The most efficient approach for
therapy selection to inhibit the deregulated
kinases in
cancer tissues is to measure their phosphorylation status prior to the treatment. The aim of our study was to evaluate the influence of pre-analytical parameters (cold ischemia time, temperature before and during tissue fixation, and sample type) on the levels of
proteins and
phosphoproteins in
breast cancer tissues, focusing on the
PI3 kinase/AKT pathway. The BALB-neuT mouse
breast cancer model expressing HER2 and pAKT
proteins and human biopsy and resection specimens were analyzed. By using quantitative reverse phase
protein arrays (RPPA), 9
proteins and 16
phosphoproteins relevant to
breast cancer biology were assessed. Cold temperatures before and during fixation resulted in a marked improvement in the preservation of the reactivity of
biological markers (eg, ER, HER2) in general and, specifically, pHER2 and pAKT. Some
phosphoproteins, eg, pHER2 and pAKT, were more sensitive to prolonged cold ischemia times than others (eg, pS6RP and pSTAT5). By comparing the
phosphoprotein levels in core needle biopsies with those in resection specimens, we found a marked decrease in many
phosphoproteins in the latter. Cold conditions can improve the preservation of
proteins and
phosphoproteins in
breast cancer tissues. Biopsies ≤ 1 mm in size are the preferred sample type for assessing the activity of deregulated
kinases for personalized
cancer treatments because the
phosphoprotein levels are better preserved compared with resection specimens. Each potential new (phospho)
protein biomarker should be tested for its sensitivity to pre-analytical processing prior to the development of a diagnostic assay.