC 3 + vs IHC2 + and FISH +) | 1.017 | 0.355–2.687 | 0.972 | ||||
| Hormone-receptor status (ER and/or PR positive vs ER and PR negative) | 0.639 | 0.299–1.367 | 0.248 | ||||
| Previous anti-HER2 antibody treatment in metastatic setting (yes vs no) | 0.947 | 0.225–4.037 | 0.953 | ||||
| Previous anti-HER2 TKI treatment (yes vs no) | 0.958 | 0.451–2.033 | 0.911 | ||||
| Brain metastases (yes vs no) | 1.877 | 0.843–4.179 | 0.123 | ||||
| Liver or/and lung metastases (yes vs no) | 0.322 | 0.132–0.831 | 0.018 | 0.281 | 0.093–0.876 | 0.026 | |
| Number of previous treatment lines (≤ 2 vs > 2) | 1.190 | 0.548–2.585 | 0.660 | ||||
| Combined chemotherapeutic drug (Vinorelbine vs Cepecitabine vs Nab-paclitaxel) | 1.197 | 0.834–1.717 | 0.329 | ||||
| Abbreviations: HR hazard ratio; CI confidence interval; HER2 = human epidermal growth factor receptor; IHC immunohistochemistry; FISH fluorescence in situ hybridization; ER estrogen receptor; PR progesterone receptor; TKI tyrosine kinase inhibitor. | |||||||
All treatment-related adverse events (TRAEs) reported in the study were summarized in Table 4. The most common adverse events (frequency ≥ 10%) were diarrhea (85.0%), leukopenia (42.5%), neutropenia (37.5%), fatigue (37.5%), vomiting (32.5%), and nausea (12.5%) in turn. Other recorded adverse events included hand-foot syndrome (10.0%), aspartate aminotransferase increase (10.0%), anorexia (7.5%), alanine aminotransferase increase (5.0%), anemia (5.0%), peripheral neurotoxicity (5.0%), rash (2.5%) and thrombocytopenia (2.5%). The most frequent grade 3 to 4 adverse events were diarrhea (25.0%), neutropenia (15.0%), and leukopenia (12.5%). Grade 4 adverse events occurred in 1 patient (leukopenia and febrile neutropenia). No grade 4 diarrhea or cardiac-related events were reported. No one discontinued study treatment or died owing to TRAEs. However, 14 patients (35.0%) experienced pyrotinib dose reduction because of diarrhea (10 patients reduced from 400 mg to 320 mg and 4 patients from 400 mg to 240 mg). 4 patients experienced chemotherapeutic drug dose reduction (nab-paclitaxel in 1 patient, capecitabine in 1 patient, and vinorelbine in 2 patients). Diarrhea, the most common adverse event, mostly occurred within the first two weeks after treatment initiation and could be managed by the administration of loperamide or/and pyrotinib dose reduction.
| Adverse events | Any grade (%) | Grade ≥ 3 (%) |
|---|---|---|
| Diarrhea | 34 (85.0) | 10 (25.0) |
| Leukopenia | 17 (42.5) | 5 (12.5) |
| Neutropenia | 15 (37.5) | 6 (15.0) |
| Fatigue | 15 (37.5) | 0 |
| Vomiting | 13 (32.5) | 1 (2.5) |
| Nausea | 5 (12.5) | 0 |
| Hand-foot syndrome | 4 (10.0) | 0 |
| Aspartate aminotransferase increase | 4 (10.0) | 1 (2.5) |
| Anorexia | 3 (7.5) | 0 |
| Alanine aminotransferase increase | 2 (5.0) | 0 |
| Anemia | 2 (5.0) | 0 |
| Peripheral neurotoxicity | 2 (5.0) | 0 |
| Rash | 1 (2.5) | 0 |
| Thrombocytopenia | 1 (2.5) | 1 (2.5) |
In this single-center study, pyrotinib combined with trastuzumab and chemotherapy demonstrated promising efficacy in heavily pre-treated HER2 positive MBC (92.5% of the patients with prior trastuzumab and 52.5% with lapatinib), with a median PFS of 7.5 months and an ORR of 50%, which implies that half of the patients could still respond to pyrotinib-based regimen even in later-line settings.
As is well known, efficacy and safety of pyrotinib in patients with HER2-positive MBC have been verified in several pivotal studies. In the phase I study pyrotinib monotherapy demonstrated an ORR of 83.3% in trastuzumab-naive patients and 33.3% in trastuzumab-pretreated patients [14]. The benefit of continued use of trastuzumab beyond disease progression was also confirmed in earlier studies [8, 9]. The anti-tumor activity and favorable tolerability of dual HER2 blockade with TKI plus trastuzumab has been reported previously and mechanisms of synergistic interaction may involve enhanced apoptosis of cancer cells, increased stabilization and degradation of HER2 receptors, and reversion of resistance to trastuzumab by accumulation of HER2 receptors on the surface of breast cancer cells [20, 21]. Impressively, Murthy and colleagues reported that tucatinib, which was an investigational, oral, highly selective inhibitor of the HER2 tyrosine kinase, was an alternative regimen in heavily pre-treated metastatic breast cancer when combined with trastuzumab and capecitabine, with a median PFS of 7.8 months in the study population and 7.6 months in the brain metastases subgroup [19]. Another single-arm exploratory phase II trial has demonstrated that pyrotinib plus trastuzumab and albumin-bound paclitaxel generated an encouraging pCR rate and an ORR of 100% in the neoadjuvant setting [18]. To our knowledge, there exists no published data investigating the activity and safety of pyrotinib plus trastuzumab and chemotherapy in MBC patients previously treated. Our findings are fairly consistent with and further consolidate the above-mentioned data. The PFS duration reported here is similar with that in the HER2CLIMB trial. Noteworthy, the median PFS of BrM subset that have presumably worse prognosis, was numerically superior to that of the total population. The previous therapeutic lines may predominantly account for this result, since 32.5% of the total population versus 20.0% of BrM subgroup received ≥ 2 lines of therapy previously. Besides, concerning the patients with intracranial metastasis, the PFS duration estimated here (9.4 months) was also longer than that in the HER2CLIMB trial (7.6 months). The potent activity of pyrotinib per se for intracranial metastases may partly account for this finding. Another possible explanation is that local therapy for intracranial metastases (46.7% of the patients with brain metastases received local therapy) improved blood brain barrier permeability and thus the intracranial concentration of anti-tumor drugs increased.
Up to now, evidences on head-to-head comparison of TKI plus trastuzumab versus TKI alone are lacking. A randomized phase II trial, which was designed to assess the efficacy and tolerability of pyrotinib plus capecitabine versus lapatinib plus capecitabine in women with HER2-positive MBC previously treated with taxanes, anthracyclines, and/or trastuzumab, reported that the ORR was 78.5% and the median PFS was 18.1 months in the pyrotinib arm [15]. Similarly, PHENIX study showed that the ORR and median PFS of pyrotinib plus capecitabine were 68.6% and 11.1 months, respectively [17]. Numerically, these findings were superior to the data reported in our study, which might be partly attributed to different distribution of patient characteristics. The phase II study and PHENIX study enrolled patients with no more than 2 lines of prior treatment in the metastatic setting, and nearly 70% of the patients were trastuzumab-naïve. However, 67.5% of the patients in our study received ≥ 2 lines of therapy previously, and almost all patients had received prior trastuzumab for their metastatic disease [15]. Considering that 52.5% of our patients had been exposed to prior lapatinib, our data further implies the activity of pyrotinib beyond progression on lapatinib treatment. It follows that pyrotinib combined with trastuzumab and chemotherapy may offer a promising alternative for patients with heavily treated HER2-positive MBC, including those with BrM.
In addition, we further analyzed the predictive factors of PFS and found that liver or/and lung metastases was independent adverse factor for PFS, which suggested that patients without liver or/and lung metastases might better benefit from this combination therapy. This finding could provide guidance for patient selection and optimize clinical management.
Referring to safety, the overall incidences of TRAEs were similar to that of previous report and no new TRAEs were reported. The majority of TRAEs were Grade 1 or 2 in severity and most of TRAEs could be managed by dose reduction and supportive treatment. No one discontinued study treatment or died due to TRAEs. Diarrhea, usually occurring on days 4–14 after the first dose, is the most common TRAE which is mainly induced by pyrotinib and could be managed by dose reduction and loperamide, turning mostly tolerable after treatment administered for one month. The majority of diarrhea events were Grade 1 or 2, and 25% of them were Grade 3 or 4. Prophylaxis use of loperamide could effectively reduce incidence of diarrhea caused by neratinib. However, evidence is scarce supporting the prophylaxis use of loperamide in pyrotinib treatment. Of note, no cardiac-related events were recorded in our study. The low incidence of severe adverse events demonstrated the safety of pyrotinib plus trastuzumab in heavily pretreated MBC patients.
After all, the study is an initial single-center investigation based on a small Chinese cohort. Another limitation is the relatively low rate of standard care administration in previous lines of treatment among the enrolled patients. Only 6 patients received pertuzumab and trastuzumab as first-line therapeutic regimen and none of them had received TDM-1 treatment owing to financial and drug accessibility factors. Multi-center randomized controlled trials in larger cohorts are needed to further validate the efficacy and safety of this combination regimen.
In conclusion, pyrotinib in combination with trastuzumab and chemotherapy offer an active option with a favorable safety profile in heavily pre-treated patients with HER2-positive MBC, including those with brain metastases. Multi-center randomized controlled trials are warranted to validate the results.
Conflict of Interest
The authors declare that they have no conflict of interest.
Acknowledgements
We are grateful to the patients and all the researchers, including the physicians, pathologists, and technicians, who participated in this study.
Funding
This work was supported by funds from ShenZhen basic research program (2018, JCYJ20180306171227129). Cai-Wen Du has received research support from Shenzhen Science and Technology Innovation Commission.
Competing interests
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Author contributions statement
All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Xiao-Feng Xie, Qiu-Yi Zhang, Jia-Yi Huang, Li-Ping Chen, Si-Jia Guo, Shui-Ling Xiong and Xiao-Feng Lan. Writing-reviewing and editing were performed by Xie-Xiao Feng, Xue-Bai, Lin Song. The first draft of the manuscript was written by Xiao-Feng Xie and Cai-Wen Du. Supervision and project administration was performed by Cai-Wen Du. All authors are responsible for the contents and have read and approved the manuscript for submission.
Data availability
The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.
Ethical approval
All procedures performed involving human participants were in accordance with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. This study was also approved by institutional review boards and ethics committees (ethical code: 2019-35) of National Cancer Center/ National Clinical Research Center for Cancer/Cancer Hospital & Shenzhen Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College.
Informed consent
Informed consent was obtained from all individual participants included in the study. The authors affirmed that human research participants provided informed consent for publication.