Circulating proteins in response to combined-modality therapy in rectal cancer identified by antibody array screening

Abstract

Background: The increasingly complex programs of contemporary cancer therapy emphasize the need for biological indicators of both therapeutic response and adverse effects. One example is combined-modality treatment aimed at improving long-term outcome in patients with locally advanced rectal cancer, which commonly comes at the price of extended limits of patient tolerance.

Methods: In a prospective study with intensified neoadjuvant treatment of rectal cancer patients, using an antibody array, the profiling of approximately 500 proteins was performed in serial serum samples collected at different stages of the treatment course.

Results: The small number of proteins whose levels significantly changed after induction neoadjuvant chemotherapy (NACT) expanded substantially following the sequential chemoradiotherapy (CRT) and persisted four weeks later at treatment evaluation before pelvic surgery. Serum levels of proteins selected for validation of the experimental design, lipocalin-2 and matrix metalloproteinase-9, declined after NACT and gradually reverted to baseline values during the remaining neoadjuvant course. Of note, the greater the decline in post-NACT and post-CRT matrix metalloproteinase-9 levels, the more favorable progression-free survival. No correlation was found, however, with diarrhea scores, the clinical correlate of adverse therapeutic effects.

Conclusions: Even though the findings were indicative of only tumor and not normal tissue effects, multiplex immunoassay analysis of circulating proteins in patients undergoing combined-modality therapy may in principle dissect the contribution of the individual modalities to overall systemic responses in patient outcome and tolerance.

Trial registration: ClinicalTrials.gov NCT00278694 ; registration date: January 16, 2006, retrospective to enrollment of the first 10 patients of the current report.

Keywords: Chemotherapy; Outcome; Protein array; Radiotherapy; Rectal cancer; Serum proteins.

Figures

Fig. 1

The timing of blood sampling (red arrows) within the treatment protocol. Black arrows indicate the start of each cycle of induction neoadjuvant chemotherapy (NACT) and of each consecutive week of the sequential chemoradiotherapy (CRT). A study-specific evaluation was undertaken before surgery, which was accomplished when the patient had recovered from the neoadjuvant therapy (commonly 2–4 weeks after evaluation)

Fig. 2

Functional coupling between proteins that changed in patients’ circulation during neoadjuvant therapy. Proteins are depicted by their gene symbols. Yellow nodes: proteins whose serum levels significantly differed from baseline following induction neoadjuvant chemotherapy (post-NACT) and sequential chemoradiotherapy (post-CRT) and at treatment evaluation. Non-yellow nodes: proteins not present in the query list but predicted as interacting with the significantly altered proteins at the specific sampling point. Encircled nodes: proteins further analyzed

Fig. 3

Correlations between array fluorescence (FL) intensities and single-parameter immunoassay measurements. Values (transformed to natural logarithms) of lipocalin-2 (LCN2) and matrix metalloproteinase-9 (MMP9) levels in serum samples obtained at baseline, following induction neoadjuvant chemotherapy (post-NACT) and sequential chemoradiotherapy (post-CRT), and at treatment evaluation from 24 randomly chosen patients were compared

Fig. 4

Serum lipocalin-2 (LCN2) and matrix metalloproteinase-9 (MMP9) levels during neoadjuvant therapy. Array fluorescence intensities relative to the individual patient’s baseline values following induction neoadjuvant chemotherapy (post-NACT; n = 50 for LCN2 and n = 61 for MMP9) and sequential chemoradiotherapy (post-CRT; n = 48 for LCN2 and n = 57 for MMP9) and at treatment evaluation (n =50 for LCN2 and n =54 for MMP9); lines, median group values; *p < 0.01; **p < 0.001; ***p < 0.0001