Vaccination with Dendritic Cells Loaded with Autologous Leukemic Cells in Combination with Low-Dose Lenalidomide Induced Immune Responses in Chronic Lymphocytic Leukemia (CLL) Patients
Authors: Hansson Lotta , Palma Marzia , Adamson Lars , Ryblom Harriet , Näsman-Glaser Barbro , Eriksson Ingrid , Heimersson Kia , Gentilcore Giusy , Mellstedt Håkan , Österborg Anders 
Published: October 2019

Abstract
Background and aim. We have previously demonstrated that immunization with ex-vivo generated autologous dendritic cells loaded with apoptotic tumor cells (Apo-DC) induces specific immune responses in CLL patients especially when combined with GM-CSF and low-dose cyclophosphamide (CTX) (Palma et al, CII 2012). In this study we evaluate the safety and immunogenicity of Apo-DC vaccination in combination with low-dose lenalidomide alone, or in combination with GM-CSF and low-dose CTX in CLL patients. In patients with multiple myeloma, lenalidomide boosted the response to a pneumococcal vaccine (Noonan et al, Clin Ca Res 2012); lenalidomide may thus be a useful adjuvant also with tumor vaccines but has not yet been explored in man. In patients with CLL, lenalidomide has immunomodulatory properties including NK and T cell stimulation, as well as enhanced immunoglobulin production. Lenalidomide also induced a clinical "flare" reaction, upregulated adhesion molecules and facilitated synapse formation between CLL and T cells (Ramsay et al, J Clin Invest 2008; Shanafelt et al, Blood 2013)i.e. effects of advantage in tumor vaccination. We here report on the first results using lenalidomide as an adjuvant in tumor vaccination in man.
Methods. Ten patients with slowly progressive but asymptomatic CLL were included. The first five patients were immunized intradermally five times during fourteen weeks with a mean of 16x106 Apo-DCs. Low-dose lenalidomide was given daily as an adjuvant at a dose of 2.5 mg from start of vaccination, for four weeks, and then escalated to 5 mg daily until week 24 (cohort I). The next five patients were treated in the same way but also received 300 mg/m2 CTX at day -3 and GM-CSF sc day 1-4 (cohort II). Clinical and immune effects of the vaccination were evaluated at regular time intervals for 1 year. A vaccine-induced immune response was defined as a ≥ 2-fold increase compared to pre-immunization values in either 3H-thymidine incorporation (proliferation) assay or ELISpot assay as described (Palma et al, CII 2012). Changes in the numbers of lymphocyte subpopulations including regulatory T cells (Tregs) as well as in T-cells expressing activation (CD69, CD137) and regulatory markers (CD103) were also evaluated.
Results. To date, the 8 first included patients have completed treatment as planned and passed study week 52. The remaining 2 patients are between study week 24 and 52. No adverse events (AE) > grade 2 have been observed with the exception of one patient who had grade 3 hemolysis (AIHA) at study week 6 (cohort I) and one patient who had grade 4 thrombocytopenia at study week 5 (cohort II). AIHA was treated with steroids and lenalidomide was temporarily withdrawn for 4 weeks. The grade 4 thrombocytopenia required a 10 weeks withdraw of lenalidomide, upon which the thrombocytopenia resolved to grade 1. A decrease in the lymphocyte count fulfilling the criteria for partial response (clinical PR) was seen in 5 patients during lenalidomide treatment (one in cohort I and four in cohort II), but the lymphocyte count increased again to pre-vaccination levels at withdrawal of lenalidomide at week 24. Vaccine-induced immune responses were noted in 4/5 patients in cohort I and in 2/5 patients in cohort II. An inverse correlation between Tregs numbers and proliferation assay values was observed (r = -0.46, p=0.0015) i.e. high proliferative response was associated with low Tregs count. Changes in T cells expressing activation markers (CD69 and CD137) as well as the regulatory marker CD103 were also observed.
Summary/conclusions. This is the first study on lenalidomide as an immunomodulatory agent in tumor vaccination. The results indicate that immunization of CLL patients with autologous tumor-loaded DCs combined with low-dose lenalidomide induced specific immune responses in CLL patients. Lenalidomide as a low-dose immune adjuvance had acceptable tolerability. This therapeutic approach should be explored further to define an optimal combination of vaccination schedule, lenalidomide dose and combination with other immunomodulatory agents interfering with the tumor-microenvironment with the aim to induce a potent immune response with a clinical impact.

Disclosures
Off Label Use: Lenalidomide is used as an adjuvant in this CLL vaccination study. Österborg:Celgene: DMC memeber of Celgene-initiated phase 3 trials Other.