AUTHOR=Scatena Cristian , Roncella Manuela , Di Paolo Antonello , Aretini Paolo , Menicagli Michele , Fanelli Giovanni , Marini Carolina , Mazzanti Chiara Maria , Ghilli Matteo , Sotgia Federica , Lisanti Michael P. , Naccarato Antonio Giuseppe 

TITLE=Doxycycline, an Inhibitor of Mitochondrial Biogenesis, Effectively Reduces Cancer Stem Cells (CSCs) in Early Breast Cancer Patients: A Clinical Pilot Study

JOURNAL=Frontiers in Oncology

VOLUME=8

YEAR=2018

URL=https://www.frontiersin.org/journals/oncology/articles/10.3389/fonc.2018.00452

DOI=10.3389/fonc.2018.00452

ISSN=2234-943X

ABSTRACT=<p><bold>Background and objectives:</bold> Cancer stem cells (CSCs) have been implicated in tumor initiation, recurrence, metastatic spread and poor survival in multiple tumor types, breast cancers included. CSCs selectively overexpress key mitochondrial-related proteins and inhibition of mitochondrial function may represent a new potential approach for the eradication of CSCs. Because mitochondria evolved from bacteria, many classes of FDA-approved antibiotics, including doxycycline, actually target mitochondria. Our clinical pilot study aimed to determine whether short-term pre-operative treatment with oral doxycycline results in reduction of CSCs in early breast cancer patients.</p><p><bold>Methods:</bold> Doxycycline was administered orally for 14 days before surgery for a daily dose of 200 mg. Immuno-histochemical analysis of formalin-fixed paraffin-embedded (FFPE) samples from 15 patients, of which 9 were treated with doxycycline and 6 were controls (no treatment), was performed with known biomarkers of “stemness” (CD44, ALDH1), mitochondria (TOMM20), cell proliferation (Ki67, p27), apoptosis (cleaved caspase-3), and neo-angiogenesis (CD31). For each patient, the analysis was performed both on pre-operative specimens (core-biopsies) and surgical specimens. Changes from baseline to post-treatment were assessed with MedCalc 12 (unpaired <italic>t-</italic>test) and ANOVA.</p><p><bold>Results:</bold> Post-doxycycline tumor samples demonstrated a statistically significant decrease in the stemness marker CD44 (<italic>p</italic>-value &lt; 0.005), when compared to pre-doxycycline tumor samples. More specifically, CD44 levels were reduced between 17.65 and 66.67%, in 8 out of 9 patients treated with doxycycline. In contrast, only one patient showed a rise in CD44, by 15%. Overall, this represents a positive response rate of nearly 90%. Similar results were also obtained with ALDH1, another marker of stemness. In contrast, markers of mitochondria, proliferation, apoptosis, and neo-angiogenesis, were all similar between the two groups.</p><p><bold>Conclusions:</bold> Quantitative decreases in CD44 and ALDH1 expression are consistent with pre-clinical experiments and suggest that doxycycline can selectively eradicate CSCs in breast cancer patients <italic>in vivo</italic>. Future studies (with larger numbers of patients) will be conducted to validate these promising pilot studies.</p>