Figure 3.

Antibody response against extracellular domain (ECD) and transmembrane domain (TMD) of HER2 after vaccination using peptide array (from left to right; patients 6, 9, 11, 12, 16, 17, 18, 21, 26, 27, and 29 with the two best responders—patients 9 and 17 marked with light orange). Sequential serum samples from the patients (baseline, white; weeks 8–12, green; weeks 20–24, magenta; weeks 48–52, blue; >52 weeks, dark orange) and IgG (control) were used to display the intensity distribution of the binding to the peptide on the microarray. The peptide sequences of HER2 ECD and TMD are listed on the left column of the heatmap. HER2 ECD immunogenic domains are marked on the right side of the heatmap: I (residues 42–186; white bar), II (residues 236–363; gray bar), III (residues 324–530; black bar), and IV (residues 531–626; blue stripes). Of note, trastuzumab binds to domain IV and pertuzumab binds to domain II, respectively. Patient samples that showed ≥2.5-fold response over baseline; the samples were marked with blue-lined squares in the rows of corresponding peptides. Patient 17 showed polyclonal responses to numerous peptides throughout HER2 ECD immunogenic domains. Patient 9 showed a ≥2.5-fold increase compared with the baseline reaction against HER2 peptides 11 and 137. The two best responders on this study shared many peptides they had reactions to at baseline or after the vaccination, and many of those appeared to be unique to these two patients only (marked with green dotted lines; peptides 14, 35, 63, 116, 117, 131, 132, and 148). Patient 29 also showed a ≥2.5-fold increase compared with baseline against the peptides in domain IV as patient 9 did, but the responses were not accompanied by the corresponding clinical responses. Nearly all patient samples from all timepoints showed reactions to multiple peptides (marked with black solid lines; peptides 22, 83, 84, 85, and 122) across the previously described immunogenic domains.