Self-assembly formed by a short DNA probe pair: Application for highly sensitive mRNA species detection without reverse transcription
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Authors: Usui, Mitsugu; Fujikawa, Toshihiko; Osawa, Masako; Hakii, Chikako; Ikumi, Natsumi; Nozaki, Takamasa; Kitamura, Noboru; Hatta, Yoshihiro; Fujiwara, Shigeyoshi; Takei, Masami
Abstract
We describe a novel technology for detecting nucleic acids: Probe Alteration Link Self-Assembly Reactions (PALSAR). PALSAR comprises DNA self-assembly of pairs of short DNA probes formed by alternate hybridization of three complementary regions in a pair of honeycomb probes (HCPs). Self-assembly occurs at designated salt concentrations and reaction temperatures and requires no enzymes. We prepared pairs of HCPs to detect mRNAs encoded by the GAPDH gene beta-actin (BA) gene, CD3D gene, CD4 gene, major vault protein (MV) gene and the signalling lymphocytic activation molecule-associated protein (SAP) gene, and succeeded in quantitatively detecting these mRNAs. PALSAR could detect mRNA directly without synthesizing cDNA. Moreover, multiple mRNAs could be detected simultaneously in a single reaction tube and there was a good correlation between the results obtained PALSAR and those by real-time PCR. (C) 2015 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Tumor-Associated Lymphocytes As an Independent Predictor of Response to Neoadjuvant Chemotherapy in Breast Cancer
JOURNAL OF CLINICAL ONCOLOGY
Authors: Denkert, Carsten; Loibl, Sibylle; Noske, Aurelia; Roller, Marc; Mueller, Berit Maria; Komor, Martina; Budczies, Jan; Darb-Esfahani, Silvia; Kronenwett, Ralf; Hanusch, Claus; von Toerne, Christian; Weichert, Wilko; Engels, Knut; Solbach, Christine; Schrader, Iris; Dietel, Manfred; von Minckwitz, Gunter
Abstract
Purpose Preclinical data suggest a contribution of the immune system to chemotherapy response. In this study, we investigated the prespecified hypothesis that the presence of a lymphocytic infiltrate in cancer tissue predicts the response to neoadjuvant chemotherapy. Methods We investigated intratumoral and stromal lymphocytes in a total of 1,058 pretherapeutic breast cancer core biopsies from two neoadjuvant anthracycline/taxane-based studies (GeparDuo, n = 218, training cohort; and GeparTrio, n = 840, validation cohort). Molecular parameters of lymphocyte recruitment and activation were evaluated by kinetic polymerase chain reaction in 134 formalin-fixed, paraffin-embedded tumor samples. Results In a multivariate regression analysis including all known predictive clinicopathologic factors, the percentage of intratumoral lymphocytes was a significant independent parameter for pathologic complete response (pCR) in both cohorts (training cohort: P = .012; validation cohort: P = .001). Lymphocyte-predominant breast cancer responded, with pCR rates of 42% (training cohort) and 40% (validation cohort). In contrast, those tumors without any infiltrating lymphocytes had pCR rates of 3% (training cohort) and 7% (validation cohort). The expression of inflammatory marker genes and proteins was linked to the histopathologic infiltrate, and logistic regression showed a significant association of the T-cell-related markers CD3D and CXCL9 with pCR. Conclusion The presence of tumor-associated lymphocytes in breast cancer is a new independent predictor of response to anthracycline/taxane neoadjuvant chemotherapy and provides useful information for oncologists to identify a subgroup of patients with a high benefit from this type of chemotherapy.