Early intervention with lenalidomide in patients with high-risk chronic lymphocytic leukemia

Abstract

Purpose: Infectious complications constitute a leading cause of morbidity and mortality in chronic lymphocytic leukemia (CLL). Patients respond poorly to vaccines, particularly pneumococcal polysaccharide and influenza vaccines. Additionally, patients with genetically high risk disease are at increased risk for early disease progression and death. Lenalidomide, an oral immunomodulatory agent with demonstrated clinical activity in CLL, can potentially restore immune system dysfunction associated with CLL while improving disease outcomes. Experimental Design: Phase 2 study randomized 49 patients with genetically high-risk CLL or SLL (defined as unmutated IGHV, deletion(17p) or (11q), and/or complex abnormal karyotype), to receive lenalidomide either concurrent (Arm A) or sequential to (Arm B) 2 doses of 13-valent protein-conjugated pneumococcal vaccine (PCV13) administered 2 months apart, in patients not meeting IWCLL treatment criteria. Results: Four serotypes (3, 4, 5, 6B) achieved the additional seroprotection definition of a 4-fold increase in arm A, and 6 serotypes (3, 4, 5, 6B, 19A, 19F) in arm B. All patients achieved the defined concentration of 0.35ug/ml for at least one serotype tested. No significant difference was observed with the addition of lenalidomide. At median time on treatment of 3.6 years, median PFS was 5.8 years (95% CI 3.1-NR). PFS at 1, 2 and 3 year was 85% (95% CI 72–93), 79% (64–88), and 72% (95% CI 57–83) respectively. Conclusion: Lenalidomide is efficacious with manageable toxicities as an early intervention strategy in patients with high-risk CLL, but did not enhance humoral response to PCV13 vaccine. This trial was registered with ClinicalTrials.gov Identifier: NCT01351896

Introduction

Infectious complications constitute the leading cause of morbidity and mortality in patients with Chronic Lymphocytic Leukemia (CLL).(1–3) This increased risk is attributable to both patient-related factors such as age, and disease-specific factors including hypogammaglobulinemia, impaired cell-mediated immunity, and poor response to preventative vaccines. The immune defect in CLL is characterized by both humoral and cellular deficits. Hypogammaglobulinemia is often present at diagnosis and typically increases in incidence with disease progression and ultimately affects 75% of patients.(4,5) Impaired immunoglobulin (Ig) production significantly increases the risk for bacterial infection, particularly those caused by encapsulated organisms such as Streptococcus pneumoniae. Defects in both cellular and humoral immunity likely account for the poor response to vaccination in CLL patients, and developing strategies to enhance vaccine response represents an important area in CLL research.(5)

Early studies in previously untreated CLL patients who were challenged with antigens to mumps, diphtheria, influenza, and typhoid, demonstrated a correlation between the level of gamma-globulin deficiency, response to antigenic stimulation, and development of subsequent infections.(6) A correlation between immunologic responses to vaccine and absolute numbers of CD4+/CD45RA+ naïve T-cells has also been identified, which may suggest a role of this subset of cells in antibody response.(7) In addition, the use of a booster dose resulted in an increase in the response rate from 5% to 15% for influenza A and from 15% to 30% for influenza B, but this did not have a significant impact on protection rates.(8) This poor response was confirmed in an open-label, randomized study, where the response rates with two doses compared to one was 18% vs 22% for H1N1, 26% vs 14% for H3N2 serotypes, and 25% vs 22% for influenza B.(9) Purified polysaccharide antigens result in T-cell independent type-2 antibody formation. In protein conjugated vaccines, the polysaccharide antigen is conjugated to a toxoid as a carrier protein, resulting in antibody responses to the polysaccharide antigens in a T cell-dependent manner. T cell-dependent antigens induce immunological memory, resulting in the possibility of response augmentation with booster vaccinations. Studies have also indicated that conjugation of polysaccharides may render the antigen more immunogenic in the CLL population, and there is evidence that patients with CLL have a more significant immune response to Haemophilus influenzae b conjugate than to plain polysaccharide antigen.(10) However, age, disease stage, and IgG levels all play a role in the degree of response to immunization.(11) Similarly, administration of a 7-valent conjugated pneumococcal vaccine demonstrated responses in 39% of previously untreated patients, however, responses in more advanced stage of disease were only 5%.(12)

Lenalidomide inhibits proliferation of CLL cells through its targeting of cereblon,(13) and importantly, results in multiple immunomodulatory effects. This includes repair of the immunologic synapse formation between T-cells and CLL B-cells,(14) improvement of NK-cell mediated cytotoxicity,(15) and stimulation of the production of Igs by B-cells.(16–18) Clinical use of lenalidomide has demonstrated its activity in CLL, and its efficacy as maintenance therapy hints at an immune mediated mechanism that results in disease control albeit without impacting clinical responses.(19–21) Moreover routine preventative vaccinations in conjunction with lenalidomide has been determined to be a safe and effective in multiple myeloma.(22)

Herein, we present results from an National Cancer Institute/Cancer Therapy Evaluation Program (NCI/CTEP)-sponsored, randomized phase 2 study (NCI 8834) of low dose lenalidomide designed to assess the ability of lenalidomide to restore immune synapse response and humoral immunity, as well as delay progression of asymptomatic, genetically high-risk, early-stage CLL/small lymphocytic lymphoma (SLL) who did not meet the criteria for treatment. The trial was registered at clinicaltrials.gov (ClinicalTrials.gov Identifier: NCT01351896).

Methods:

Inclusion and exclusion criteria:

Treatment-naïve, adult patients aged ≥ 18 years and < 80 years with histologically identified chronic lymphocytic leukemia (CLL) or small lymphocytic lymphoma (SLL), as defined by the WHO classification of hematopoietic neoplasms were enrolled on the study starting in 2011. Patients were required to demonstrate one or more of the following high-risk genomic features: del(17)(p13.1) as detected by fluorescence in-situ hybridization (FISH) in >20% of cells; del(11)(q22.3) as detected by FISH in >20% of cells; complex karyotype (≥3 cytogenetic abnormalities on stimulated karyotype); or unmutated immunoglobulin heavy chain variable region (IGHV) (≥98% sequence homology compared with germline sequence).

Patients were excluded if they met any of the following consensus International Workshop on Chronic Lymphocytic Leukemia (IWCLL) 2008 criteria for initiating treatment: progressive splenomegaly and/or lymphadenopathy identified by physical examination or radiographic studies; progressive lymphocytosis with total white blood cell (WBC) count ≥ 300,000/μL; anemia (<11g/dL) or thrombocytopenia (<100,000/μL) due to bone marrow involvement; presence of unintentional weight loss > 10% over the preceding 6 months; NCI Common Terminology Criteria for Adverse Events (CTCAE) grade 2 or 3 fatigue; fevers >100.5° or night sweats for >2 weeks without evidence of infection; progressive lymphocytosis with an increase of >50% over a 2 month period or an anticipated doubling time of <6 months. All patients provided written informed consent and the study was conducted in accordance with rules specified under the Declaration of Helsinki, after approval by an institutional review board.

Study design

In this randomized, open-label, phase II trial, patients were randomized to receive lenalidomide either concurrent with (Arm A) or sequential to (Arm B) 2 doses of 13-valent protein-conjugated pneumococcal vaccine (PCV-13) administered 2 months apart (supplementary figure 1) in a 1:1 manner. The booster dosing was specifically employed to evaluate the efficacy and safety of repeated PCV13 vaccination given the historically poor responses in patients with CLL. Lenalidomide was dosed at 2.5 mg/day during the first 28-day cycle to reduce risk for tumor flare and increased to 5 mg/day for the second and subsequent cycles as tolerated. Treatment continued for at least 24 cycles in the absence of disease progression or irreversible Grade ≥ 3 adverse event (AE). AEs were summarized by and across treatment arms, and include the type, severity, and perceived attribution to study treatment according to NCI’s CTCAE (version 4). Descriptive statistics are provided for all correlative laboratory parameters.

Study Endpoints:

The primary endpoint of this study was the proportion of patients who achieve an antibody response to PCV13 vaccination. Following the definitions used by the World Health Organization (WHO) for serotype-based response as well as recent definitions of antibody response in the literature for multi-valent pneumococcal conjugate vaccine, we defined an antibody response in the context of this trial as achieving at least a four-fold increase in post-vaccination serotype-specific IgG titers or serotype-specific IgG concentrations of ≥ 0.35 μg/mL for 12 of 13 serotypes measured (1, 3, 4, 5, 6B, 7F, 9V, 14, 18C, 19A, 19F, 23F) by a standard enzyme-linked immunosorbent assay (ELISA) assay. These were measured as changes from baseline compared to repeat assessment 1 month after the second (booster) dose of PCV13. While it is not typical to employ a direct comparison of primary endpoints in the phase II setting, we believed that a randomized phase II design better controls for cohort and sampling variability necessary to assess the primary endpoint. The secondary endpoint of the study was to evaluate the clinical efficacy of treatment with lenalidomide in patients vaccinated with PCV13, where clinical efficacy was primarily determined through the proportion of patients who achieved a complete response (CR) within two years. Previous experience with lenalidomide suggested that late responses may be observed and hence a relatively substantial 2-year treatment regimen was explored. (19).

Sample size

Assuming that vaccine success in each arm was binomially distributed, we used a two sided chi-square test to compare the difference in proportions. We constrained the Type I error rate in the comparison of vaccine response rates to 0.10. Using a two-sided test, we had at least 90% power to detect a difference of 0.40 between the two rates (0.20 vs. 0.60) with a total of 44 evaluable patients (22 in each arm). The 40% cut-off was derived from earlier reports demonstrating up to a 39% response to protein conjugate vaccinations. Comparing these two vaccine success rates, if pA≠pB and the resulting p-value < 0.10, we considered the arm with the higher antibody response rate to have a significantly higher rate of vaccine success than the other arm. The difference in proportions allowed for a practical sample size, while comparing historical poor rate of vaccine efficacy with rates seen in non-immunocompromised patients.(23) This design also allowed for an interim analysis to be conducted after 11 patients have been accrued to each treatment arm (i.e. 50% of total planned accrual). Under a Lan-DeMets alpha spending function to constrain the Type I error rate, we considered it sufficient evidence that one arm had a significantly higher antibody response rate than the other arm if the observed p-value was <0.0056 (design generated using East version 5.3).

Independent of the comparison of vaccine-related efficacy, we also formally evaluated clinical efficacy using the two-year CR rate. Therefore, to effectively capture evidence of regimen efficacy (beyond the primary endpoint analysis) we assessed the CR rate in a pooled analysis across both arms since the main component of the regimen is lenalidomide therapy. For the purposes of this trial, if a patient had a documented CR on at least two consecutive evaluations during the first two years, they were deemed a clinical success. Given the extended time required to observe this clinical endpoint in these patients, we used a one-stage design. With 44 patients, we had at least 90% power and a Type I error of 0.10 to be able to detect a 25% or greater true CR rate versus the null hypothesis that the true CR rate is at most 10%. If 8 or more patients achieved a clinical success as defined above, then we considered this evidence of efficacy. With an estimated 10% drop out prior to administration of the 2nd vaccination, the final sample size was 22+2=24 in each arm for a total of 48 patients. Additional endpoints included progression free survival (PFS), time to next treatment (TTNT), and overall survival (OS). Time to event outcomes were summarized and explored between treatment arms using Kaplan-Meier methods. Formal comparison of these endpoints between the two arms were not done due to lack of statistical power.

Results:

Patient Demographics:

Forty-nine patients were randomized in a 1:1 manner to either arm; 24 in the concurrent arm (Arm A) and 25 in the sequential arm (Arm B) (Supplementary Figure 1). Baseline clinical and genetic factors were similar between the groups (Table 1). Median ages at diagnosis and study entry were 58 (34–70) and 59 (40–70) years respectively, with a median time from diagnosis of 1.3 (range 0.2–9.0) years. At the time of last follow up (up to Oct. 31^st, 2018), 24% of the patients were still continuing on lenalidomide with a median time on treatment of 3.7 years (Table 1). The median follow up was 5.5 years (range 4.3–7.0 years). Median CLL-International Prognostic Index (IPI) score at study entry was 2, with 14% of the patients harboring del(17)(p13.1) and 31% with del(11)(q22.3) on fluorescent in-situ hybridization (FISH) testing. Ninety-four percent of patients had unmutated IGHV and 47% had complex karyotype. There was no difference in baseline serotype specific serum titers (Supplementary Table 1).

Table 1:

Baseline Demographics

Characteristics	Arm A (Concurrent) (n=24)	Arm B (Sequential) (n=25)	All Patients (n=49)
Age at diagnosis, median(range)	58.5 (47–69)	57 (34–70)	58 (34–70)
Age at study entry, median(range)	61 (48–69)	57 (40–70)	59 (40–70)
On treatment, n(%)	7 (29)	5 (20)	12 (24)
Days on lenalidomide, median (range)	1229 (33–2487)	1365 (59–2261)	1346 (33–2487)
Reason for Discontinuation, n(%)
Adverse Events	8 (47)	7 (35)	15 (41)
Disease Progression Other	5 (29)	8 (40)	13 (35)
Other	4 (24)	5 (25)	9 (24)
ECOG PS, n(%)
0	21 (88)	23 (92)	44 (90)
1	3 (12)	2 (8)	5 (10)
Rai Stage at study entry, n(%)
0	13 (54)	13 (52)	26 (53)
1	11 (46)	11 (44)	22 (45)
2	0	1 (4)	1 (2)
Gender, n(%)
Male	16 (67)	18 (72)	34 (69)
Female	8 (33)	7 (28)	15 (31)
Race, n(%) Caucasian	22 (92)	25 (100)	47 (96)
IGHV unmutated, n(%)	23 (96)	23 (92)	46 (94)
B2 microglobulin, median (range)	2.4 (1.2–7.2)	2.4 (1.2–3.4)	2.4 (1.2–7.2)
FISH at study entry, n(%)
Del 17p13	1 (4)	6 (24)	7 (14)
Del 11q22	8 (33)	7 (29)	15 (31)
Complex Karyotype (≥3 cytogenetic abnormalities)	12 (50)	11 (44)	23 (47)
IPI score, median(range)	2 (0–6)	2 (0–7)	2 (0–7)
Low, n(%)	1 (4)	1 (4)	2 (4)
Intermediate, n(%)	19 (79)	18 (72)	37 (76)
High, n(%)	4 (17)	4 (16)	8 (16)
Very High, n(%)	0 (0)	2 (8)	2 (4)

Pharmacokinetic studies

A total of 320 concentration-time points were obtained from 41 patients who all received 5 mg per day of lenalidomide during the time at which PK samples were collected. One patient’s data was excluded from analysis because no 24-hour sample was available, and reliable pharmacokinetic parameter estimates could not be determined. Plots showing individual and mean concentration-time profiles are shown in Supplementary Figure 2A, 2B and 2C, and resulting pharmacokinetic parameter estimates from 40 patients are listed in Supplementary Table 2. Estimates were similar to those previously reported for CLL patients receiving 2.5–15 mg per day,(24) where reported clearance (CL) was 14.43 ± 9.05 L/hr, and in other disease populations receiving higher doses (25–75 mg) with CL ranging from 8.04 – 12.35 L/hr.(25,26) As has been demonstrated previously,(26) lenalidomide CL was correlated with creatinine CL, as calculated using the Cockroft-Gault formula.(27) No differences in pharmacokinetic parameters (maximum concentration (Cmax) and area under the curve (AUC)) were observed between patients with different clinical responses.

Safety and Tolerability

All patients experienced at least one AE but treatment with lenalidomide resulted in a significant incidence of grade ≥ 3 AEs (Table 2). Most common treatment emergent grade ≥3 AEs were hypertension (in 37.5% and 24% of patients in Arm A and Arm B respectively), neutropenia (in 25% and 16% of patients in Arm A and Arm B respectively), and thrombocytopenia (in 12.5% and 4% of patients in Arm A and Arm B respectively). Other notable AEs included diarrhea, infections (mostly grades 1–2), anemia, and fatigue (all grades 1–2). There were total of 190 incident infections of any type and any grade, occurring in 43 patients (99 infections occurred in 20 patients from arm A, 91 infections occurred in 23 patients from arm B. 6 patients (4 in arm A, 2 in arm B) did not have any infections during the study. Tumor flare was observed in 20% of all patients (all grades 1–2). Treatment related AE profiles were similar between arms and are detailed in Supplemental Table 3. Vaccination with the PCV13 was well tolerated with only injection site reactions the most notable AEs (Supplemental Table 4). There were no thromboembolic events. Sixty-one percent of patients required dose reductions to 2.5mg daily or every other day, most commonly for neutropenia and diarrhea (Supplemental Table 5). Secondary neoplasms occurred in 5 patients on arm A and 8 patients on arm B and were mostly cutaneous malignancies, with one episode of melanoma in situ, as well as a case of metastatic non-small cell lung cancer. There were no secondary hematologic malignancies.

Table 2:

All treatment emergent adverse events

	Arm A (n=24 )			Arm B (n=25 )
Adverse Event	Grades 1–2	Grades 3–4	All Grades	Grades 1–2	Grades 3–4	All Grades
	Number of patients (percent)
Hyperglycemia	23 (95.8)	1 (4.2)	24 (100)	23 (92)	1 (4)	24 (96)
Neutropenia	16 (66.7)	6 (25)	22 (91.7)	20 (80)	4 (16)	24 (96)
Hypertension	14 (58.3)	9 (37.5)	23 (95.8)	18 (72)	6 (24)	24 (96)
Diarrhea	16 (66.7)	2 (8.3)	18 (75)	17 (68)	4 (16)	21 (84)
Thrombocytopenia	15 (62.5)	3 (12.5)	18 (75)	18 (72)	1 (4)	19 (76)
Maculopapular rash	14 (58.3)	2 (8.3)	16 (66.7)	10 (40)	0	10 (40)
Anemia	15 (62.5)	0	15 (62.5)	15 (60)	0	15 (60)
Lymphocyte count increased	11 (45.8)	2 (8.3)	13 (54.2)	12 (48)	7 (28)	19 (76)
Upper respiratory tract infection	13 (54.2)	0	13 (54.2)	15 (60)	0	15 (60)
Headache	13 (54.2)	0	13 (54.2)	17 (68)	0	17 (68)
Fatigue	11 (45.8)	0	11 (45.8)	20 (80)	0	20 (80)
Alanine aminotransferase increased	11 (45.8)	0	11 (45.8)	13 (52)	2 (8)	15 (60)
Cough	11 (45.8)	0	11 (45.8)	22 (88)	0	22 (88)
Sinus Bradycardia	11 (45.8)	0	11 (45.8)	7 (28)	0	7 (28)
Lymphocyte count decreased	8 (33.3)	2 (8.3)	10 (41.7)	6 (24)	2 (8)	8 (32)
Sore throat	10 (41.7)	0	10 (41.7)	14 (56)	0	14 (56)
Myalgia	10 (41.7)	0	10 (41.7)	13 (52)	0	13 (52)
Hypokalemia	7 (29.2)	3 (12.5)	10 (41.7)	10 (40)	0	10 (40)
Allergic rhinitis	10 (41.7)	0	10 (41.7)	10 (40)	0	10 (40)
Aspartate aminotransferase increased	10 (41.7)	0	10 (41.7)	12 (48)	1 (4)	13 (52)
Generalized pain	9 (37.5)	0	9 (37.5)	7 (28)	0	7 (28)
Lung infections	5 (20.8)	4 (16.7)	9 (37.5)	0	0	0
Back pain	9 (37.5)	0	9 (37.5)	7 (28)	0	7 (28)
Weight loss	8 (33.3)	0	8 (33.3)	8 (32)	1 (4)	9 (36)
Creatinine increased	8 (33.3)	0	8 (33.3)	5 (20)	0	5 (20)
Hyperuricemia	7 (29.2)	0	7 (29.2)	10 (40)	0	10 (40)
Hypocalcemia	7 (29.2)	0	7 (29.2)	9 (36)	0	9 (36)
Fever	7 (29.2)	0	7 (29.2)	9 (36)	0	9 (36)
Sinusitis	7 (29.2)	0	7 (29.2)	9 (36)	0	9 (36)
Dysesthesia	7 (29.2)	0	7 (29.2)	7 (28)	0	7 (28)
Dyspnea	6 (25)	0	6 (25)	12 (48)	0	12 (48)
Productive cough	6 (25)	0	6 (25)	9 (36)	0	9 (36)
Lymph node pain	6 (25)	0	6 (25)	8 (32)	0	8 (32)
Nausea	5 (20.8)	0	5 (20.8)	10 (40)	0	10 (40)
Blood bilirubin increased	4 (16.7)	1 (4.2)	5 (20.8)	6 (24)	1 (4)	7 (28)
Dizziness	4 (16.7)	1 (4.2)	5 (20.8)	9 (36)	0	9 (36)
Hypoglycemia	5 (20.8)	0	5 (20.8)	9 (36)	0	9 (36)
Hypophosphatemia	2 (8.3)	2 (8.3)	4 (16.7)	6 (24)	7 (28)	13 (52)
Bruising	4 (16.7)	0	4 (16.7)	10 (40)	0	10 (40)
Abdominal pain	3 (12.5)	1 (4.2)	4 (16.7)	7 (28)	2 (8)	9 (36)
Infections and infestation	4 (16.7)	0	4 (16.7)	8 (32)	0	8 (32)
Alkaline phosphatase increased	4 (16.7)	0	4 (16.7)	7 (28)	0	7 (28)
Benign and malignant neoplasms (including cysts and polyps)	4 (16.7)	0	4 (16.7)	7 (28)	1 (4)	8 (32)
Urinary tract infection	2 (8.3)	1 (4.2)	3 (12.5)	4 (16)	0	4 (16)
Falls	2 (8.3)	1 (4.2)	3 (12.5)	9 (36)	0	9 (36)
Anorexia	3 (12.5)	0	3 (12.5)	3 (12)	1 (4)	4 (16)
Hypomagnesemia	3 (12.5)	0	3 (12.5)	0	0	0
Vomiting	0	0	0	6 (24)	1 (4)	7 (28)
Constipation	0	0	0	11 (44)	0	11 (44)
Oral mucositis	0	0	0	7 (28)	0	7 (28)
Neck pain	0	0	0	3 (12)	0	3 (12)
Postnasal drip	0	0	0	9 (36)	0	9 (36)

Vaccination efficacy

Seroprotection against 12 out of the 13 pneumococcal serotypes included in the PCV13 vaccine (1, 3, 4, 5, 6B, 7F, 9V, 14, 18C, 19A, 19F, and 23F) was measured 4 weeks after the second dose of vaccine (Table 3). Baseline serotype specific mean titers were >0.35ug/ml for all serotypes tested and similar across both arms (Supplemental table 1). Four serotypes (3, 4, 5, 6B) achieved the additional apriori seroprotection definition of a 4-fold increase in arm A, and 6 serotypes (3, 4, 5, 6B, 19A, 19F) achieved the 4-fold increase in arm B (Table 3). All patients achieved the defined concentration of 0.35ug/ml for at least one serotype tested. No significant difference was observed with the addition of lenalidomide.

Table 3:

Serotype specific vaccine titers

Specific Serotypes	Arm A			Arm B
	Cycle 3 titers	Cycle 6 titers	Fold Change	Cycle 1 titers	Cycle 4 titers	Fold change
	Mean serotype specific titers (μg/ml)			Mean serotype specific titers (μg/ml)
1	13.66	16.9	1.24	7.3	17.15	2.35
3	2.05	9.47	4.62	2.9	20.11	6.93
4	0.5	6.06	12.12	1.39	18.25	13.13
5	2.91	12.91	4.44	4.35	26.93	6.19
6B	4.05	17.78	4.39	6.66	60.18	9.04
7F	4.87	5.7	1.17	7.64	17.04	2.23
9V	3.97	6.23	1.57	4.45	14.02	3.15
14	2.34	3.65	1.56	6.85	12.9	1.88
18C	3.76	9.57	2.55	3.8	10.55	2.78
19A	5.06	12.13	2.40	6.792	36.96	5.44
19F	6.2	24.65	3.98	6.04	34.46	5.71
23F	9.35	16.83	1.80	15.79	45.28	2.87

Mean IgG/IgM/IgA levels at baseline were 722/109/49 mg/dL, and improved to 820/136/51 mg/dL and 947/197/59 mg/dL after 12 and 24 cycles of treatment, respectively. IgG increased from baseline to cycle 24 for both arms (arm A: p=0.04; arm B: p=0.01) with no difference in the amount of increase across arms (p=0.15). IgA increased from baseline to cycle 12 (p=0.008) and then to cycle 24 in Arm A (p=0.049), but not in Arm B (p=0.81, p=0.27). No significant increase in IgM was observed for either of arm A or B, Supplemental Figure 3). This suggests that while administration of lenalidomide did have a positive impact on restoration of humoral immunity, the sequence of administration with the vaccine had no influence on this outcome. The use of lenalidomide was also associated with a substantial increase in Th2 cytokines and TNF-α during the early phase of treatment, with a plateau and decline in concentration in the latter part of treatment. A similar but lower increase was observed for Th1 cytokines with lenalidomide treatment (Figure 1). In addition, lenalidomide induced modulation of known proteins was also not impacted by vaccination (Supplemental Figure 4).

Figure 1: Serum cytokine expression levels in CLL patients treated with lenalidomide:

Cytokine levels were measured in the serum of 10 different patients prior to starting therapy (C1), 3 months for arm A and 4 months for arm B, 6 months for arm A and 7 months for arm B, 12 months for arm A and 16 months for arm B, after lenalidomide treatment. In arm A the levels of the cytokines shown increases significantly on treatement with lenlidomide but not in arm B. *p < 0.05.

Efficacy

IWCLL 2008 response was assessed at the completion of 24-months of treatment. Overall response rate was 56% (partial response (PR) in 53% and CR in 3%). An additional 36% of patients achieved stable disease. Only 2 patients experienced disease progression during the first 2-years of treatment with lenalidomide. No significant differences in the response were observed between two arms. At a median time on treatment of 3.6 years, median PFS was 5.8 years (95% CI 3.1-NR). PFS at 1 year 85% (95% CI 72–93), 2 year PFS was 79% (64–88), and estimated 3 year is PFS 72% (95% CI 57–83) (Figure 2, and Supplemental Table 6). TTNT and OS was also similar between the 2 arms (Figure 3 and Supplemental Figure 5). Additionally, patients with del(17)(p13.1), those with a CLL-IPI score of 3 or higher were also more likely to have an inferior PFS and TTNT (Supplementary Tables 7, 8 and 9) on multivariable analyses.

Figure 2: Progression free survival:

No difference was observed in progression free survival between patients in the concurrent arm (Arm A) vs. the sequential arm (Arm B), p=0.5

Figure 3: Time to next treatment per arm:

Cumulative incidence of starting next treatment was similar in patients treated on the concurrent arm (Arm A) vs. the sequential arm (Arm B), p=0.88

Discussion

Our study suggests that low-dose lenalidomide can be administered to asymptomatic, genetically high-risk early-stage CLL patients with modest toxicity and high rates of durable clinical response. Since infections constitute the leading cause of morbidity for patients with CLL, our study was also designed to evaluate the impact of an immunomodulatory therapeutic approach to enhance vaccine efficacy in an effort to decrease the infectious complications. The addition of lenalidomide did not appear to enhance vaccine responses as measured by serotype specific vaccine responses. This effect could also be explained by the high baseline anti-serotype specific titers reflecting the increased utilization of pneumococcal vaccine in patients with CLL. A limitation of our study is the lack of detail of prior vaccination history of our study population since administration records are not accurate or readily available. Nevertheless, baseline serotype specific data suggests a high incidence of vaccination in our population which could have blunted the impact of PCV13 vaccinations. In addition, all patients demonstrated a response to at least one of the serotypes included in the PCV13 vaccine, and a 4-fold increase in serotype specific titers were observed for a number of serotypes in both arms. These results suggest that patients early in their disease process appear to have a fairly robust response to 2 doses of PCV13 vaccine administered 2-months apart, a result that is consistent with recent reports, and this response is not significantly impacted by the use of lenalidomide.(28) However, these results can also be partly explained by the generally younger patient population included in our study as compared to the average CLL patient population and the resultant robust immune responses. Nevertheless, the vaccination approach appeared to be safe and well tolerated. These results are also consistent with earlier studies demonstrating an improved response to protein-conjugated vaccines in patients with CLL and provides evidence for the safety and utility of this vaccination approach for patients with CLL. Patients also had a sustained improvement in serum IgG levels, consistent with prior reports, and suggesting restoration of humoral immunity, and differentiating this therapy from experience gleaned from the use of chemoimmunotherapy or BTK inhibitors.(3,29) Interestingly, while lenalidomide treatment resulted in increase in multiple serum cytokines, a definite trend towards Th1 polarization was not observed. This is contrary to recent reports demonstrating Th1 polarization in the CLL lymph node microenvironment and is perhaps reflective of the limited utility of the assessment of serum cytokines as a marker for Th polarization.(30)

Recent reports detailing patient perception regarding CLL treatment suggest a high rate of anxiety and stress related to the conventional approach of “watchful waiting” for patients with early stage disease, and earlier intervention might result in lower stress levels and improved quality of life provided the therapy is well tolerated.(31,32) While these historical “watchful waiting” recommendations are based on established data demonstrating a lack of improvement in survival outcomes with the early use of traditional chemotherapeutic options, it is open for debate in the modern era where well tolerated and less toxic options are available.(33) However, given the heterogeneity of the CLL disease process, an early intervention approach can be considered at this time only for patients with high-risk disease, since treatment free and overall survival outcomes are excellent in patients with low risk disease. In addition, early data from the CLL12 trial suggests improvement in PFS and event-free survival with ibrutinib utilized as an early intervention strategy in patients with high-risk disease, as compared to placebo.(34) Our study was performed prior to the development of the CLL-IPI score and more formal risk stratification parameters, and utilized prevalent criteria of del(17)(p13.1), del(11)(q22.3), and unmutated IGHV as definition of high risk disease.(35) The majority of the patients had intermediate risk disease according to the CLL-IPI score and this partly explains the high response rates and PFS outcomes observed in our cohort of patients. Nevertheless, this does suggest an improvement in TTNT (median not reached at 5.5 years of median follow up) compared to a similar population of intermediate CLL-IPI score (median TTNT of 55 months).(36) These results compare favorably with previous reports of lenalidomide as single agent therapy, however, it failed to reach our pre-specified efficacy criteria based on CR rates, possibly secondary to the low dose of lenalidomide used in our study and despite the fact that median age from diagnosis to treatment was 1.3 years (range 0.2–9 years) and arguably included patients who were not clinically behaving as having high risk disease.

Recent data has demonstrated the efficacy of lenalidomide for CLL in multiple settings including patients with previously untreated disease, with relapsed disease, in the maintenance setting, and in combination with chemotherapeutic approaches.(19–21) While our study demonstrates the safety and efficacy of lenalidomide as early treatment for patients with high-risk CLL who did not meet the conventional IWCLL criteria for treatment, a substantial number of patients did experience adverse events requiring dose reductions. This limits its utility as standard treatment in this setting. Nevertheless, despite the fact that high rate of CRs were not observed, we did observe prolonged disease control rates with manageable toxicity with the use of lenalidomide. With the advent of exciting new and better tolerated therapeutic agents like bruton tyrosine kinase (BTK) inhibitors and bcl-2 antagonists, early intervention has the potential to become a standard therapeutic option.

Our study provides the feasibility of an early intervention strategy for patients with high risk CLL. While lenalidomide was efficacious with manageable toxicities in this setting, it failed to achieve the primary endpoint of improving response to PCV13 vaccine. Additional well-controlled clinical trials are required of selected high risk patients, including the elderly and those with documented lack of responsiveness to vaccines; with quality of life and survival end points; in addition to evaluating strategies designed to reduce the burden of common comorbidities that affect our patients.

Key Points:

• - Early intervention strategy with lenalidomide in patients with high-risk CLL is efficacious and results in durable remissions

• - Lenalidomide does not enhance vaccination response to pneumococcal vaccines

Statement of Translational Relevance.

Our randomized phase II study evaluated clinical outcomes of patients with genetically high-risk CLL or SLL (defined as unmutated IGHV, deletion(17p) or (11q), and/or complex abnormal karyotype), who were treated with lenalidomide as an early intervention strategy (patients not meeting conventional IWCLL criteria for treatment) in an attempt to augment pneumococcal vaccine directed humoral responses. Extensive laboratory correlates established pharmacokinetics of lenalidomide in this population along with its safety and efficacy. Additional serological testing delineated serotype specific humoral responses to pneumococcal vaccination in our patients. These translational efforts were employed to develop better strategies to mitigate infectious morbidity and mortality in patients with CLL.

List of Abbreviations:

CLL

chronic lymphocytic leukemia

PCV13

protein-conjugated pneumococcal vaccine

Ig

immunoglobulin

NCI/CTEP

National Cancer Institute/Cancer Therapy Evaluation Program

SLL

small lymphocytic lymphoma

FISH

fluorescence in-situ hybridization

IGHV

immunoglobulin heavy chain variable region

IWCLL

International Workshop on Chronic Lymphocytic Leukemia

WBC

white blood cell

CTCAE

Common Terminology Criteria for Adverse Events

ECOG

Eastern Cooperative Oncology Group

ULN

upper limit of normal

ALT

alanine aminotransferase

AST

aspartate aminotransferase

H

Histamine

AE

adverse event

WHO

World Health Organization

ELISA

enzyme-linked immunosorbent assay

CR

complete response

PFS

progression free survival

TTNT

time to next treatment

OS

overall survival

EDTA

Ethylenediaminetetraacetic acid

LC/MS

liquid chromatography/mass spectroscopy

SDS-PAGE

sodium dodecyl sulfate–polyacrylamide gel electrophoresis

IPI

International Prognostic Index

FISH

Fluorescent in-situ hybridization

CL

clearance

Cmax

maximum concentration

AUC

area under the curve

PR

partial response

BTK

bruton tyrosine kinase