Dynamics Changes in CCL3 and CCL4 Plasma Chemokine Levels in Patients with Chronic Lymhocytic Leukemia (CLL) Managed with Observation

Background: CCL3 and CCL4, previously called macrophage inflammatory protein-1 alpha (MIP-1α) and MIP-1β, are chemokines of the CC subfamily, which are secreted by CLL cells in response to B cell receptor (BCR) stimulation and by co-culture with nurse-like cells (NLC), presumably to attract T lymphocytes and monocytes that can deliver pro-survival signals. We previously reported, that CLL patients have elevated plasma levels of CCL3 and CCL4, which rapidly normalize after treatment with kinase inhibitors that target BCR signaling, such as the BTK inhibitor ibrutinib and the PI3K inhibitor idelalisib (Blood 119:1182-9, 2012)(Blood 118:3603-12, 2011). Moreover, in untreated CLL patients, high CCL3 levels are an independent prognostic marker for short time to disease progression (Blood 117:1662-9, 2011), emphasizing the clinical relevance of these chemokines in CLL.

Aim: To better understand the dynamic changes, we analyzed plasma levels of CCL3 and CCL4 in a large cohort of serial samples from untreated CLL patients. Additionally, we analyzed separately from the entire cohort, CCL3 and CCL4 changes overtime in patients who subsequently received therapy and patients who were still on observation at time of analysis.

Methods: CCL3 and CCL4 plasma levels were measured by ELISA in serial samples collected over time in untreated CLL patients. A total of 193 individual patients with 603 plasma samples were included in the analysis (entire cohort), of the 193 patients, 81 of them required treatment at a later time, although all the samples analyzed were collected prior any treatment, 112 patients did not need any intervention over the period of sample collection and were managed with observation. The duration between first and subsequent samples ranged between 5 months to 12 years. The duration between the date of the last sample and the date of the initiation of treatment ranged between less than a month to 88 months with a median of 6 months. Mixed linear models with repeated measure were used to determine whether there was a difference in mean levels of CCL3 and CCL4 over time.

Results: The median and range of CCL3 and CCL4 levels for the initial sample over all 193 patients were 11.8 pg/mL (2.5 – 1977.1 pg/mL) and 88.8 pg/mL (6.9 – 2601.3 pg/mL) respectively. There were significant associations of time between first and subsequent serial samples (p<0.0001) and log of baseline CCL3 (p<0.0001) with log of CCL3 measurements over time, in both the watchful waiting and treated patient's cohort; same associations were found for CCL4. Overall, we found that there was an increase by a factor of 1.009/month in CCL3 and 1.006/month for CCL4 in the group of patients managed with observation, and 1.01/month in CCL3 and 1.011/month in CCL4 in the group of patients that were eventually treated.

Conclusions: These studies demonstrated that there is a slight but statistically significant increase in the levels of the chemokines CCL3 and CCL4 over time in CLL patients, regardless of whether the patients required therapy at a later time. We conclude that CCL3 and CCL4 plasma levels presumably reflect the dynamic changes of CLL pathogenesis over time, strengthening the relevance of CCL3 and CCL4 chemokine production in CLL.