Radiofrequency Thermal Ablation of Hepatocellular Carcinoma Liver Nodules Can Activate and Enhance Tumor-Specific T-Cell Responses Alessandro Zerbini,1 Massimo Pilli,1 Amalia Penna,1 Guido Pelosi,1 Claudia Schianchi,1 Atim Molinari,1 Simona Schivazappa,1 Carlo Zibera,2 Francesco F. Fagnoni,2 Carlo Ferrari,1 and Gabriele Missale1 1Unit of Infectious Diseases and Hepatology, Azienda Ospedaliero- Universitaria di Parma, Parma, Italy and 2Laboratory of Experimental Oncology, Scientific Institute of Pavia, Fondazione Salvatore Maugeri- Clinica del Lavoro e della Riabilitazione, IRCCS, Pavia, Italy

Relevance 1.Is the objective of the article comparing therapeutic interventions similar to your clinical dilemma? Yes, the aim of the study was to find out if radiofrequency thermal ablation can prevent recurrences in HCC

Validity Guides for treatment: 1. Was the assignment of patients to treatment randomized? Was randomization concealed? No, there was no randomization of patients to control and treatment group. The comparison was made between the T-cell response of the patient before and after radiofrequency ablation.

Validity Guides for treatment: 2.Was the follow-up sufficiently long and complete? Follow up was done 6 months after therapy which may not be enough.

Validity Guides for treatment: 3.Was the data analyzed on an intention-to- treat basis? Yes 4. Was there adequate blinding of subjects and researchers? No

Validity Guides for treatment: 5.Were there similar baseline characteristics in each group? All patients were treated for HCC in cirrhosis Other baseline characteristics were not uniform in all patients

Validity Guides for treatment: 6. What is the magnitude of treatment effect? 1 months after radioablation, T-cell response was measured Final outcome measured was recurrence of the tumor Over all is the study valid? No

External validity 1.Are the results of the study applicable to my patient? Yes 2.Were all clinically important outcomes considered? Yes 3.Is treatment feasible? Yes