This retrospective, single-center study enrolled patients with HER2+ MBC treated with inetetamab at Shandong Frist Medical University and Shandong Academy of Medical Sciences between July 2020 and June 2022. The Ethics Committee and Institutional Review Board of Shandong First Medical University and Shandong Academy of Medical Sciences approved this study (SDTHEC2022012020). All investigations were conducted in accordance with the Declaration of Helsinki.

The inclusion criteria for participants were as follows: female sex, age ≥18 years, histologically or cytologically confirmed MBC with documentation of HER2 overexpression, prior trastuzumab therapy with or without other HER2-targeted treatment, at least one cycle of inetetamab, and complete medical records. The exclusion criteria were non-measurable or non-evaluable lesions and those lost to follow-up. There were no limits to the number of prior cytotoxic regimens for metastatic diseases. The last follow-up was conducted in November 2022. Until the last follow-up date, patients who were lost to follow-up were considered as censored data. All data were retrospectively collected from medical records and laboratory results. Patients or their family members (for patients who already died at the study initiation) provided signed informed consent or oral agreement with tape recording.

The characteristics of the patients at the time of initial diagnosis (including age, ECOG performance status, and menstrual status), tumor characteristics (including tumor size, lymph node involvement, grade, histology, and receptor status), treatment regimen in the (neo)adjuvant and metastatic settings (including chemotherapy, anti-HER2, endocrine regimen, surgery, radiotherapy, dose reductions or delays, etc.) were extracted from electronic medical records. Hormone receptor (HR) was defined as estrogen receptor (ER) and/or progesterone receptor (PR) positivity (ER and PR were determined by at least 10% of positively stained nuclei). HER2 positivity was defined as an immunohistochemistry(IHC) score of 3+ or 2+ together with HER2 gene amplification verified by fluorescence in situ hybridization(FISH+). Disease-free interval (DFI) was defined as the time from surgery to diagnosis of metastasis.

Clinical response was evaluated using computed tomography, magnetic resonance imaging, and physical examination according to the Response Evaluation Criteria in Solid Tumors, version 1.1. The main endpoint was progression-free survival (PFS), defined as the time from treatment initiation until disease progression or death. Other endpoints included the objective response rate (ORR), clinical benefit rate (CBR), and safety. ORR was defined as the proportion of patients who achieved a complete response (CR) or partial response (PR). CBR was defined as the proportion of patients who achieved CR, PR, or stable disease (SD). Adverse events (AEs) were graded based on the National Cancer Institute Common Terminology Criteria for AEs, version 4.0.

The median (range) or percentage of patients was used to represent the clinicopathological characteristics. Continuous variables were analyzed by One-way ANOVA. Categorical variables were assessed by the Pearson’s chi-squared test or Fisher’s exact test. The Kaplan–Meier method was used to estimate PFS. Additionally, Cox univariable model was employed to assess the covariate effects on PFS, and then Cox multivariate models were used to assess the factors with relative significant p-values(p ≤ 0.1) in univariate analysis to PFS with hazard ratios(HR.) and corresponding 95% confidence intervals (CIs). Statistical significance for all analyses was set at p<0.05. GraphPad Prism 9.3.1 software was used to perform all statistical analyses.

A total of 69 patients with HER2+ MBC treated with inetetamab were recruited. After considering the exclusion criteria, 64 (92.8%) patients were included in the study ( Figure 1 ).

The baseline characteristics are presented in Table 1 . The median age of the patients at diagnosis was 46 years (range: 27–67 years), 54 (84.4%) underwent surgery, 14 patients (21.9%) had stage IV BC as their first diagnosis. Moreover, 53.1% of the patients had more than two metastatic sites, with the four most common metastatic sites being the lymph node (48.4%), local sites (35.9%), bone (32.8%), and liver (32.8%). Half of the patients had visceral metastases, whereas 13 (20.3%) had brain metastases. All patients had been exposed to anti-HER2 therapy, with 64.1% prescribed pyrotinib and 18.8% with lapatinib. More than four-fifths of the patients received trastuzumab during salvage treatment. Furthermore, 62.5% of patients received two or more lines of systemic therapy before inetetamab. These results suggest that, in a real-world setting, patients receiving inetetamab are more likely to be heavily pretreated.

The treatment regimens are shown in Table 2 . Most patients were treated with inetetamab in combination with chemotherapy and/or other HER2-targeted therapies. The most common chemotherapy regimens were vinorelbine (n = 39, 60.9%) and abraxane (n = 15, 23.4%). Pyrotinib and inetetamab in combination were administered to 40 (62.5%) patients. Meanwhile, three (3.1%) patients received inetetamab and brain radiotherapy but did not receive any other anti-cancer drugs.

All patients were evaluated for PFS. The median follow-up time was 14.3(12.7–15.9) months. The median progression-free survival (mPFS) was 5.6 (4.6–6.6) months and the ORR was 26.6% ( Figure 2A ).

Patients who received inetetamab‐based therapy as first and later lines of metastatic treatment had a median PFS of 5.7 (1.9–9.5) and 5.3 (3.8–6.8) months, respectively ( Figure 2B ). Thirty-two patients with visceral metastases showed a median PFS of 5.3 (3.9–6.7) months. A total of 31 patients with lymph node metastases, 23 patients with local metastases, 21 patients with bone metastases and 13 patients with brain metastases had median PFS of 5.7 (0.5–10.9) months, 7.0 (0.8–13.2) months, 4.1(3.4–4.8) months and 4.2 (2.0–6.4) months, respectively ( Figure 2C ).

To determine the best combination for inetetamab, we firstly investigated the anti-HER2 treatment in the overall cohort. Baseline characteristics were analyzed in Supplementary Tables 1-3 . The median PFSs among inetetamab plus pyrotinib, inetetamab plus pertuzumab and inetetamab alone were 6.1 (2.5–9.7) months, 2.5 (1.9–3.1) months, and 5.3 (4.6–6.6) months, respectively. Inetetamab plus pyrotinib was the best combination among the three groups (p=0.005) ( Figure 3A ). However, the age of patients treated with inetetamab plus pertuzumab is relatively old than other two cohorts(p=0.001) and 6 patients(100%) received at least 3 lines of rescue treatment(p=0.016), which led to the unbalanced baseline characteristics among three cohorts. As vinorelbine and abraxane were the most common combined cytotoxic drugs, we compared the PFS of different chemotherapies. The median PFSs among inetetamab plus vinorelbine, inetetamab plus abraxane and inetetamab plus other therapeutic agents were 5.7 (3.7-7.7) months, 5.7 (4.4–7.0) months, and 4.0 (1.4–6.6) months, respectively ( Figure 3B ). Furthermore, we compared the efficacy of the combination treatments. The median PFS of inetetamab plus pyrotinib plus vinorelbine was 9.3 (3.1–15.5) months; inetetamab plus pyrotinib plus abraxane, 5.6(0-13.6) months; and inetetamab plus other therapeutic agents, 4.1 (3.0–5.2) months. There were statistically significant differences among the three groups (p = 0.048) ( Figure 3C ). These findings indicate that inetetamab plus pyrotinib plus vinorelbine may be the most effective inetetamab-based regimen. Thirty-one (48.4%) patients received inetetamab plus pyrotinib plus vinorelbine. The subgroup of patients achieved an ORR of 35.5% and CBR of 48.4%, with CR achieved in three patients, PR achieved in eight patients, and SD achieved in four patients ( Figure 3C ).

Thirty-five patients received vinorelbine before inetetamab-based therapy. The mPFS of patients with versus without vinorelbine pretreatment was 5.7 (4.0–7.4) months versus 5.6 (3.5–7.7) months, respectively (p=0.750) ( Figure 4A ). Forty-one patients received pyrotinib before inetetamab‐based therapy. The mPFS of patients with versus without pyrotinib pretreatment was 5.7 (3.5–7.9) months versus 5.3 (3.3–7.3) months, respectively (p=0.988) ( Figure 4B ). These results indicate that the medication history of vinorelbine and/or pyrotinib had no influence on the efficacy of the drug.

We also analyzed the PFS of the patients pretreated with pyrotinib ( Figure 4C ). A total of 41 patients were included in this subgroup analysis, with an ORR of 29.3%. Two patients achieved CR and 10 patients achieved PR. Patients exposed to inetetamab plus vinorelbine plus pyrotinib agent had significantly longer PFS (10.3 (5.2–15.4) months) than those exposed to other therapeutic agents (4.0 (2.0–6.0) months) (p=0.018).

The univariate analysis indicated that age group (<40 vs. ≥40 years), menstrual status (pre vs. post), hormone receptor status (negative vs. positive), and regimens (inetetamab plus vinorelbine plus pyrotinib vs. other therapeutic agents) were correlated with PFS (p<0.05). Next, we constructed a multivariate model with the above factors, ECOG performance status(0-1 vs ≥2) and visceral metastasis (yes vs. no) as covariates for PFS ( Table 3 ). After adjustment, Cox multivariate regression analysis showed that the regimens (inetetamab plus vinorelbine plus pyrotinib vs. other therapeutic agents) and visceral metastasis (yes vs. no) were independent predictors of PFS ( Table 3 ).

Thirty-two patients (50.0%) exhibited visceral metastasis. Patients with and without visceral metastases had PFS times of 5.3 months and 7.0 months, respectively ( Figure 5A ). Of the 32 patients, 18 received inetetamab plus vinorelbine plus pyrotinib treatment, with an ORR of 27.8% and a CBR of 33.3%. One patient achieved CR, four achieved PR, and one achieved SD. The median PFS was significantly different for patients who underwent inetetamab plus vinorelbine plus pyrotinib or other therapeutic agents (6.1 (5.1–7.1) vs. 2.9-(0.9–4.9) months, p=0.002; Figure 5B ).

The safety assessments of etamab-based therapy are listed in Table 4 . After the initial etamab-based therapy, 10 (15.6%) patients in the inetetamab group experienced a dose reduction, and two (3.1%) patients interrupted the treatment. The most common grade 3/4 AEs were leukopenia (4.7%) and neutropenia (3.1%). No treatment-related deaths were reported. Overall, the results show that the safety of etamab-based therapy is controllable and tolerable.