br STAR Methods br Acknowledgments The authors

STAR★Methods
Acknowledgments The authors would like to acknowledge Genentech DNA Sequencing, Biologics Resource, Oligo, FACS, and Bioinformatics groups for their help with the project. Our thanks to Craig Cumming and Sophia Maund for facilitating our access to large-scale mutation data. We also want to thank Gerard Manning and Gabriele Schaefer for providing critical input during the course of this work. We wish to thank Allison Bruce for her help with the artwork. This work was supported by grants from the National Institute of General Medical Sciences to N.J. (R01 GM109176), Susan G. Komen Foundation Career Award to N.J. (CCR14299947), National Cancer Institute to T.M.T. (F32 CA216928), HHMI Gilliam Fellowship to M.M., UCSF Discovery Fellowship to M.M., and National Institute of General Medical Sciences to W.F.D. (R35 GM122603 and R01 GM117593).
Introduction Human epidermal growth factor receptor 2 (HER2) overexpression/amplification is observed in 7%–34% of patients with gastric cancer [1], [2], [3], [4], [5], [6]. In the phase III ToGA trial of trastuzumab with cisplatin and fluoropyrimidine-based doublet chemotherapy, a significant improvement in survival was observed for patients with HER2-positive gastric cancer [7]. Therefore, addition of trastuzumab to chemotherapy is now the standard first-line treatment for patients with HER2-positive advanced or recurrent gastric cancer. Although trastuzumab prolongs survival in this population, responses are rarely complete and resistance develops invariably. On the other hand, in the second-line setting, anti-HER2 therapy has not demonstrated a survival advantage until now. In the TyTAN trial, the addition of lapatinib to second-line paclitaxel was not superior to the placebo plus paclitaxel [8]. In the GATSBY trial, trastuzumab emtansine (T-DM1) did not demonstrate superiority to taxane monochemotherapy [9]. There are several possible explanations for these unexpected results. In breast cancer, therapies which include trastuzumab may preferentially eradicate HER2-positive cancer cells, allowing HER2-negative clones to become renin inhibitor [10], [11], [12]. This phenomenon may be more frequently observed in gastric cancer because of the heterogeneity of HER2 expression [13]. It is also possible that there are coexisting oncogenic drivers that may function as negative predictors of trastuzumab benefit and could potentially be exploited as therapeutic cotargets. Several candidate alterations (EGFR and/or MET amplification, EGFR/MET/HER3/PI3K/PTEN mutations, etc.) may co-occur with HER2 overexpression/amplification and putatively confer trastuzumab resistance [14]. However, the biological mechanisms of acquired resistance to anti-HER2 therapy after use of trastuzumab for gastric cancer are not well understood.
Materials and methods
Results
Discussion HER2 loss is proposed to be one of the biological reasons explaining the failure of anti-HER2 therapy after resistance to trastuzumab in HER2-positive gastric cancer. Janjigian et al. [17] reported silica HER2 loss was found in 8 of 23 cases (34.8%). Pietrantonio et al. [18] reported loss of HER2 positivity and HER2 overexpression after trastuzumab-based therapy in 6 of 19 (31.6%) and 7 of 22 (31.8%) samples, respectively. In our study, HER2 loss was found in approximately 60% of 33 patient samples, which represents a relatively high frequency. Pietrantonio et al. [18] also showed that HER2 loss preferentially occurred in cases with initial IHC 2+. Consistently, in our study, HER2 loss tended to be more frequent in patients with initial biopsies with IHC 2+ than in those with IHC 3+ (77.8% vs 54.2%). This observational study demonstrated a considerable rate of HER2 loss after acquired resistance to trastuzumab. Our data raise some important clinical issues. First, we emphasise that a dynamic change in HER2 status may be induced by trastuzumab-based chemotherapy, although re-biopsy after progression is not routinely performed in clinical practice. Importantly, as shown in Table 2, no clinicopathological factors including regimen of chemotherapy, duration of therapy and response to therapy were associated with susceptibility to HER2 loss. This indicates that clinicians should re-assess the HER2 status during administration of the first-line trastuzumab-based chemotherapy to accurately determine whether trastuzumab treatment should continue. Unnecessary trastuzumab treatment should be avoided to minimise the development of trastuzumab-induced adverse events and to reduce unneeded costs [19]. Next, knowledge of HER2 loss in the setting of acquired resistance to trastuzumab might help to increase the likelihood of positive results from a second-line clinical trial. In fact, in both the TyTAN and GATSBY trials, the re-assessment of HER2 status after the development of resistance was not mandatory [8], [9]. However, re-biopsies are often clinically difficult, especially in cases with postoperative recurrence. In the present study, all cases analysed were advanced gastric cancer, not recurrent cancer after gastrectomy. Liquid biopsy may be a helpful tool to predict the efficacy of anti-HER2 therapy by providing a comprehensive description of acquired resistance to trastuzumab [20].