INTRODUCTION Currently available prognostic factors determining the course of chronic lymphocytic leukemia (CLL) are not fully efficient, especially for newly diagnosed patients. Investigation of molecular changes may help clarify the reasons for the heterogeneity of the disease. Apart from already confirmed TP53 mutations, the novel candidates: NOTCH1, SF3B1, and MYD88 might represent clinically relevant biomarkers. OBJECTIVES The aim of this study was to evaluate the mutational status of NOTCH1, MYD88, and SF3B1 and to compare the results with confirmed prognostic factors: ZAP‑70, CD38, and immunoglobulin heavy‑chain variable region (IGHV) mutation in CLL. The study assessed also prognostic significance in terms of the time to first treatment (TTFT) and subset analysis. PATIENTS AND METHODS The study was conducted on samples of 370 newly diagnosed patients with CLL. The analysis was performed using high‑resolution melting, Sanger sequencing, and polymerase chain reaction methods. RESULTS Patients harboring the NOTCH1 mutation were significantly more often found among patients with an unmutated IGHV gene status and high expression of CD38 and ZAP‑70. The MYD88 mutation was equally distributed in patients with mutated and unmutated IGHV status (5 vs 7 patients). For MYD88 and SF3B1, there were no significant differences in the levels of CD38 and ZAP‑70 expression. The tendency for lower median TTFT was revealed in patients with mutated SF3B1 (P = 0.08). The analysis showed the presence of 14 different types of the subsets of IGHV in 50 of 345 patients (14.5%). The most frequent were subsets #1 and #2. CONCLUSIONS The NOTCH1 and SF3B1 mutations accompany biological markers of unfavorable prognosis in patients with CLL. The mutations may contribute to the identification of patients with high‑risk CLL.