Repeated-sprint training (RST) is a common training method for enhancing physical fitness in athletes. To advance RST prescription, it is important to understand the effects of programming variables on physical fitness and physiological adaptation. This study (1) quantifies the pooled effects of running RST on changes in 10 and 20m sprint time, maximal oxygen consumption (VO2max), Yo-Yo Intermittent Recovery Test Level 1 (YYIR1) distance, repeated-sprint ability (RSA), countermovement jump (CMJ) height and change of direction (COD) ability in athletes, and (2) examines the moderating effects of program duration, training frequency, weekly volume, sprint modality, repetition distance, number of repetitions per set and number of sets per session on changes in these outcome measures. Pubmed, SPORTDiscus and Scopus databases were searched for original research articles up to 04 July 2023, investigating RST in healthy, able-bodied athletes, between 14 and 35years of age, and a performance calibre of trained or above. RST interventions were limited to repeated, maximal running (land-based) sprints of ≤ 10s duration, with ≤ 60s recovery, performed for 2-12weeks. A Downs and Black checklist was used to assess the methodological quality of the included studies. Eligible data were analysed using multi-level mixed-effects meta-analysis, with standardised mean changes determined for all outcomes. Standardised effects [Hedges G (G)] were evaluated based on coverage of their confidence (compatibility) intervals (CI) using a strength and conditioning specific reference value of G = 0.25 to declare an improvement (i.e. G > 0.25) or impairment (i.e. G < - 0.25) in outcome measures. Applying the same analysis, the effects of programming variables were then evaluated against a reference RST program, consisting of three sets of 6 × 30m straight-line sprints performed twice per week for 6weeks (1200m weekly volume). 40 publications were included in our investigation, with data from 48 RST groups (541 athletes) and 19 active control groups (213 athletes). Across all studies, the effects of RST were compatible with improvements in VO2max (G 0.56, 90% CI 0.32-0.80), YYIR1 distance (G 0.61, 90% CI 0.43-0.79), RSA decrement (G - 0.61, 90% CI - 0.85 to - 0.37), linear sprint times (10m: G - 0.35, 90% CI - 0.48 to - 0.22; 20m: G - 0.48, 90% CI - 0.69 to - 0.27), RSA average time (G - 0.34, 90% CI - 0.49 to - 0.18), CMJ height (G 0.26, 90% CI 0.13-0.39) and COD ability (G - 0.32, 90% CI - 0.52 to - 0.12). Compared with the reference RST program, the effects of manipulating training frequency (+ 1 session per week), program duration (+ 1 extra training week), RST volume (+ 200m per week), number of reps (+ 2 per set), number of sets per session (+ 1 set) or rep distance (+ 10m per rep) were either non-substantial or comparable with an impairment in at least one outcome measure per programming variable. Running-based RST improves speed, intermittent running performance, VO2max, RSA, COD ability and CMJ height in trained athletes. Performing three sets of 6 × 30m sprints, twice per week for 6weeks is effective for enhancing physical fitness and physiological adaptation. Additionally, since our findings do not provide conclusive support for the manipulation of RST variables, further work is needed to better understand how programming factors can be manipulated to augment training-induced adaptations. Open Science Framework registration https://doi.org/10.17605/OSF.IO/RVNDW .