Editor's Note: The 2025 European Hematology Association (EHA) Annual Meeting was grandly held both online and in person, gathering top experts and cutting-edge research from the global hematology community. At this conference, a revolutionary study on the origin of Chronic Lymphocytic Leukemia (CLL) attracted widespread attention. A research team from Spain, using multi-omics single-cell sequencing technologies, revealed that CLL does not, as traditionally believed, originate from a single clone. Instead, in some patients, multiple independent minor clones with the biological characteristics of CLL exist, having emerged and begun to evolve decades before the disease is diagnosed. This discovery not only profoundly reshapes our understanding of CLL pathogenesis but also provides a completely new theoretical basis for future early diagnosis and intervention strategies.

The conventional view holds that Chronic Lymphocytic Leukemia (CLL) is a monoclonal disease, where all tumor cells derive from a single founding cell and therefore share the same B-cell Receptor (BCR). However, oligoclonality is observed in approximately 5% to 10% of clinical CLL cases and in an even higher proportion of its precursor condition, Monoclonal B-cell Lymphocytosis (MBL). These clues suggest that the origin of CLL may be more complex than previously thought, possibly involving a polyclonal process. To investigate this question, a young scholar from the laboratory of the renowned Spanish hematopathologist Professor Elias Campo systematically reported their team’s latest research findings at this year’s congress.

Immunogenetic Features: A Pervasive CLL Bias Exists in the Patient’s B-cell Lineage

The research team first performed deep sequencing of immunoglobulin genes on samples from over 550 individuals with CLL, MBL, and healthy blood donors. They found that the incidence of oligoclonality was as high as 60% in low-count MBL, which has low malignant potential, 28% in high-count MBL, and 12% in CLL. Interestingly, in nearly 70% of cases with these co-existing, abnormally expanded clones, the immunoglobulin genes shared the same mutational status (i.e., either all mutated or all unmutated), suggesting a bias at the immunogenetic level.

A more surprising discovery came from the analysis of non-expanded B-cell clones. The team found that within the “background” B-cell repertoire of patients with CLL and MBL, the frequency of CLL-specific stereotyped BCRs was 18 times higher than in healthy individuals. This powerful data reveals that the entire B-cell system of a CLL patient appears to have a “preset” predisposition towards CLL, not just the single clone that eventually becomes dominant. As the scholar emphasized in the presentation: “This bias is not limited to the expanded clones but is pervasive throughout the patient’s entire B-cell lineage.”

Transcriptomic and Genomic Evidence: Minor Clones Already Possess the Complete Biological Imprint of CLL

To investigate whether these minor clones with CLL immunogenetic features also possess the biological essence of CLL, the research team utilized single-cell transcriptomic sequencing for an in-depth analysis of 38 samples. They developed a gene expression signature model capable of distinguishing between mutated CLL, unmutated CLL, and normal B cells.

The results showed that both the dominant, large clones and the extremely low-frequency minor clones (at levels equivalent to very low-count MBL) clearly displayed the CLL transcriptomic signature corresponding to their immunoglobulin mutational status. In one MBL case involving six clones of different sizes, five of the minor clones exhibited CLL features consistent with their respective mutational statuses. Among the 10 CLL patients studied, all clones in 9 patients showed a CLL-biased transcription profile; in 4 of the 6 MBL patients, the minor clones also carried the CLL transcriptomic imprint.

Subsequently, the team further confirmed through single-cell DNA sequencing that these minor clones also harbored CLL driver gene mutations. The study identified CLL driver alterations in the minor clones of all nine analyzed cases, including del(13q), a key early event in CLL, and even some mutations associated with aggressive disease, such as TP53 mutations. Strikingly, in some cases, a minor clone carried a clear driver mutation while the dominant tumor clone at that time had no detectable known driver alterations. This indicates that these minor clones are truly malignant-potential tumor cells, not benign bystanders.

Tracing the Roots: Whole-Genome Sequencing Reveals Independent Origins and Long-Term Evolution of CLL Clones

Do these CLL clones co-existing within the same patient originate from a common ancestor, or did they arise independently? This was the ultimate question the study sought to answer. To this end, the research team employed cutting-edge single-cell whole-genome sequencing. By analyzing the complete set of somatic mutations within single cells, they constructed precise phylogenetic trees for 6 oligoclonal CLL patients.

The results clearly demonstrated that in all analyzed cases, the different clones were located on independent branches of the phylogenetic tree and did not share any somatic mutations with each other. This definitive evidence proves that they originated from entirely different cells. Furthermore, the phylogenetic trees also revealed the phenomenon of “convergent evolution,” where, for example, clones on different branches independently acquired the key driver event of del(13q).

Using genomic mutations as a “molecular clock,” the research team further calculated the evolutionary timeline of these clones. The analysis yielded a stunning conclusion: the first driver mutation in CLL may be acquired within the first 30 years of a patient’s life, and the divergence and expansion of different clones begin silently, decades before the disease is clinically diagnosed.

Summary and Outlook

This study systematically demonstrates that, at least in a subset of CLL patients, the disease is fundamentally a co-expansion of multiple clones possessing the immunogenetic, transcriptomic, and genomic features of CLL. These clones originate from different ancestral cells and undergo a long evolutionary process spanning decades before diagnosis.

This work completely subverts the monoclonal origin theory of CLL, painting a new picture of the disease’s pathogenesis: the “seeds” of CLL may be widely sown in the B-cell system early in life, and through a prolonged period of evolution and selection, they eventually form a clinically apparent disease. This discovery is not only a milestone in understanding the ontogeny of CLL but also points the way toward developing entirely new clinical strategies based on early risk prediction and disease intervention, truly embodying the scientific spirit of exploring disease from its source to lead the future with wisdom.