Application of High-Content Analysis in Clinical Cytology for Translational Safety Biomarkers of Drug-Induced Toxicity for Lymphoma Chemotherapy

Drug cytotoxicity detected by high-content analysis (HCA) of live, cultured, human hepatocytes concurs with human toxic potential. We assessed HCA effectiveness in vivo using human, lymphoma line (HuT-78) and blood cells from healthy dogs and lymphomatous dogs treated with anticancer drugs: cytarabine, arsenic trioxide (AT), doxorubicin and mitoxantrone. DNA was stained with Hoechst-33342, calcium with fluo-4, mitochondria with TMRM and cell permeability with toto-3. HuT-78 and canine blood lymphocytes were treated 24hr with 10 times increasing drug concentrations to determine concentration-response relationships to test agents. These showed hormesis (biphasic response) for TMRM and nuclear area. Fluorescence intensity was decreased with AT and mitoantrone but increased with cytarabine for TMRM, decreased from cell injury or anthracycline interaction for Hoechst-33342 and decreased with AT and interfered with by doxorubicin for fluo-4. Nuclear area increased with anthracyclines and AT. Mitochondria, calcium and nuclear area were affected in peripheral blood lymphocytes of lymphomatous dogs receiving chemotherapy: mitochondria were inhibited by acute treatment, but increased 2weeks after treatment, as were ionized calcium, nuclear area and DNA. We conclude (i) concentration-response relationships are determinable in vitro for human and canine lymphocytes by HCA, (ii) cytotoxicity is detectable in vivo during and after anticancer drug treatment using canine blood lymphocytes: mitochondria are most affected, with smaller changes in calcium and nuclei and (iii) blood cell viability may be a biomarker for anticancer drug toxicity.