Highlights
– Venetoclax monotherapy rapidly reduces circulating CLL subpopulations but surviving cells upregulate prosurvival BCL-2 family proteins (BCL-2, BCL-XL, MCL-1) and show reduced drug sensitivity.
– Venetoclax treatment (and anti-CD20 therapy) is associated with an early increase in plasma BAFF levels; BAFF signaling is required for the survival-program induction in preclinical models.
– These findings support testing strategies that co-target BAFF signaling or complementary prosurvival pathways (for example, MCL-1) to deepen and prolong responses, and raise the possibility that short-term homeostatic responses during dose ramping may limit long-term efficacy.
Background: clinical context and unmet need
Chronic lymphocytic leukemia (CLL) is characterized by the clonal accumulation of mature B lymphocytes that depend on antiapoptotic signaling for survival. Venetoclax is a selective small-molecule inhibitor of B-cell lymphoma-2 (BCL-2) and has transformed CLL treatment by inducing deep remissions, including undetectable minimal residual disease (MRD) in many patients. Depth of MRD early during therapy is a strong predictor of durable responses and progression-free survival.
Despite transformative efficacy, not all patients achieve deep, durable remissions and acquired resistance remains a clinical challenge. Understanding early, therapy-induced adaptive responses in CLL cells can identify mechanisms that blunt initial depth of response and suggest rational combinations to prevent or overcome resistance.
Study design
Luo and colleagues performed a longitudinal, single-cell proteomic analysis of peripheral blood from patients with CLL treated with venetoclax monotherapy, focusing on the first 5 weeks of therapy during dose escalation. The authors used mass cytometry (CyTOF) to profile proliferative, metabolic, and survival-related proteins at single-cell resolution, resolving intra-patient CLL heterogeneity and dynamics of surviving populations. Complementary murine models and genetic perturbations probed mechanistic requirements, including the role of apoptosis and B-cell activating factor (BAFF). Patient plasma BAFF levels were measured longitudinally and changes were contextualized against treatment with venetoclax or the anti-CD20 antibody obinutuzumab.
Key findings
Baseline heterogeneity and early depletion
At baseline, mass cytometry resolved multiple distinct CLL subpopulations within patients, characterized by differences in proliferation markers, metabolic proteins, and expression of survival factors. Following venetoclax initiation and dose escalation, there was a significant and rapid reduction in the abundance of all identified CLL subpopulations in peripheral blood, consistent with the drug’s potent pro-apoptotic activity in sensitive cells.
Survivors rapidly upregulate prosurvival proteins and show decreased sensitivity
Notably, the small pool of surviving CLL cells exhibited a rapid and marked upregulation of prosurvival BCL-2 family proteins, including BCL-2 itself as well as BCL-XL and MCL-1. Functionally, these surviving cells demonstrated reduced sensitivity to venetoclax ex vivo. The pattern indicates an immediate adaptive survival program in residual cells despite early cytoreduction.
BAFF elevation and requirement for survival-program induction
Across patient samples, plasma BAFF levels rose markedly after venetoclax initiation. Similar BAFF increases and upregulation of prosurvival proteins were seen after treatment with obinutuzumab, suggesting that B-cell targeting and the resulting cell death or perturbation of the B-cell compartment provoke systemic homeostatic cytokine responses. In murine models, authors recapitulated the venetoclax-induced elevation of survival proteins in both normal B cells and CLL-like cells. Genetic experiments showed that extensive apoptosis and access to BAFF were essential for the observed prosurvival program: blocking BAFF signaling blunted the induction of prosurvival proteins in surviving cells.
Mechanistic interpretation
BAFF (B-cell activating factor) binds receptors on B cells (BAFF-R, TACI, BCMA) and activates downstream NF-κB and other survival pathways that increase expression of BCL-2 family members. The data suggest a model in which venetoclax-mediated cell death and disruption of B-cell homeostasis triggers a compensatory increase in circulating BAFF that, through receptor signaling, upregulates multiple antiapoptotic proteins (including MCL-1 and BCL-XL) in surviving cells. This homeostatic response reduces the fractional killing achieved by venetoclax and creates an early reservoir of drug-tolerant cells with higher expression of alternative survival factors.
Clinical translation and implications
The findings offer several clinically relevant insights. First, the commonly used venetoclax dose ramp—implemented to reduce tumor lysis risk—occurs during a period of rapid adaptive survival signaling; whether ramp duration influences the magnitude of BAFF induction or compensatory prosurvival upregulation merits investigation. Second, BAFF elevation was not unique to venetoclax; anti-CD20 therapy produced a similar effect, implying that homeostatic cytokine responses are a general consequence of acute B-cell depletion. Third, upregulation of MCL-1 and BCL-XL identifies these proteins as candidate co-targets to prevent early adaptation and deepen MRD responses.
Safety and secondary observations
The study was focused on mechanistic and cell-signaling endpoints rather than long-term clinical outcomes. Reports of adverse events or safety signals were not the central focus; however, any strategy that targets BAFF signaling or additional prosurvival pathways will need careful assessment for on-target immune effects, such as increased risk of infections or perturbations of normal B-cell homeostasis.
Expert commentary and limitations
These data are biologically plausible and align with established roles for BAFF in B-cell survival and homeostasis. The single-cell proteomic approach provides high-resolution insight into early adaptive responses that are difficult to capture with bulk assays. Demonstration of the phenomenon in mouse models and genetic perturbation strengthens causal inference.
Limitations include the relatively short sampling window (first 5 weeks) and focus on peripheral blood. Lymph node and bone marrow microenvironments are important niches for CLL and may exhibit different kinetics or additional survival cues. The patient cohort size and heterogeneity (e.g., prior therapies, disease stage) may limit generalizability. Clinical impact—whether BAFF-driven adaptation measurably reduces long-term progression-free survival or can be reversed by targeted combination therapy—remains to be tested prospectively in trials.
Therapeutic translation presents challenges: BAFF neutralization (for example, agents used in autoimmune disease such as belimumab) has not been routinely used in CLL and may carry immunologic risks. Direct inhibition of MCL-1 is an active area of drug development; early-phase MCL-1 inhibitors are being tested in hematologic malignancies and may offer a more proximal approach to prevent venetoclax escape. Combination timing, sequencing, and safety will require careful clinical testing.
Practical implications and research priorities
For clinicians and investigators, the study suggests the following priorities:
- Consider measuring BAFF as a potential early pharmacodynamic biomarker in venetoclax studies to explore correlations with MRD depth and relapse kinetics.
- Design clinical trials that test venetoclax combined with agents targeting BAFF signaling (for example, BAFF-neutralizing agents/decoy receptors) or complementary antiapoptotic proteins, particularly MCL-1 inhibitors, with close monitoring for infectious and immunologic adverse events.
- Investigate whether shorter or different venetoclax ramp strategies alter the magnitude of BAFF induction or the emergence of prosurvival signatures, balancing tumor-lysis safety with potential adaptive resistance.
- Include tissue-based correlative studies (lymph node, bone marrow) to define microenvironment contributions and confirm peripheral blood observations in relevant niches.
Conclusion
Luo et al. provide compelling single-cell and preclinical evidence that venetoclax dose escalation rapidly activates a BAFF-driven prosurvival program in CLL cells that survive early therapy. This homeostatic response increases expression of BCL-2 family proteins including MCL-1 and BCL-XL, producing cells with reduced venetoclax sensitivity. The findings identify BAFF signaling and alternative antiapoptotic proteins as rational co-targets to deepen responses and delay resistance, and underscore the value of integrating early pharmacodynamic profiling into trials of targeted therapies.
Funding and clinicaltrials.gov
The primary study cited is supported and reported by Luo MX et al. (Blood 2024). Specific funding sources and trial registrations for patient cohorts should be consulted in the original publication (Luo et al., Blood 2024) and on ClinicalTrials.gov for ongoing combination studies involving venetoclax and agents targeting BAFF or MCL-1.
References
1. Luo MX, Tan T, Trussart M, et al. Venetoclax dose escalation rapidly activates a BAFF/BCL-2 survival axis in chronic lymphocytic leukemia. Blood. 2024 Dec 26;144(26):2748-2761. doi: 10.1182/blood.2024024341. PMID: 39471335; PMCID: PMC11738032.
2. Hallek M, Cheson BD, Catovsky D, et al. iwCLL guidelines for diagnosis, indications for treatment, response assessment, and supportive management of CLL. Blood. 2018;131(25):2745-2760. doi:10.1182/blood-2017-09-806398.
3. Roberts AW, Davids MS, Pagel JM, et al. Targeting BCL2 with venetoclax in relapsed chronic lymphocytic leukemia. N Engl J Med. 2016;374:311-322. doi:10.1056/NEJMoa1513257.
4. Mackay F, Schneider P. Cracking the BAFF code. Nat Rev Immunol. 2009;9(7):491-502. doi:10.1038/nri2552.
Suggested next steps for clinicians and researchers
Clinicians should be aware of early adaptive survival signaling after venetoclax initiation and consider enrolling patients in trials testing rational combinations. Investigators should prioritize correlative biomarker studies of BAFF and BCL-2 family proteins during therapy and pursue preclinical and early-phase clinical trials that evaluate BAFF pathway inhibitors or MCL-1 inhibitors combined with venetoclax, with careful attention to infectious risk and immune consequences.
