Capecitabine

Fresh-frozen tumor specimens from 293 consecutive CRC patients were retrieved from the tissue banks of the Royal Melbourne Hospital, Western Hospital and Peter MacCallum Cancer Center in Australia, and the H. Lee Moffitt Cancer Center in the United States; individuals who had received preoperative chemo- and/or radiotherapy or for whom tumor-derived total RNA was inadequate for microarray analysis (RIN < 6) were excluded. All patients gave informed consent, and this study was approved by the medical ethics committees of all sites. Patient median age at diagnosis was 67 years (range 26 to 92 years). All specimens were derived from primary carcinomas and were snap-frozen in liquid nitrogen immediately after surgery for storage at −80°C. Cases comprised 44 stage A, 95 stage B, 93 stage C and 61 stage D cancers; 252 were localized to the colon and 40 to the rectum, with one case missing this information. 22 of 94 patients who had stage B disease and 64 of 91 patients who had stage C disease had received standard adjuvant chemotherapy (either single agent 5-fluouracil/capecitabine or 5-fluouracil and oxaliplatin) or postoperative concurrent chemoradiotherapy (50.4 Gy in 28 fractions with concurrent 5-fluorouracil) according to hospital protocols. All patients were assessed annually. For stage B and C patients, follow-up and additional clinical data including patient gender and TNM staging were collected by Biogrid Australia 1 for Australian patients and the Moffitt Cancer Center Tumor Registry for US patients. The median duration of follow-up was 47.8 months (range 0.9 to 118.6 months) for the 140 patients without recurrence, and 19.1 months (range 1.6 to 93.7 months) for the 48 patients with local or distant recurrence. The median follow-up for all 188 patients was 37.2 months (range 0.9 to 118.6 months).
Total RNA was extracted using Trizol reagent (Invitrogen) from CRC samples containing >60% tumor cells. All samples included showed good integrity of 18S and 28S ribosomal bands (RIN > 6) using a 2100 Bioanalyzer (Agilent Technologies). Total RNA was labeled and hybridized to HG-U133Plus2.0 GeneChip arrays (Affymetrix) according to the manufacturer’s instructions. The microarray data on a subset of 174 tumors have been published previously (NCBI Gene Expression Omnibus, GSE5206 and GSE13067).
In addition, published gene expression data were retrieved for 42 stage A CRCs, 83 stage B, 73 stage C and 62 stage D CRCs analyzed as part of the Expression Project for Oncology (expO) 2 using HG-U133Plus2.0 GeneChip arrays (Affymetrix) (Supplementary Table S1). Of the 62 stage D CRCs, 32 were primary cancer and 30 were metastectomy specimens. None of the primary cancer patients had received preoperative therapy, but 17 metastectomy specimens were from patients who had received adjuvant chemotherapy treatment prior to resection. Data processing and analysis were performed using the statistical software package R (15 ) and appropriate Bioconductor packages (16 (link)).

The EMILIA study is a randomized, open-label, international trial involving patients with HER2-positive, unresectable, locally advanced or metastatic breast cancer who were previously treated with trastuzumab and a taxane. The study was conducted in accordance with the International Conference on Harmonization Good Clinical Practice standards and the Declaration of Helsinki. Patients provided written informed consent; the study was approved by the relevant institutional review board or independent ethics committee.
Patients were randomly assigned in a 1:1 ratio to T-DM1 or lapatinib plus capecitabine with the use of a hierarchical, dynamic randomization scheme through an interactive voice-response system. Stratification factors were world region (United States, Western Europe, or other), the number of prior chemotherapy regimens for unresectable, locally advanced or metastatic disease (0 or 1 vs. >1), and disease involvement (visceral vs. nonvisceral).
The primary end points were progression-free survival assessed by independent review, overall survival, and safety. Progression-free survival was defined as the time from randomization to progression or death from any cause. Progression was assessed according to modified Response Evaluation Criteria in Solid Tumors (RECIST), version 1.0^{12 (link)}; the modified criteria are specified in the Supplementary Appendix, available with the full text of this article at NEJM.org. Overall survival was defined as the time from randomization to death from any cause. Prespecified secondary end points included progression-free survival (investigator-assessed), the objective response rate, the duration of response, and the time to symptom progression. The objective response rate was determined according to modified RECIST on the basis of an independent review of patients with measurable disease at baseline; responses were confirmed at least 28 days after the initial documentation of a response. The time to symptom progression was defined as the time from randomization to the first decrease of 5 points or more from baseline scores on the Trial Outcome Index of the patient-reported Functional Assessment of Cancer Therapy–Breast (FACT-B TOI, on which scores range from 0 to 92, with higher scores indicating a better quality of life)^{13 (link)} in women with a baseline score and at least one postbaseline score. Safety was monitored by an independent data monitoring committee and a cardiac review committee.

We conducted a randomized, two-group, open-label, phase 3 trial involving patients with HER2-low, unresectable or metastatic breast cancer. Trial enrollment was planned for 480 patients with hormone receptor–positive disease (immunoreactive for estrogen or progesterone receptor in ≥1% of tumor-cell nuclei according to local testing) and 60 patients with hormone receptor–negative disease, approximating the proportions of receptor subtype observed in HER2-low breast cancer.^{1 (link)} Patients were randomly assigned in a 2:1 ratio to receive trastuzumab deruxtecan or the physician’s choice of capecitabine, eribulin, gemcitabine, paclitaxel, or nab-paclitaxel. Randomization was stratified according to HER2-low status (IHC1+ vs. IHC 2+ and ISH-negative), the number of previous lines of chemotherapy for metastatic disease (one vs. two), and hormone-receptor status (positive [with vs. without previous CDK4/6 inhibitor therapy] vs. negative).
IHC scores for HER2 expression were determined through central testing of adequate archived or recent tumor-biopsy specimens with the use of an investigational IHC assay, the VENTANA HER2/neu (4B5) IUO (investigational use only) Assay system, according to an algorithm adapted from the 2018 American Society of Clinical Oncology/College of American Pathologists testing guidelines (Table S1 in the Supplementary Appendix, available with the full text of this article at NEJM.org).^{22 (link)} Specimens that yielded central HER2 IHC scores of 2+ were reflexed to ISH testing with the use of the investigational INFORM HER2 Dual ISH DNA Probe Cocktail IUO Assay system.
Eligible patients must have received chemotherapy for metastatic disease or have had disease recurrence during or within 6 months after completing adjuvant chemotherapy; patients with hormone receptor–positive disease must have received at least one line of endocrine therapy. Patients with treated, stable brain metastases were eligible; patients were ineligible if they had a history of noninfectious interstitial lung disease requiring treatment with glucocorticoids or had suspected interstitial lung disease on imaging at screening.
Trastuzumab deruxtecan was administered intravenously every 3 weeks at a dose of 5.4 mg per kilogram of body weight, and the physician’s choice of chemotherapy was administered in accordance with local label or the National Comprehensive Cancer Network guidelines.⁶ More details are provided in the Supplementary Appendix.