Introduction <p>Adrenocorticotropic hormone (ACTH) is a first-line drug and standard therapy in infantile epileptic spasm syndrome (IESS) with different response rates. The mechanism of ACTH for treatment of IESS remains incompletely understood. This study aimed to evaluate short-term outcomes and immunologic mechanisms of ACTH in treating pediatric patients with IESS.</p> Methods <p>A multicenter prospective trial of pediatric patients with IESS treated with ACTH over 2&#xa0;weeks was performed. Short-term responders were defined as those with resolution of epileptic spasms and hypsarrhythmia (if present) at day&#xa0;14 without epileptic spasm recurrence over 30&#xa0;days after ACTH therapy, while non-responders included those with persistent spasms or hypsarrhythmia after treatment, or recurrence within 30&#xa0;days. The risk factors for short-term efficacy were analyzed. The receiver operating characteristic (ROC) curve analysis was adopted to reveal the optimal critical value of peripheral blood lymphocyte subpopulations before ACTH treatment.</p> Results <p>Among 151 included pediatric patients, 44 (29.1%) were short-term responders. The presence of hypsarrhythmia on pre-therapy electroencephalogram (EEG) was a risk factor for the short-term efficacy (<i>p</i> = 0.029). A total of 81 patients accepted lymphocyte subpopulation investigation. The short-term responder group had a lower percentage of CD3⁺CD8⁺ T&#xa0;cells (<i>p</i> = 0.080) and a higher CD3⁺CD4⁺/CD3⁺CD8⁺ ratio (<i>p</i> = 0.077) compared to the non-responder group, but the differences were not statistically significant. ROC analysis yielded area under the curve (AUC) values of 0.599 for CD3⁺CD8⁺ T&#xa0;cell percentage and 0.600 for the CD3⁺CD4⁺/CD3⁺CD8⁺ratio. Patients with a CD3⁺CD8⁺ T&#xa0;cell percentage below 19.495% (<i>p</i> = 0.04) and a CD3⁺CD4⁺/CD3⁺CD8⁺ above 2.585 (<i>p</i> = 0.013) had a significantly higher short-term responder rate to ACTH therapy.</p> Conclusion <p>About 30% children with IESS were short-term responders to ACTH therapy. The presence of hypsarrhythmia on the pre-therapy EEG was a risk factor for short-term efficacy of ACTH administration. The CD3⁺CD8⁺ T&#xa0;cell percentage and CD3⁺CD4⁺/CD3⁺CD8⁺ ratio may exhibit a predictive capacity.</p>