Activity of ceritinib in crizotinib-resistant ROS1-rearranged non-small-cell lung cancer patients

As a second-generation selective oral anaplastic lymphoma kinase inhibitor, ceritinib is an effective first-line treatment for c-ros oncogene 1 (ROS1)-rearranged non-small-cell lung cancer (NSCLC). Its efficacy and safety for the treatment of crizotinib-resistant ROS1-rearranged NSCLC were explored in the study. A retrospective single-center study was conducted to investigate the efficacy of ceritinib in crizotinib-resistant ROS1-rearranged NSCLC. The objective response rate was the primary objective, while the disease control rate, progression-free survival and adverse events were secondary objectives. From December 2015 to October 2021, a total of 246 patients with ROS1-rearranged NSCLC were screened, 12 (4.9%) of whom were treated with ceritinib after the development of crizotinib resistance. Among the 12 crizotinib-resistant patients included, 3 displayed the efficacy of partial response and 3 had the efficacy of stable condition. The objective response rate, disease control rate and median progression-free survival of all patients were 25% (95% confidence interval [CI]: −3.7% to 53.7%; 3 of 12 patients), 50% (95% CI: 16.8% to 83.2%; 6 of 12 patients), and 10.5 months (95% CI, 5.7 to 15.3 months), respectively. In addition, of the 6 patients with brain metastases, an intracranial disease control rate of 66.7% (95% CI:12.5% to 120.9%) was obtained. The research results reveal that ceritinib can be a treatment option for ROS1-rearranged NSCLC patients after the development of crizotinib resistance.


Introduction
3] Reportedly, lung adenocarcinoma patients who are young, nonsmokers, and Asians are subject to a higher incidence of ROS1 rearrangement. [4]The ROS1 rearrangement partners include but are not limited to CD74, CCDC6, EZR, FIG, and KDELR2. [5]ecause ROS1 and anaplastic lymphoma kinase (ALK) are structurally similar and share 49% amino acid sequence homology in their kinase domains, ROS1-positive NSCLC is generally sensitive to ALK inhibitors. [6]Crizotinib, an inhibitor of Mesenchymal epithelial transforming factor/ALK/ROS1 tyrosine kinase (TKI), was the first inhibitor approved by the US Food and Drug Administration in 2016 for the treatment of ROS1-positive NSCLC. [7,8]Crizotinib is effective in the initial stage but is eventually resisted by most patients. [9]In addition, its low blood-brain barrier penetration leads to a high risk of brain recurrence. [10,11]ritinib is a second-generation selective oral ALK inhibitor and proves to be effective in the first-line treatment for ROS1positive NSCLC. [12,13]It is recommended in the NCCN guidelines for the treatment of ROS1-positive metastatic NSCLC, based on an open-label, multicenter, phase II study. [14]This study confirmed the effectiveness of ceritinib in ROS1-positive NSCLC patients who were not treated with crizotinib.In addition, the brain-blood contact rate of ceritinib crossing the blood-brain barrier was found to be approximately 15%. [15]A high rate of intracranial response in patients with asymptomatic brain metastases was also identified.However, due to the inclusion of only 2 crizotinib-resistant patients, the efficacy of ceritinib in crizotinib-resistant ROS1-rearranged NSCLC patients remains unknown.
Given financial considerations, crizotinib is the first choice for most Chinese patients. [16]However, no approved second-line targeted therapies are available once resistance to crizotinib has developed among patients. [17]As a second-generation ALK inhibitor, ceritinib can overcome crizotinib resistance in ALKpositive NSCLC patients, but its efficacy in the treatment of The authors have no conflicts of interest to disclose.
The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.Medicine crizotinib-resistant ROS1-rearranged NSCLC patients remains unclear. [13,18]n the present retrospective study, patients who developed the development of resistance to crizotinib and were then treated with ceritinib were included to investigate the activity of ceritinib in crizotinib-resistant ROS1-rearranged NSCLC patients.

Patients and samples
In this retrospective study, ROS1-rearranged NSCLC patients who were treated with ceritinib after the development of crizotinib resistance at the First Affiliated Hospital of Zhengzhou University between January 2016 and December 2021 were enrolled.Their medical records were reviewed and the TNM stage was determined according to the International Association for the Study of Lung Cancer (version 8).This study was approved by the Ethics Review Board of the First Affiliated Hospital of Zhengzhou University.The need for written informed consent was waived owing to the retrospective design of the study.

Study design
A retrospective single-center study was conducted to investigate the activity of ceritinib in crizotinib-resistant ROS1-rearranged NSCLC patients with the primary objective of evaluating the objective response rate (ORR) and secondary objectives of evaluating the disease control rate (DCR), progression-free survival (PFS) and adverse events (AEs).PFS was defined as the time from the beginning of ceritinib treatment to the documented tumor progression or death.Patients who had not progressed/died at the data cutoff in April 2022 were considered to be censored at the last follow-up.ORR was defined as the proportion of patients who experienced a complete or partial response to ceritinib therapy, according to RECIST 1.1.DCR represents the proportion of patients with complete response, partial response (PR) or stable condition (SD).Moreover, AEs were graded according to the Common Terminology Criteria for Adverse Events (CTCAE) (version 5.0).

Data collection and assessment
The data of this study were collected from clinical records on medical history.

Statistical analysis
The median PFS and the number of patients at risk were estimated using the Kaplan-Meier method and associated 95% confidence intervals (CIs).The confidence intervals of the ORR equal proportions were calculated using the Clopper-Pearson method.A Log-rank test was conducted to analyze the differences between survival curves, and univariate Cox proportional hazards regression analyses were performed to examine the impact of clinicopathological factors on PFS.All statistical analyses were conducted using the SPSS version 26.0 software (IBM Corp), where P value <.05 was considered statistically significant.

Results
From January 2016 to December 2021, a total of 246 patients with ROS1-positive NSCLC were screened out, 12 (4.9%) of whom were treated with ceritinib after the development of crizotinib resistance.The included patients received oral ceritinib at the recommended dose of 450 mg/day in continuous 28-day treatment cycles.These 12 patients were composed of 5 females and 7 males, with a median age of 54 years old at diagnosis and an age range of 38 to 70 years old.All the patients were in stage IV.The majority of patients (75%) were never-smokers, and all had adenocarcinoma histology.Among the 12 patients included in the study, 1 was diagnosed with ROS1 gene fusion by immunohistochemistry, 2 were detected by RT-PCR, and the other patients were detected by NGS.Clinicopathological characteristics of the patients are presented in Table 1.

Response
To assess intracranial response, intracranial lesions in patients with brain metastases were investigated (Table 4).At study entry, 6 patients had brain metastases, including 2 with PR, who had not received prior brain radiotherapy and presented a response time of 5 months and 7 months, respectively.In addition, 2 patients were assessed as having SD, and 2 other patients had intracranial lesion progression at the time of the first assessment.Overall, the ORR of intracranial lesions was 33.3% (95% CI: −20.9% to 87.5%) and disease control was achieved in 66.7% (95% CI: 12.5%-120.9%).Overall, the above intracranial results were consistent with systemic responses (Table 3).
The incidence and types of AEs are summarized in Table 5.All 12 patients experienced at least 1 adverse event associated with ceritinib.As expected, gastrointestinal AEs were the most frequent.Notably, the incidences of nausea and vomiting were high (50% and 41.7%, respectively).In addition, 2 patients presented with rash, 1 with liver injury, 1 with vision disorder, and all with grades I to II.Four patients presented with grade 3 or higher toxicity, including nausea, vomiting, decreased appetite and constipation.The incidence of grade 3 and above vomiting was the highest, with severe vomiting occurring in 2 of the 12 patients (16.7%).All these AEs were relieved after symptomatic supportive treatment, and no ceritinib-related fatal AEs occurred.

Discussion
Both crizotinib and ceritinib are recommended by the NCCN guidelines as first-line treatment for patients with advanced ROS1-positive NSCLC. [19]In East Asian patients with ROS1positive advanced NSCLC, the ORR and mPFS after crizotinib treatment were 71.7% and 15.9 months, respectively. [20]Therefore, crizotinib remains the best first-line treatment option for such patients, especially for those from China, followed by the next generation of TKIs when the disease progresses. [21]owever, no approved second-line targeted therapies are available once resistance to crizotinib has developed.
[24] Lorlatinib is a highly brain-penetrant third-generation ALK and ROS1 inhibitor, which has been approved by the US Food and Drug Administration for the treatment of ALK-positive NSCLC. [25]In a recent multicenter, single-arm, Phase I to II study, lorlatinib demonstrated efficacy in ROS1-positive patients previously treated with crizotinib with an ORR of 35% and mPFS of 13.8 months. [22]Therefore, it has been recommended by NCCN guidelines as a follow-up option for ROS1-positive lung cancer with crizotinib or Entrectinib disease progression.Few studies have been conducted on the efficacy of the second-generation ALK inhibitor, brigatinib, for the treatment of ROS1-positive NSCLC after the development of crizotinib resistance.One retrospective study included a total of 7 patients, of whom 2 showed partial responses (ORR 29%) and 1 (14%) was in a stable condition. [26]o the best of our knowledge, this is the first retrospective study to analyze the efficacy of ceritinib in the treatment of  crizotinib-resistant ROS1-positive NSCLC patients.Among the 12 crizotinib-resistant patients included, 3 had an efficacy of PR and 3 had an efficacy of SD, with a disease control rate of 50%, an overall response rate of 25%, and a median PFS of 10.5 months.Because of the blood-brain barrier, the brain is considered a major pharmacological refuge.In fact, approximately 50% of the patients treated with ROS1-TKIs develop disease progression due to central nervous system metastases. [17]A total of 6 patients with brain metastases were included in this study, 2 with a PR response to brain lesions.Overall, the ORR of intracranial lesions was 33.3% (95% CI: −20.9% to 87.5%) and intracranial disease control was achieved in 66.7% (95% CI: 12.5%-120.9%).The study results also indicated that ceritinib might have high activity in the central nervous system.
This study confirms that ceritinib is a treatment option for ROS1-positive NSCLC patients with crizotinib resistance.Apparently, intracranial and systemic activity is related to the underlying molecular mechanisms of crizotinib resistance.Two mechanisms of acquired crizotinib resistance have been reported, namely the ROS1-dependent mechanism and the ROS1-independent mechanism. [27,28]Reportedly, mutations in the ROS1 kinase domain and activation of bypass signaling pathways are the most important mechanisms of crizotinib resistance among the identified mechanisms so far. [29]In addition, approximately 50% to 60% of crizotinib resistance is caused by on-target mutations, of which G2032R is the most frequently observed mutation in ROS1-positive patients. [30,31]Notably, ceritinib, entrectinib, and lorlatinib were found by preclinical studies to have no strong activity against G2032R. [32]Also, in vitro studies have demonstrated that L2026M gatekeeper mutations maintain sensitivity to ceritinib and brigatinib. [26]In this retrospective study, the crizotinib resistance pathway that might be more sensitive to ceritinib was not explored.This should be considered in future research.
This study is still subject to some limitations.First, all enrolled patients were from the same hospital, therefore the results could not represent the entire population.Second, this study was retrospective, and complete clinical information of the patients might not have been collected.Third, progressive and post-progressive biopsies, which are critical for understanding the mechanisms of response and/or resistance, are lacking.Moreover, the sample size was limited.Therefore, the results should be validated in prospective studies with larger sample sizes.

Conclusion
Based on a retrospective analysis of the efficacy of ceritinib in the treatment of ROS1-positive NSCLC patients after the development of crizotinib resistance, ceritinib can be used as a treatment option after the development of crizotinib resistance.However, further research is needed to find out how to screen ceritinib-sensitive patients.

Figure 1 .
Figure 1.Best percentage change from baseline in tumor volume in patients with at least 1 postbaseline measurement.PD = progressive disease, PR = partial response, SD = stable disease.

Figure 2 .
Figure 2. Kaplan-Meier curve of progression-free survival in all patients.

Table 4
Best clinical response in cranial.

Table 5
Adverse events.