Induction chemotherapy followed by camrelizumab plus apatinib and chemotherapy as first-line treatment for extensive-stage small-cell lung cancer: a multicenter, single-arm trial

ABSTRACT Chemo-immunotherapy is the current first-line treatment for patients with extensive-stage small cell lung cancer (ES-SCLC), but survival benefits are modest. We aimed to evaluate

the safety, antitumor activity and biomarkers of first-line camrelizumab and apatinib plus chemotherapy in untreated ES-SCLC patients. In this single-arm trial (ClinicalTrials.gov

NCT05001412), eligible patients received 2 cycles of etoposide and carboplatin (EC) as induction treatment followed by 2–4 cycles of camrelizumab, apatinib plus EC, then maintenance

camrelizumab plus apatinib. Primary endpoint was safety. Secondary endpoints included objective response rate (ORR), duration of response, progression-free survival (PFS), and overall

survival (OS). Targeted sequencing and whole transcriptome sequencing were performed to explore biomarkers. All enrolled 40 patients were treated and analyzed for safety. During the entire

treatment, treatment-emergent adverse events (TEAEs) occurred in 40 patients (100%), and 30 (75.0%) were grade ≥3. The most common grade ≥3 TEAEs were neutropenia (35.0%), anemia (15.0%) and

increased alanine aminotransferase (15.0%). No treatment-related deaths occurred. Among 36 evaluable patients, ORR was 88.9% (95% CI: 73.9%–96.9%), median PFS was 7.3 months (95% CI:

6.6–9.2) and median OS was 17.3 months (11.8-not reached). Mutations in RB1, high levels of tumor mutation burden, natural killer cells, and interferons, and low levels of cancer-associated

fibroblasts, correlated with prolonged PFS. Induction chemotherapy followed by camrelizumab, apatinib plus EC demonstrated acceptable safety and promising antitumor activity in untreated

ES-SCLC patients. The identified biomarkers need further validation. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT05001412. SIMILAR CONTENT BEING VIEWED BY OTHERS BIOMARKER-DIRECTED,

PEMBROLIZUMAB-BASED COMBINATION THERAPY IN NON-SMALL CELL LUNG CANCER: PHASE 2 KEYNOTE-495/KEYIMPACT TRIAL INTERIM RESULTS Article 10 July 2023 CEMIPLIMAB PLUS CHEMOTHERAPY VERSUS

CHEMOTHERAPY ALONE IN NON-SMALL CELL LUNG CANCER: A RANDOMIZED, CONTROLLED, DOUBLE-BLIND PHASE 3 TRIAL Article Open access 25 August 2022 NEOADJUVANT CAMRELIZUMAB (AN ANTI-PD-1 ANTIBODY)

PLUS CHEMOTHERAPY OR APATINIB (A VEGFR-2 INHIBITOR) FOR INITIALLY UNRESECTABLE STAGE II–III NON-SMALL-CELL LUNG CANCER: A MULTICENTRE, TWO-ARM, PHASE 2 EXPLORATORY STUDY Article Open access

14 June 2024 INTRODUCTION Small-cell lung cancer (SCLC) is an aggressive and rapidly progressing subtype of lung cancer, comprising about 15% of all lung cancer diagnoses.1 SCLC is

associated with a dismal prognosis, with a median overall survival (OS) limited to just 7 months.2 At diagnosis, approximately 70% of SCLC patients are found to have extensive-stage disease

(ES-SCLC).3 In 2019, the FDA approved a chemo-immunotherapy regimen combining platinum-based chemotherapy and the PD-L1 inhibitor atezolizumab, based on the results from the IMpower133

trial.4 This marked a significant step forward in the treatment landscape of ES-SCLC and established chemo-immunotherapy as the standard of care for newly diagnosed ES-SCLC patients.

Subsequent phase 3 trials also substantiated the clinical benefits of incorporating PD-L1 inhibitors into first-line regimens, demonstrating improved outcomes in this aggressive

disease.5,6,7,8 Despite these advancements, the survival improvements are modest, with median OS extending by just 2.0 to 4.7 months.5,6,7,8 Hence, novel therapies are urgently needed to

further improve outcomes for patients with ES-SCLC. Recent studies have revealed the highly heterogeneous and immunosuppressive microenvironment of SCLC, and insufficient CD8 + T cell

infiltration is a key reason for limited response to immune checkpoint inhibitors (ICIs) in SCLC.9 The aggressive growth and invasiveness of SCLC are driven by angiogenesis, and vascular

endothelial growth factor (VEGF) overexpression correlates with poor prognosis in patients with SCLC.10,11 The overexpressed VEGF downregulates endothelial adhesion molecules, like ICAM-1

and VCAM-1, thereby reducing immune cell adhesion and migration.12 Thus, targeting the VEGF pathway might increase CD8 + T cells infiltration and reduce neovascularization, potentially

enhancing the antitumor response in ES-SCLC. Combining ICIs with anti-VEGFR agents has synergistic effects via increasing T-cell infiltration in the tumor microenvironment.13 Preclinical

data have shown that apatinib, a VEGFR inhibitor, can modulate the tumor microenvironment by decreasing tumor hypoxia, enhancing CD8 + T cell infiltration, and decreasing the accumulation of

tumor-associated macrophages in lung cancer tissues.14 In lung cancer mouse models, the combination of apatinib with anti-PD-L1 antibodies led to significant suppression of tumor growth and

metastasis while prolonging mouse survival.14 The combination strategy of ICIs, anti-VEGFR agents, and chemotherapy has shown promising efficacy in patients with lung cancer. Specifically,

in the IMpower 150 trial, first-line therapy with atezolizumab plus bevacizumab and platinum-based chemotherapy notably improved OS (median OS: 19.2 months versus 14.7 months; hazard ratio

[HR] = 0.78) and objective response rate (ORR; 63.5% versus 48.0%) compared with bevacizumab combined with platinum-based chemotherapy in patients with non-small cell lung cancer (NSCLC).15

Additionally, the incorporation of an anti-VEGFR agent, bevacizumab, into cisplatin and etoposide also enhanced progression-free survival (PFS; median PFS: 6.7 months versus 5.7 months; HR =

 0.72) and ORR (55.3% versus 58.4%; odds ratio=1.13) compared with cisplatin and etoposide alone in newly diagnosed ES-SCLC patients.16 Recently, the ETER 701 trial demonstrated the efficacy

of combining ICIs, anti-VEGFR agents, and platinum-based chemotherapy in ES-SCLC patients in the first-line setting.17 This combination regimen, comprising benmelstobart, anlotinib,

carboplatin, and etoposide significantly prolonged OS (median OS: 19.3 months versus 11.9 months; HR = 0.61) and increased ORR (81.3% versus 66.8%) compared with carboplatin and etoposide

alone in patients with ES-SCLC.17 However, in the clinical setting, most SCLC cases are centrally located near the hilum and large blood vessels,2 which carry a high risk of bleeding.

Previous trials of anti-VEGFR agent combinations for SCLC typically excluded patients with large vessel invasion or high bleeding risk.17,18 This exclusion poses a challenge to the clinical

application or optimization of anti-angiogenic combination strategies in this subset of ES-SCLC patients. Addressing this gap is essential, as these patients represent a considerable

proportion of the ES-SCLC population. Apatinib, a VEGFR2-targeting tyrosine kinase inhibitor, and camrelizumab, a PD-1 inhibitor, has each shown notable antitumor activity in ES-SCLC

patients beyond the first-line treatment.19,20 The PASSION study reported that camrelizumab plus apatinib yielded promising antitumor activity and was well-tolerable in ES-SCLC patients,

including those with responsive and resistant to chemotherapy, in the second-line setting.21 A retrospective study showed that first-line camrelizumab plus chemotherapy, then maintenance

camrelizumab plus apatinib, provided better survival benefits compared with PD-L1 inhibitors plus chemotherapy and exhibited strong antitumor activity.22 At the ASCO 2024 meeting, this

combination also showed efficacy (ORR: 82.14%; median PFS: 7.56 months) and tolerable safety in untreated ES-SCLC patients.18 In this study, we aimed to evaluate the safety and antitumor

activity of first-line induction etoposide and carboplatin (EC), followed by a combination of camrelizumab, apatinib, and EC in ES-SCLC patients, including those with large vessel invasion.

The choice of induction chemotherapy was based on its high ORR of 97% after 2 treatment cycles in limited-disease SCLC.23 This suggests that for most patients, this approach could promote

tumor shrinkage and separation from surrounding vasculature, thereby reducing the bleeding risk associated with anti-VEGFR agents. Moreover, this approach could induce immunogenic cell

death, thereby enhancing the efficacy of subsequent immunotherapy.24 When we designed this study, chemotherapy alone was also the standard recommended first-line regimen for patients with

ES-SCLC.25 Concurrently, we identify potential biomarkers that could predict clinical response. RESULTS PATIENTS AND TREATMENT Between 21 January 2021 and 20 August 2022, 40 patients were

included and received induction EC (Fig. 1). After one cycle of induction EC, 4 patients withdrew informed consent and were unevaluable for tumor response. Thus, 40 patients were evaluable

for safety, and 36 patients were evaluable for tumor response. Of 40 patients, the median age was 60 years (range: 40-73), and 36 (90.0%) were male. All 40 patients (100%) presented with

stage IV disease. Most patients had central SCLC (33/40, 82.5%) and had an Eastern Cooperative Oncology Group (ECOG) performance status (PS) of 1 (31/40, 77.5%). Table 1 presents the

baseline characteristics. Vascular invasion was assessable in 35 patients, and all of them had large vessel invasion (Table 1, Supplementary Table 1). In total, 36 patients completed initial

2 cycles of induction EC. Subsequently, one patient discontinued treatment due to disease progression. Thus, 35 patients received camrelizumab plus apatinib plus EC. During this phase,

additional 5 patients discontinued treatment due to progressive disease (_n_ = 4) and withdrawal of informed consent (_n_ = 1). Finally, 30 patients entered the maintenance treatment phase

with camrelizumab plus apatinib. The reasons for treatment discontinuation during maintenance treatment were progressive disease (_n_ = 24) and adverse events (_n_ = 2). At the data cut-off

on May 30, 2023, the median duration of follow-up was 20.6 months (range: 4.1–27.5). 3 (8.3%) of 36 patients were still on the study treatment. 30 patients (83.3%) received 6 cycles of EC

(Supplementary Table 2). The median treatment cycles of camrelizumab were 7.0 (range: 2.0–35.0). The median treatment duration of apatinib was 5.23 months (range: 0–24.2). The reasons for

not completing 4 cycles of camrelizumab plus apatinib plus EC are provided in Supplementary Table 3. SAFETY During the entire treatment phase, any grade treatment-emergent adverse events

(TEAEs) occurred in 40 patients (100%), with the most common being leukopenia (31 [77.5%]), anemia (28 [70.0%]), and neutropenia (25 [62.5%]) (Table 2). Grade 3/4 TEAEs occurred in 30

patients (75.0%), with the most common being neutropenia (14 [35.0%]), anemia (6 [15.0%]), and increased alanine aminotransferase (6 [15.0%]). No deaths were considered related to study drug

by the investigator. Serious adverse events occurred in 6 (15.0%) patients. Any grade immune-related adverse events (irAEs) occurred in 30 patients (85.7%) and grade 3/4 irAEs occurred in 9

patients (25.7%). The most common irAEs were hyperthyroidism (13 [37.1%]), increased alanine aminotransferase (8 [22.9%]), and increased thyroid stimulating hormone (7 [20.0%])

(Supplementary Table 4). Adverse events (AEs) related to apatinib are presented in Supplementary Table 5. No grade ≥ 3 bleeding events related to apatinib were observed. AEs led to treatment

discontinuation of camrelizumab in 4 patients (11.4%) and apatinib in 2 patient (5.7%), including grade 3 pneumonitis, grade 3 hyponatremia, grade 2 muscle spasm, and grade 2 pulmonary

tuberculosis (_n_ = 1 each). No AEs led to treatment discontinuation of chemotherapy. AEs led to treatment delay of camrelizumab in 12 patients (34.3%) and treatment interruption of apatinib

in 7 patients (20.0%). AEs led to dose reduction of apatinib in 2 patients (5.7%) and chemotherapy in 3 patients (7.5%). EFFICACY After 2 cycles of induction EC, 24 of 36 patients had an

objective response (66.7%, 95% CI: 49.0–81.4); 35 of 36 patients had disease control (97.2%, 95% CI: 85.5–100) (Table 3). Among them, 24 (66.7%) patients had partial response (PR), and 11

(30.6%) had stable disease (SD). Notably, 72.7% (8/11) of those with initial SD then achieved PR after combination treatment with camrelizumab, apatinib plus EC, followed by maintenance

camrelizumab plus apatinib. After the entire treatment, 32 of 36 patients had an objective response (88.9%, 95% CI: 73.9–96.9); 35 of 36 patients had disease control (97.2%, 95% CI:

85.5–100) (Table 3). Among them, 32 (88.9%) had PR, and 3 (8.3%) had SD. In total, 34 patients (94.4%) had a decrease in tumor size of target lesions from the baseline. Median best change

from baseline was −63.3% (Fig. 2a). Median duration of response (DoR) was 5.4 months (95% CI: 4.2–7.7; Fig. 2b–d). Median time to response (TTR) was 1.5 months. The overall ORR in all 40

patients was provided in the Supplementary Table 6. At the data cut-off on May 30, 2023, the median duration of follow-up was 20.6 months (range: 4.1–27.5). Of the 36 evaluable patients, 30

(83.3%) had disease progression or deaths (_n_ = 21). The median PFS was 7.3 months (95% CI: 6.6–9.2, Fig. 2e). The median OS was 17.3 months (95% CI: 11.8-NR). The 12-month OS rate was

63.4% (95% CI: 45.4%–76.9%) (Fig. 2f). EXPLORATORY BIOMARKERS We evaluated the association between genomic alteration and clinical outcomes. Baseline tissue samples were available from 30

patients for targeted gene sequencing and from 21 patients for whole transcriptome sequencing (WTS). TP53 (97%) and RB1 (90%) were the most frequently mutated genes (Fig. 3a). No significant

correlation between genomic mutations and response (complete response/PR) during the induction treatment and the entire treatment was observed. Mutations in RB1 were associated with longer

PFS (_P_ < 0.001; HR, 0.10; 95% CI: 0.02–0.45), whereas mutations in PTPRD (_P_ = 0.02; HR, 3.85; 95% CI: 1.17–12.67) and the mTOR signaling pathway genes (_P_ = 0.005; HR, 3.97; 95% CI:

1.43–11.06) were associated with shorter PFS (Fig. 3b–d). A trend toward shorter PFS was observed in patients with mutations in SPTA1 (P = 0.06; HR, 2.22; 95% CI: 0.94–5.25) (Fig. 3e). We

observed that tumor mutation burden (TMB) status and homologous recombination deficiency (HRD) score did not significantly correlate with response (complete response/PR) during the induction

treatment and the entire treatment (Fig. 4a, b). High TMB (TMB ≥ 7.0) correlated with longer PFS (_P_ = 0.008; HR, 0.26; 95% CI: 0.09–0.74) (Fig. 4c). Patients with high HRD score (HRD ≥ 

34.0) had trend toward longer PFS (_P_ = 0.07; HR, 0.40; 95% CI: 0.15–1.12) (Fig. 4d). Immune cell infiltration analysis revealed that high levels of NK cells (_P_ = 0.002; HR, 0.08; 95% CI:

0.01–0.61) and interferons (_P_ = 0.004; HR, 0.13; 95% CI: 0.03–0.63) correlated with longer PFS, whereas high cancer-associated fibroblasts levels correlated with shorter PFS (_P_ = 0.001;

HR, 5.55; 95% CI: 1.74–17.69) (Fig. 4e–g). DISCUSSION As far as we are aware, this is the first report to investigate the safety, antitumor response and potential biomarkers of PD-1

inhibitors plus anti-VEGFR agents and chemotherapy in ES-SCLC patients. In this single-arm trial, induction EC followed by a combination of camrelizumab, apatinib and EC, and subsequent

maintenance with camrelizumab plus apatinib, had a manageable safety profile in untreated ES-SCLC patients. Notably, this regimen showed promising antitumor activity, with ORR of 88.9% and

disease control rate (DCR) of 97.2%. Median PFS was 7.3 months and median OS was 17.3 months. Our findings offered valuable insights for future clinical trials incorporating

chemo-immunotherapy with anti-VEGFR agents in ES-SCLC. The safety profile observed for our combination regimen of EC, camrelizumab, and apatinib aligns with previously reported toxicity

profiles for these agents as monotherapies. We did not observe unexpected toxicities. The most common grade ≥3 TEAEs were leukopenia, anemia and increased alanine aminotransferase, which

were mostly chemotherapy related. Most toxicities were manageable with dose adjustments and supportive care. Given that most cases of SCLC were centrally located and near major blood

vessels, the use of anti-angiogenic agents could increase the risk of bleeding.2,26 In this study, 82.5% of patients enrolled were central ES-SCLC, and all patients evaluable for vascular

invasion had large vessel invasion. However, no grade ≥ 3 bleeding events were observed. These findings suggest that first-line combination of EC, camrelizumab, and apatinib was tolerable in

ES-SCLC patients. The IMpower133 and CASPIAN studies reported that first-line atezolizumab or durvalumab plus EP had ORRs of 60.2% and 79%, median PFS of 5.2 and 5.1 months, and median OS

of 12.3 and 13.0 months, respectively.5,6 The CAPSTONE-1 and ASTRUM-005 studies reported that adebrelimab or serplulimab plus EC had ORRs of 70.4% and 80.2%, median PFS of 5.8 and 5.8

months, and median OS of 15.3 and 15.4 months, respectively.7,8 A retrospective analysis showed that first-line camrelizumab plus EP/EC had an ORR of 65.7%, DCR of 74.3%, median PFS of 7.4

months, and median OS of 12.5 months.27 In this study, the ORR was 88.9%, with a DCR of 97.2%, median PFS of 7.3 months, and median OS of 17.3 months. Moreover, this study consisted of

patients with more severe disease, all at stage IV. These results suggest that the combination provides favorable data over first-line chemotherapy plus ICIs in ES-SCLC, despite potential

confounding factors from crossover trial comparisons such as study design, patient selection and chemotherapy regimens. This is consistent with high antitumor response reported for such a

combination in SCLC and other solid tumors.17,28,29,30,31 This also aligns with evidence that anti-VEGFR agents can synergically improve the efficacy of immunotherapy in cancer treatment.32

Overall, combining chemo-immunotherapy with anti-VEGFR agents may enhance clinical outcomes for patients with previously untreated ES-SCLC. Due to bleeding concerns, studies like the ETER

701 trial on anti-angiogenic drug combinations typically excluded patients with large vessel invasion.17 This exclusion challenges the optimization of therapeutic strategies for ES-SCLC with

large vessel invasion, highlighting the need for broader inclusion criteria in clinical trials. Unlike the ETER701 trial, we used induction chemotherapy before the combination therapy to

reduce bleeding events. At baseline, all 35 evaluable patients had large vessel invasion and were at high risk of bleeding. Nonetheless, the incidence of hemoptysis was similar to that

reported in the ETER701 trial,17 with no grade ≥3 hemoptysis or other severe bleeding events. Moreover, our regimen achieved clinical outcomes comparable to benmelstobart and anlotinib plus

EC in the ETER701 trial (ORR: 81.3%; median PFS: 6.9 months).17 These findings suggest that induction chemotherapy combined with camrelizumab and apatinib demonstrates promising efficacy in

ES-SCLC. Only one patient experienced PD during the 2 cycles of induction chemotherapy, suggesting a low risk of PD during extended induction therapy. However, given the limited patient

number, the risk-benefit of induction therapy in ES-SCLC requires further investigation. Overall, this study provides important data for future research to broaden the population eligible

for first-line anti-angiogenic combinations in ES-SCLC. Consistent with the ETER701 trial and other anti-angiogenesis, immunotherapy, and chemotherapy combination trial,17,31 disease

progression was the main reason for treatment discontinuation in this study. This suggests that despite the clinical benefits of such a combination, most ES-SCLC patients may develop

therapeutic resistance. Thus, new first-line treatments for ES-SCLC are still needed to overcome therapeutic resistance. Additionally, given that 82.8% (24/29) of disease progression

occurred during the maintenance phase, more robust maintenance treatments may be feasible. Currently, predictive biomarkers for ICIs in SCLC remain lack. The CASPIAN study demonstrated that

PD-L1 expression level, a predictive biomarker for ICIs in various cancers, did not correlate with response to ICIs in SCLC.5 Only about 25% of SCLC patients had PD-L1 expression on ≥ 1% on

tumor cells.33 This necessitates the exploration for new biomarkers. Our study identified several candidate predictive biomarkers via targeted sequencing and WTS. The most frequent mutation

occurred in TP53 and RB1, consisting with prior reports.34 Previous studies indicated that TMB seemed to correlate with clinical activity in patients treated with single agent immunotherapy

(nivolumab monotherapy) or PD-1 plus CTLA-4 blockage (nivolumab plus ipilimuma), but not for chemo-immunotherapy combinations in ES-SCLC.6,35,36 In the current study, patients with high TMB

had better survival outcomes. These controversial results may be due to small sample size, the lack of standardized assays and score methods. The role of TMB in predicting

chemo-immunotherapy in SCLC remains to be determined and warrants further investigation. Additionally, our analysis revealed longer PFS in patients with high NK cells, interferons, and RB1

mutations, whereas high cancer-associated fibroblasts levels and mutations in the mTOR signaling pathway and PTPRD were indicators of shorter PFS. The positive correlation between NK cells

levels and PFS aligns with previous findings in ES-SCLC patients treated with first-line chemo-immunotherapy.37 However, the negative correlation between mutations in the mTOR signaling

pathway and PTPRD and PFS differs from previous studies, which reported that mutations in the mTOR signaling pathway and PTPRD correlated with longer PFS in NSCLC patients treated with

chemo-immunotherapy.38,39 These discrepancies may be due to the small sample size or different treatment regimens. Given the significant heterogeneity and the complex regulation of tumor

immune microenvironment in SCLC, single biomarker seems inadequate to accurately predict the efficacy of chemo-immunotherapy. This underlines the need to develop predictive models and

monitor biomarker dynamics during treatment in the further studies. Altogether, the identified biomarkers may help guide patient selection for camrelizumab plus apatinib and chemotherapy.

However, further validation studies are needed before clinical application. This study has several limitations. First, our study provides novel and valuable data into the safety and clinical

activity of platinum-based chemotherapy combined with camrelizumab and apatinib plus as first-line regimen for ES-SCLC. However, the study design was not randomized with no comparative

groups; thus, the antitumor activity was preliminary and cannot be analyzed for causality. Further randomized clinical studies are warranted to confirm the exact contribution of adding

anti-VEGFR agents. Second, although incorporating biomarker analysis, limited availability of tissue sample warrants cautious interpretation of these results. Future studies of camrelizumab

and apatinib plus chemotherapy in ES-SCLC, including body fluids biomarkers, are expected to build on our findings. In conclusion, induction EC followed by a combination of camrelizumab,

apatinib and EC, and subsequent maintenance camrelizumab plus apatinib, showed a tolearble safety and promising antitumor activity, suggesting its potential as a first-line therapy option

for patients with ES-SCLC. Identified predictive biomarkers including gene mutations, TMB, and tumor microenvironment components. METHODS AND MATERIALS The present study was approved by the

ethical committee of the First Affiliated Hospital of Guangzhou Medical University (Guangzhou, China, No. 2020-189, 2021-1-13). The study was performed in accordance with the Declaration of

Helsinki and Good Clinical Practice Guidelines. Written informed consent was provided by every participant before study entry. This trial was registered on ClinicalTrials.gov (NCT05001412).

PATIENTS In this multicenter, single-arm study, enrolled patients were aged 18-75 years with histologically confirmed ES-SCLC who were treatment-naïve for ES-SCLC. Prior radiotherapy and

chemotherapy for limited-stage SCLC were acceptable if a minimum of 6 months interval had elapsed between the last treatment and the diagnosis of ES-SCLC. Key inclusion criteria were an ECOG

PS of 0 or 1, measurable lesions per RECIST v1.1, a minimum life expectancy of 12 weeks, and normal organ functionality. Patients were eligible if they were previously treated with

apatinib, anti-PD-1 or anti-PD-L1 therapies. Patients with asymptomatic brain metastases not requiring immediate radiotherapy were permitted to participate. PROCEDURES Eligible patients

received 2 cycles of induction etoposide (100 mg/m2, IV, on days 1 to 3) and carboplatin (AUC 5 mg/mL/min, IV, on day 1), then 2-4 cycles of camrelizumab (200 mg, IV, on day 1) and apatinib

(250 mg, orally, daily) plus etoposide (100 mg/m2, IV, on days 1 to 3) and carboplatin (AUC 5 mg/mL/min, IV, on day 1). Patients continued maintenance treatment with camrelizumab (200 mg,

IV, on day 1) and apatinib (250 mg, orally, daily) until disease progression, intolerable toxicity, death, consent withdrawal or up to 24 months. Each treatment cycle was 3 weeks. Safety was

evaluated at each treatment cycle. AEs were graded according to CTCAE v5.0 and reported from the day of informed consent signing until 30 days post the final study drug dose. Tumor response

was assessed by investigators according to RECIST v1.1, using computed tomography or magnetic resonance imaging. Assessments were conducted at baseline, on day 21 of cycles 1 and 2, and

then every 2 cycles until disease progression or the start of a new treatment. Subsequent to treatment discontinuation, survival was follow-up every 3 months, until the patient was dead or

lost to follow-up. OUTCOMES The primary endpoint was safety, graded according to CTCAE v5.0. The ORR, DCR, DoR, TTR, PFS, and OS were the secondary endpoints (SUPPLEMENT). Exploratory

endpoints analyzed the association between treatment response and pretreatment biomarkers, including TMB, gene mutation, HRD score and immune cells. Biomarker analysis was performed for

patients who had sufficient tissue samples at baseline using targeted sequencing and WTS. DNA ISOLATION AND CAPTURE-BASED TARGETED DNA SEQUENCING The methods for DNA isolation and DNA

sequencing were as previously described in ref. 40. In brief, tissue DNA was extracted from formalin-fixed, paraffin-embedded (FFPE) tumor tissues using QIAamp DNA FFPE tissue kit (Qiagen,

Hilden, Germany). DNA fragments between 200 and 400 bp were then purified using Agencourt AMPure XP Kit (Beckman Coulter, CA, USA). A commercial panel consisting of 520 genes (OncoScreen

Plus), spanning 1.86 megabases of the human genome was used for target capture. Indexed samples were sequenced on Nextseq 500 (Illumina, Inc., CA, USA) with paired-end reads and average

sequencing depth of 1000× for tissue samples. All the procedures were performed in a commercial clinical laboratory (Burning Rock Biotech) accredited by the College of American Pathologist

and certified by the Clinical Laboratory Improvement Amendments (CLIA). DNA SEQUENCING DATA ANALYSIS The sequencing data were first mapped to the reference human genome (hg19) using

Burrows-Wheeler Aligner version 0.7.10. Local alignment optimization, duplication marking and variant calling were performed using Genome Analysis Tool Kit version 3.2, and VarScan version

2.4.3, respectively. Sequencing data of corresponding white blood cells were used to filter out germline variants and clonal haematopoiesis. Base calling in tissue samples required at least

8 supporting reads for single nucleotide variations (SNVs) and 5 supporting reads for insertion-deletion variations (Indels), respectively. The mutation status of mTOR pathway was determined

by the presence or absence of any mutation in the mTOR-related genes (Supplementary Table 7). Patients with any mutation in the mTOR-related genes were classified as mTOR pathway mutant,

whereas those without mutations were classified as wild-type. TMB per patient was computed as a ratio between the total number of non-synonymous mutations detected and the total coding

region size of the panel. Loss of heterozygosity (LOH), telomere allelic imbalance (TAI) and large-scale state transitions (LST) were calculated as previously described.41 The HRD score was

calculated as the sum of LOH, TAI and LST scores. RNA SEQUENCING AND DATA ANALYSIS The methods for RNA sequencing were as previously described in ref. 42. RNA was isolated from FFPE samples

using an AllPrep DNA/RNA FFPE Kit (Qiagen, Hilden, Germany). Fragmented RNA was subjected to strand-specific cDNA synthesis, followed by dA-tailing, unique molecular identifier (UMI) adaptor

ligation, PCR amplification, and hybridization with capture probe baits. The prepared NGS libraries were sequenced on a NovaSeq 6000 system (Illumina, Inc., San Diego, CA, USA). The immune

cell scores were generated using single-sample Gene Set Enrichment Analysis (ssGSEA) of the corresponding gene set obtained from previously published literature (Supplementary Table

7).43,44,45 The upper quartile of each immune cell score was used as the threshold to classify patients into high- and low-expression groups. STATISTICAL ANALYSIS This study was exploratory

and designed to detect signals for further investigation, with safety as the primary endpoint. No statistical hypothesis was made, and no formal sample size calculation was performed.

Enrollment of approximately 40 patients was planned and considered sufficient to capture preliminary safety signals. Safety was assessed in all patients who received at least one dose of

study drug. Efficacy was assessed in patients who had at least one post-treatment tumor evaluation. All statistical analyses were performed using R version 3.4.1. Safety outcomes were

summarized with descriptive statistics. The 95% CIs for ORR and DCR were calculated using the Clopper-Pearson method. Survival outcomes were assessed by the Kaplan-Meier method. We estimated

95% CIs for survival outcomes using the Brookmeyer-Crowley method. DATA AVAILABILITY The data collected in this study are available from the corresponding author on reasonable request. Raw

sequence data of this paper have been stored in the China National Center for Bioinformation (https://ngdc.cncb.ac.cn/omix, OMIX ID: OMIX008706 and OMIX008707). REFERENCES * Nicholson, A. G.

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CAS  PubMed  PubMed Central  Google Scholar  Download references ACKNOWLEDGEMENTS The authors thank the patients and their families for their participation in this study, and also the study

teams at each of the study sites. We thank Aichun Chen and Xinwen He for data interpretation, and You Tian for medical writing assistance, all from the Department of Medical Affairs,

Jiangsu Hengrui Pharmaceuticals Co., Ltd. We thank Xinze Lv, Xiao Mi, Ting Hou, Guoqiang Wang, Zhou Zhang, and Shangli Cai for supporting data analysis, all from Burning Rock Biotech. This

study was supported by grants from National Natural Science Foundation of China (82270065), National Key Research and Development Program of China (2022YFF1203300), Natural Science

Foundation of Guangdong Province of China (2023A1515010886), Clinical and Epidemiological Research Project of State Key Laboratory of Respiratory Disease (SKLRD-L-202405), Major Project of

Guangzhou National Laboratory (GZNL2024A02004). AUTHOR INFORMATION Author notes * These authors contributed equally: Ming Liu, Guihuan Qiu, Wenhui Guan AUTHORS AND AFFILIATIONS * State Key

Laboratory of Respiratory Disease, National Clinical Research Center for Respiratory Disease, National Center for Respiratory Medicine, Department of Pulmonary and Critical Care Medicine,

Guangzhou Institute of Respiratory Health, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, China Ming Liu, Guihuan Qiu, Wenhui Guan, Xiaohong Xie, Xinqing Lin, 

Zhanhong Xie, Jiexia Zhang, Yinyin Qin, Shiyue Li, Nanshan Zhong & Chengzhi Zhou * Minimally Invasive Cancer Treatment Center, Guangdong Second Provincial General Hospital, Guangzhou,

China Haijian Du * Department of Pulmonary and Critical Care Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China Xin Chen * Department of Radiology, The First

Affiliated Hospital of Guangzhou Medical University, Guangzhou, China Yu Deng Authors * Ming Liu View author publications You can also search for this author inPubMed Google Scholar *

Guihuan Qiu View author publications You can also search for this author inPubMed Google Scholar * Wenhui Guan View author publications You can also search for this author inPubMed Google

Scholar * Xiaohong Xie View author publications You can also search for this author inPubMed Google Scholar * Xinqing Lin View author publications You can also search for this author

inPubMed Google Scholar * Zhanhong Xie View author publications You can also search for this author inPubMed Google Scholar * Jiexia Zhang View author publications You can also search for

this author inPubMed Google Scholar * Yinyin Qin View author publications You can also search for this author inPubMed Google Scholar * Haijian Du View author publications You can also

search for this author inPubMed Google Scholar * Xin Chen View author publications You can also search for this author inPubMed Google Scholar * Yu Deng View author publications You can also

search for this author inPubMed Google Scholar * Shiyue Li View author publications You can also search for this author inPubMed Google Scholar * Nanshan Zhong View author publications You

can also search for this author inPubMed Google Scholar * Chengzhi Zhou View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS Ming Liu, Shiyue

Li, Guihuan Qiu, Nanshan Zhong and Chengzhi Zhou were the principal authors in conception and design of this study. Ming Liu, Shiyue Li, Guihuan Qiu, Wenhui Guan, Xiaohong Xie, Xinqing Lin,

Zhanhong Xie, Jiexia Zhang, Yinyin Qin, Haijian Du, Xin Chen, and Yu Deng contributed to the data collection and organization. Ming Liu, Shiyue Li and Guihuan Qiu wrote the initial draft,

and all authors reviewed and edited the final version. All authors have approved the article. CORRESPONDING AUTHORS Correspondence to Shiyue Li, Nanshan Zhong or Chengzhi Zhou. ETHICS

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http://creativecommons.org/licenses/by/4.0/. Reprints and permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Liu, M., Qiu, G., Guan, W. _et al._ Induction chemotherapy followed by camrelizumab

plus apatinib and chemotherapy as first-line treatment for extensive-stage small-cell lung cancer: a multicenter, single-arm trial. _Sig Transduct Target Ther_ 10, 65 (2025).

https://doi.org/10.1038/s41392-025-02153-7 Download citation * Received: 12 August 2024 * Revised: 07 January 2025 * Accepted: 22 January 2025 * Published: 18 February 2025 * DOI:

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