Chronic Lymphocytic Leukemia Study Research Articles

Costs and health care resource utilization among Medicare beneficiaries diagnosed with chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is the most common type of leukemia. However, published studies of CLL have either only focused on costs among individuals diagnosed with CLL without a non-CLL comparator group or focused on costs associated with specific CLL treatments. An examination of utilization and costs across different care settings provides a holistic view of utilization associated with CLL. To quantify the health care costs and resource utilization types attributable to CLL among Medicare beneficiaries and identify predictors associated with each of the economic outcomes among beneficiaries diagnosed with CLL. This retrospective study used a random 20% sample of the Medicare Chronic Conditions Data Warehouse (CCW) database covering the 2017-2019 period. The study population consisted of individuals with and without CLL. The CLL cohort and non-CLL cohort were matched using a 1:5 hard match based on baseline categorical variables. We characterized economic outcomes over 360 days across cost categories and places of services. We estimated average marginal effects using multivariable generalized linear regression models of total costs and across type of services. Total cost was compared between CLL and non-CLL cohorts using the matched sample. We used generalized linear models appropriate for the count or binary outcome to identify factors associated with various categories of health care resource utilization, such as inpatient admissions, emergency department (ED) visits, and oncologist/hematologist visits. A total of 2,736 beneficiaries in the CLL cohort and 13,571 beneficiaries in the non-CLL matched cohort were identified. Compared with the non-CLL cohort, the annual cost for the CLL cohort was higher (CLL vs non-CLL, mean [SD]: $22,781 [$37,592] vs $13,901 [$24,725]), mainly driven by health care provider costs ($6,535 vs $3,915) and Part D prescription drug costs ($5,916 vs $2,556). The main categories of health care resource utilization were physician evaluation/management visits, oncologist/hematologist visits, and laboratory services. Compared with beneficiaries aged 65-74 years, beneficiaries aged 85 years or older had lower use and cost in maintenance services (ie, oncologist visits, hospital outpatient costs, and prescription drug cost) but higher use and cost in acute services (ie, ED). Compared with residency in a metropolitan area, living in a nonmetropolitan area was associated with fewer physician visits but higher ED visits and hospitalizations. The cooccurrence of lower utilization of routine care services, along with higher utilization of acute care services among some individuals, has implications for patient burden and warrants further study.

Fludarabine, cyclophosphamide, and rituximab as first-line treatment in patients with chronic lymphocytic leukemia: A long-term analysis of the German CLL Study Group (GCLLSG) registry.

Long-term data of chronic lymphocytic leukemia (CLL) patients with favorable risk who were treated with fludarabine, cyclophosphamide, and rituximab (FCR) within clinical trials show good efficacy. We here report long-term data collected within the GCLLSG registry. Altogether, 417 CLL patients who received first-line treatment with FCR were analyzed, of which 293 (70.3%) were treated outside of clinical trials. The median observation time from first-line was 95.8 (interquartile range 58.7-126.8) months. Focusing on data of 194 (46.5%) patients who received FCR first-line treatment after 2013 (start of data collection within GCLLSG registry), responses were documented in 85% of the patients, non-responses in 15%, and for 3.6% the assessment was missing. Median event-free survival (EFS, time until disease progression, subsequent treatment, or death) was 60.2 months with a 5-year EFS-rate of 50.6%. Patients with higher-risk disease, characterized by unmutated IGHV (N = 78), had a median EFS of 45.4 months with a 5-year EFS rate of 36.3%, while the median EFS was 77.5 months with a 5-year EFS rate of 60.3% in patients with mutated IGHV (N = 40). Median overall survival was not reached with a 5-year survival rate of 92.7%. In summary, first-line FCR was associated with long EFS, especially in patients exhibiting a mutated IGHV status.

Abstract P01: Impact of Germline and Somatic ATM Variants in Chronic Lymphocytic Leukemia (CLL): Clinical Implications and Response to PARP Inhibition

Abstract Background: Next-generation sequencing has revealed numerous Variants of Unknown Significance (VUS) in the ATM gene, important in DNA repair and chronic lymphocytic leukemia (CLL) pathogenesis. This study evaluates the effect of rare ATM germline variants, including VUS, on features of CLL, ATM kinase activity, and response to PARP inhibition. Methods:ATM variants were detected in 885 CLL patients (327 DFCI; 276 German CLL Study Group; 282 ICGC) using DeepVariant, and compared to 5310 ancestry-matched controls. Concurrently, 278 CLL patients with ATM aberrations from clinical sequencing were retrospectively analyzed, annotated by population allele frequency (JCO 2023, Lampson et al.). ATM kinase activity was measured by ATM and KAP1 phosphorylation following gamma-irradiation in primary CLL samples. The impact of talazoparib, a PARP inhibitor, on ATM aberrant and wild-type CLL cell proliferation was also assessed. Results: Among the 885 CLL cases, 130 ATM germline variants were identified by DeepVariant, including 87 missense variants. The top 5 most frequent rare germline missense variants with high odds ratio for association with CLL were ATM p.L2332P (AF cases vs controls, 0.328% vs 0.0188%, OR 17.46), p.L2307F (OR 16.88), p.F763L (OR 18.89), p.Y1442H (OR 35.39) and p.S99G (OR 16.28). In our retrospective clinical cohort, we identified 294 ATM variants with 146 unique variants. Our previously proposed algorithm for classifying germline vs somatic identified 74 germline, with 70 missense, and 72 somatic. Phosphorylation of ATM and KAP1 (pATM, pKAP1) in response to IR was reduced in 13 patients assessed with germline ATM variants (9 rare and 4 non-rare; ATM variant alone group) compared to those without any somatic or germline ATM aberrancy (WT group) (P<0.01). Phosphorylation levels were also significantly lower in 17 patients with rare germline ATM variants with concurrent del(11q) (del11q + ATM germline variant group) compared to 15 with del(11q) alone (del 11q group) (P<0.01). We obtained similar results from Western blot analysis. To assess the sensitivity of ATM deficient cells to talazoparib, we compared proliferation inhibition across groups cocultured with talazoparib for 14 days. The del11q with ATM-somatic group exhibited significantly greater sensitivity to talazoparib at lower dosages compared to ATM WT (talazoparib 0.1 μM, P<0.05; 0.5μM, P=0.098). The monoallelic ATM deficient groups (del11q alone or ATM-germline alone) tended to have lower proliferation, but not significantly. Higher concentrations of talazoparib (2.5μM) strongly inhibited proliferation of all CLLs including WT. Conclusion: This study highlights the prevalence of rare ATM germline variants in CLL and their impact on ATM function in CLL. Our findings also suggest that these variants affect the DNA damage response in primary CLL cells and that ATM biallelic deficiency influences sensitivity to PARP inhibitors. Our evidence could lead to more personalized therapeutic strategies for CLL patients with distinct ATM aberrancies. Citation Format: Kiyomi Mashima, Nicholas Moore, Mariia Mikhaleva, Anna Petráčková, Samantha Shupe, Stacey M Fernandes, Amaro Taylor-Weiner, Riaz Gillani, Hoyin Chu, Seunghun Han, Sabrina Camp, Eric Kofman, Gad Getz, Catherine J. Wu, Svitlana Tyekucheva, Matthew S Davids, Eliezer Mendel Van Allen, Saud AlDubayan, Jennifer R Brown. Impact of Germline and Somatic ATM Variants in Chronic Lymphocytic Leukemia (CLL): Clinical Implications and Response to PARP Inhibition [abstract]. In: Proceedings of the Blood Cancer Discovery Symposium; 2024 Mar 4-6; Boston, MA. Philadelphia (PA): AACR; Blood Cancer Discov 2024;5(2_Suppl):Abstract nr P01.

Impact of Baseline Comorbidities and Treatment on Cardiovascular (CV) Outcomes in Patients with Chronic Lymphocytic Leukemia (CLL): Results of a Systematic Literature Review (SLR) of Interventional Trials and a Targeted Literature Review (TLR) of Real-World Observational Studies in CLL

Background: Ibrutinib, the first Bruton tyrosine kinase inhibitor (BTKi) indicated for CLL, has been associated with a higher risk of CV events, including atrial fibrillation (AF) and hypertension, vs non-BTKi active comparators and placebo. In a phase 3 clinical trial of patients (pts) with CLL comparing ibrutinib with acalabrutinib-a more selective, second-generation BTKi-CV event incidence was numerically higher with ibrutinib and the incidence and cumulative risk of AF were significantly higher with ibrutinib (Byrd JC et al. J Clin Oncol. 2021;39:3441-52). Ibrutinib's toxicity profile prompted changes to its prescribing information and lowered its recommendation status in current CLL guidelines. We conducted an SLR of interventional trials in CLL followed by a supplemental TLR of observational studies in CLL to explore the extent to which CV outcomes of pts with CLL are driven by their baseline comorbidities or treatments. Methods: Embase and MEDLINE were searched to identify evidence in peer-reviewed journals (cutoff date: June 23, 2023). For the SLR, limits were English articles published in 2012 or later, and inclusion criteria specified phase 2/3 interventional trials in adults aged ≥18 y with treatment-naive or relapsed/refractory CLL. Treatments included BTKis, BCL-2 inhibitors, and chemoimmunotherapy. CV outcomes were arrhythmia, heart failure, major adverse CV events, major bleeding, myocardial infarction, stroke, sudden cardiac death, and venous thromboembolism. For the TLR, language and time limits were analogous to the SLR. Additional limits were observational studies with >70% CLL pts receiving any or no treatment. The TLR assessed baseline CV events/risk, including arrhythmia, heart failure, major adverse CV events, major bleeding, myocardial infarction, stroke, sudden cardiac death, and venous thromboembolism. Results: The SLR yielded 55 studies. Among the phase 3 randomized controlled trials, incidence ranges were 0%-16% for AF/atrial flutter, 0%-6.9% for heart failure, and 0%-11% for major bleeding; rates were generally higher with BTKis, among which ibrutinib appeared to be associated with the highest rates ( Figure 1). CV data from the SLR were sparse and heterogeneous, primarily due to limited baseline CV risk reporting and substantial variability in CV adverse event reporting. To clarify relationships of CV risk with treatment, we performed the TLR, which yielded 24 studies. Analysis of baseline CV risk reported in the observational studies yielded several key findings ( Figure 2): 1) in pts with CLL, the risk of CV events, including AF, increased with age; in pts aged 65−74 y and ≥65 y, the reported risk of AF was 2.4-fold and 2.8-fold higher, respectively, than that of pts aged <65 y (Shanafelt TD et al. Leuk Lymphoma. 2017;58:1630-9; Visentin A et al. Hematol Oncol. 2019;37:508-12); 2) baseline CV risk was similarly high in pts with CLL and matched non-CLL comparators (Larsson K et al. Am J Hematol. 2022;97:E255-7); 3) ibrutinib was associated with a higher risk of CV disease relative to chemotherapy, even when prior AF, age, and anticoagulant exposure were controlled for (Abdel-Qadir H et al. J Clin Oncol. 2021;39:3453-62). In ibrutinib-treated pts with a B-cell malignancy followed in cardio-oncology clinics (N=53; n=38 CLL), median time to new onset of ibrutinib-related AF was 5.5 mo; AF incidence was 15-fold higher in ibrutinib-treated pts overall vs that expected in an age- and sex-comparable general population and in ibrutinib-treated pts with CLL vs that expected in pts with CLL and no ibrutinib treatment (Baptiste F et al. Open Heart. 2019;6:e001049). Conclusions: CLL primarily affects the elderly, who are at higher risk of CV events vs a younger demographic. Results suggest that some pts with CLL have an increased risk of adverse CV outcomes; thus, it would be inaccurate to attribute the higher rates of CV events observed in pts with CLL solely to their treatment as it is plausible that pts with CLL already have an elevated CV risk contributing to such outcomes. However, our findings suggest that the first-generation BTKi (ibrutinib) may impact CV outcomes in these pts; data examining the CV risk of second-generation BTKis and other novel therapies (eg, venetoclax) are limited. Future research that consistently estimates baseline CV risk and closely monitors CV parameters in pts with CLL is required to ascertain the true risk of CV disease due to CLL treatment.

Aberrant Heterochromatin Silences Immune Response Genes in Chronic Lymphocytic Leukemia

Introduction: Recent studies have shown that clonal B cells derived from patients with chronic lymphocytic leukemia (CLL) have altered histone modification landscapes compared to B cells from healthy controls. We have also shown that the BCR signaling pathway impacts the chromatin landscapes of CLL B cells, implicating the interaction between oncogenic signaling and epigenetic machinery in CLL (Wang et al., Blood Cancer J. 2022; Wang et al., Cancer Res (2023) 83 (7_Supplement): 4725). However, the majority of the chromatin studies in CLL concentrate on the active enhancers associated with oncogene expression; there is a very limited understanding of lost enhancers in CLL: 1) What is the function of lost enhancers; 2) How are these lost enhancers maintained in CLL; 3) Can these lost enhancers be detected at the precursor state of CLL, monoclonal B-cell lymphocytosis (MBL); 4) if this aberrant chromatin signature affected by CLL treatment. Methods: We systematically mapped the genome-wide histone modification profile (histone marks: H3K4me1, H3K4me3, H3K9me3, H3K27ac, H3K27me3, and H3K36me3) in 22 previously untreated CLL samples, 6 high count MBL samples, and peripheral blood B cells from 6 healthy donors by Cleavage Under Targets and Tagmentation (CUT&Tag). We also analyzed B cells from 8 patients before and after one year of continuous Bruton tyrosine kinase inhibitor (BTKi) ibrutinib treatment. The genomic regions with a decrease in both H3K4me1 and H3K27ac enrichment (analyzed by Deseq2, fold change > 1, p < 0.01) were named lost enhancers. We used CRISPR-mediated epigenetic editing to study the role of gained H3K9me3 on EBF1 gene expression. Results: Compared to normal B cells, the lost enhancers (genomic regions defined by decreased enrichment of H3K4me1 and H3K27ac) in CLL B cells showed increased heterochromatin marks (H3K9me3 and H3K27me3) enrichment. Importantly, these enhancers are associated with genes downregulated in CLL and involved in the immune response, indicating the source of epigenetic impact on impaired immunity in CLL. We further found that this aberrant chromatin structure also exits in clonal B cells from the precursor stage of MBL, implicating the importance of epigenetic silencing in CLL evolution. In particular, we found the lost enhancer signature across the gene locus EBF1, an important transcription factor that defines B cell lineage and regulates normal B cell activation. Using CRISPR-mediated epigenetic editing, we demonstrated that the gain of H3K9me3 is sufficient to silence the EBF1 enhancer and its gene expression. Finally, we found that while BTKi ibrutinib treatment partially inhibits the CLL-gained enhancers in some patients, it failed to restore the lost enhancers in CLL B cells. Conclusion: In summary, we have elucidated an epigenetic silencing mechanism that may contribute to the loss of physiological function of normal B cells, which places the clonal B cells at a selection advantage during CLL evolution and increases the risk of serious infections in CLL and MBL ( Fig 1).

Cumulative Review of Hypertension in Patients with Chronic Lymphocytic Leukemia (CLL) and Other Hematologic Malignancies Treated with Acalabrutinib

Background: Patients (pts) with CLL have a high rate of comorbidities, including cardiovascular disease (CVD). Comorbid CVD is correlated with higher mortality in this pt population. Hypertension (HTN) is the most common concurrent CVD in pts with CLL and is associated with development of complications such as stroke and heart failure. The incidence of new or worsening HTN in treatment-naive CLL pts receiving placebo within the CLL12 trial was 8.3% over a 69.3-month observation period (Langerbeins et al. Hematol Oncol. 2023). New and/or worsening HTN has been observed in pts with CLL treated with Bruton tyrosine kinase inhibitor (BTKi) therapy. Studies have shown new or worsening HTN with ibrutinib (7−30%) and zanubrutinib (11−20%) (Christensen et al. Expert Rev Hematol. 2022). Acalabrutinib (acala), a selective, second-generation BTKi, demonstrated a lower rate of HTN (8.6%) vs ibrutinib (22.8%) in the head-to-head ELEVATE-RR study at a median follow-up of 40.9 months (Byrd et al. J Clin Oncol. 2021). Here, we performed a cumulative analysis of new or worsening HTN from the acala global development program and compared these data against real-world prevalence and incidence of HTN for pts with CLL prior to treatment initiation using claims data. Methods: Safety reports of new or worsening HTN were obtained from 11 clinical trials with acala (the safety database analysis comprised 2034 pts from studies in CLL [86.3%] and mantle cell lymphoma [6.1%], with the remainder from other B-cell malignancies). The clinical trials analyzed included therapy with acala (monotherapy or in combination with obinutuzumab) and active comparator groups (described in Table 1). Using the narrow Standardized MedDRA Query and the related MedDRA preferred terms for “HTN,” exposure-adjusted incidence rates in events/100 person-years were determined, and as a comparison, HTN prevalence was assessed at median treatment exposure for acala monotherapy. HTN prevalence (proportion of pts with HTN 6 months before treatment initiation) and incidence rates (number of new cases of HTN per 100 person-years during the 5 years before any treatment initiation) were also assessed based on ICD-10 codes from 3 adjudicated closed claims databases (Optum, MarketScan, and IQVIA). Results: In the claims database analysis, prevalence of HTN ranged from 47.3% to 66.1% in pts with TN CLL prior to treatment initiation. HTN prevalence with acala monotherapy at median treatment exposure (45.5 months) was 55.7%. Exposure-adjusted incidence rates of HTN were analyzed from 11 clinical trials; 1225 pts received acala monotherapy (947 pts with CLL), 223 pts received acala + obinutuzumab (all pts with CLL), 264 pts received ibrutinib as a comparator (all pts with CLL), and 322 pts received other comparators (all pts with CLL). Among the 1225 pts treated with acala monotherapy, 165 pts (147 with CLL) experienced a treatment-emergent adverse event of HTN (median age 66 years; 66.1% [n=109] male; 59.4% [n=98] with history of HTN at baseline). The exposure-adjusted incidence rates of HTN with acala monotherapy (3.844) and acala + obinutuzumab (3.926) were similar to that of other comparators (4.788 [excluding ibrutinib]) ( Table 1). In the ELEVATE-RR study specifically (subgroup of the current analysis), the exposure-adjusted incidence rate of HTN with acala (3.053) was approximately one-third that of ibrutinib (8.590). The exposure-adjusted incidence rate of HTN in pts treated with acala monotherapy without history of HTN was 3.239. Conclusions: Our data showed that HTN is a common comorbidity in pts with CLL as demonstrated in the claims database and clinical trial database analyses where 59.4% of pts who experienced HTN with acala monotherapy had history of HTN at baseline. The incidence rates of new or worsening HTN with acala monotherapy and acala + obinutuzumab were low (3.844 and 3.926, respectively). This is supported by similar incidence proportion of new or worsening HTN in pts receiving placebo (8.3%) from CLL12 (median follow-up 69.3 months) and acala monotherapy (8.9%) from ELEVATE-TN (median follow-up 58.2 months; Sharman et al. EHA. 2022), and similar HTN prevalence at median treatment exposure vs before treatment initiation in claims databases. Our analysis, conducted predominantly in patients with CLL, suggests that acala monotherapy did not worsen pre-existing HTN or increase the risk of new-onset HTN.

Ianalumab, a Novel Anti-B-Cell Activating Factor (BAFF) Receptor (BAFF-R) Monoclonal Antibody (mAb) in Development for Immune Thrombocytopenia (ITP) and Warm Autoimmune Hemolytic Anemia (wAIHA), Has Demonstrated a Favorable Safety Profile in Sjögren's Syndrome (SjS), Systemic Lupus Erythematosus (SLE) and Chronic Lymphocytic Leukemia (CLL)

Background: Current therapies for ITP and wAIHA are mostly symptomatic and rarely lead to durable responses after discontinuation; thus, chronic treatment is often needed, which may be associated with toxicity and substantial quality-of-life burden for patients. There is an unmet need for disease-modifying therapies that can induce and maintain durable responses beyond treatment completion in ITP and wAIHA. BAFF is an important regulator of B-cell differentiation, proliferation and survival through interactions with BAFF-R. Autoreactive B cells play a key role in the pathophysiology of ITP and wAIHA and are fueled by increased BAFF pathway activity; therefore, there is a strong rationale for investigating BAFF-R as a target in both diseases. Ianalumab is a fully human mAb that targets BAFF-R. It has a novel dual mechanism of action: blockade of BAFF-R-mediated signaling and potent depletion of B cells mediated by antibody-dependent cellular cytotoxicity. Ianalumab has shown favorable safety and encouraging efficacy in other indications, including SjS, SLE and CLL, and is a promising potential treatment for ITP and wAIHA. As ianalumab is being investigated in ongoing, blinded Phase III trials in ITP and wAIHA, we present the safety profile of ianalumab from completed Phase I/II studies in SjS, SLE or CLL. Methods: Data for patients with SjS were from a randomized Phase IIb trial (NCT02962895); patients (N=190) received subcutaneous (SC) placebo or ianalumab 5, 50 or 300 mg every 4 weeks (wks) for 24 wks. Patients then entered a second treatment period; patients in the ianalumab 5 or 50 mg arm continued the same dose, patients in the ianalumab 300 mg arm were randomized to continue the same dose or receive placebo, and patients in the placebo arm switched to ianalumab 150 mg. Safety data in patients with SLE were collected from a randomized Phase II trial (NCT03656562). Patients (N=67) received SC ianalumab 300 mg or placebo every 4 wks for 28 wks; patients in the placebo group could switch to ianalumab in a 20-wk open-label extension. Data for patients with CLL were from a Phase Ib trial (NCT03400176); patients (N=32) received intravenous (IV) ianalumab 0.3, 1, 3 or 9 mg/kg every 2 wks plus ibrutinib 420 mg once daily for up to 8 28-day cycles. Safety assessments included adverse events (AEs), serious AEs (SAEs) and AEs of special interest. Results: Ianalumab was safe and well tolerated in patients with SjS. Most AEs were mild or moderate in severity; severe AEs were reported in 5 patients receiving ianalumab. Three ianalumab-treated patients had SAEs; 2 SAEs in 1 patient (appendicitis and tubo-ovarian abscess) were considered related to ianalumab treatment. Frequency of infections/infestations was similar between patients treated with ianalumab and placebo. Discontinuation rates were low, with only 4 patients discontinuing ianalumab for non-serious AEs (infection [n=2], lymphopenia, local injection-site reaction [n=1 each]); no SAEs led to treatment discontinuation. Ianalumab continued to be well tolerated in the second treatment period. There were no dose-dependent differences in the rate of AEs except for injection-site reactions (mostly local and not severe). In ianalumab-treated patients with SLE, there were no serious infections during the blinded period; only 1 SAE (renal impairment) was reported. In the open-label extension, 4 SAEs were reported. No SAEs in the blinded period or open-label extension were considered treatment related. Most AEs in patients with CLL were Grade 1 or 2 and not correlated with dose. Twelve patients experienced Grade ≥3 AEs; decreased neutrophil count (n=5) was most common. No dose-limiting toxicities were reported. Discussion: Data from patients with SjS, SLE or CLL receiving ianalumab for up to 52 wks show it has a favorable safety profile. There was a low risk of infections irrespective of dosing regimen. No cases of hypogammaglobulinemia were clinically significant in any trial. New treatments are needed for ITP and wAIHA that have disease-modifying potential and allow patients to achieve and maintain high response rates after a short therapeutic course. Ianalumab has the potential to address this unmet need and is currently being assessed as an IV dose every 4 wks in Phase III trials as both a first- (VAYHIT1 [NCT05653349]) and second-line (VAYHIT2 [NCT05653219]) treatment for ITP and as a second-line treatment for wAIHA (VAYHIA [NCT05648968]).

Patient Preferences for Fixed Versus Treat-to-Progression Therapies in Chronic Lymphocytic Leukemia

The treatment landscape for chronic lymphocytic leukemia (CLL) has evolved in the last 8 years to include novel targeted therapies as alternatives to chemoimmunotherapy. These treatments vary in their benefit-risk profiles, mode and frequency of administration, and duration of treatment. Previous preference studies in CLL have evaluated preferences for treatment attributes such as efficacy, tolerability, and mode and frequency of administration. However, few have assessed preferences for treatment duration independently of mode of administration. This study sought to further understand the factors that influence patient preferences for treatment duration and to quantify the tradeoffs that patients were willing to accept to have a fixed duration therapy versus a treat-to-progression therapy. The study was conducted in 2 phases: (1) a qualitative phase of in-depth individual interviews with patients to identify the factors that influence patient preferences for treating until progression versus a fixed duration as well as the CLL treatment attributes that patients consider most important, and (2) a quantitative phase to estimate the tradeoffs that patients would accept among CLL treatment attributes. In the qualitative phase, the semistructured interview guide included open-ended questions and probes to understand perceptions of fixed-duration treatments compared with treat-to-progression regimens and to elicit a list of treatment attributes that influence CLL treatment preference. The results of the interviews informed the development of a web-based discrete-choice experiment (DCE) survey. Respondents to the online survey answered 12 DCE questions, each offering a choice between 2 hypothetical treatment profiles defined by 7 attributes with varying levels, including treatment duration, which included levels for treat to progression and fixed duration for either 6 or 12 months. Data were analyzed using a random-parameters logit model, and estimated preference weights were used to calculate the maximum acceptable risk (MAR) of treatment-related adverse events the average respondent would accept in exchange for a move from a treat-to-progression to a fixed-duration therapy. The MAR is estimated as the ratio of the relative importance of an improvement in an attribute to the relative importance of a unit change in the level of risk (i.e., tumor lysis syndrome (TLS), atrial fibrillation, or fatigue). Interviews were conducted with 20 adults with a self-reported diagnosis of CLL. The mean age was 59 years, and 55% of participants (n = 11) identified as female. The mean time since diagnosis of CLL was 3 years, and 70% of participants (n = 14) had received treatment for CLL. Overall, 93 treatment attributes were spontaneously reported, including treatment duration. When probed, treatment duration was reported by 50% of participants as “very important” in their treatment decision. Almost all participants (n = 17) preferred a treatment with a fixed duration compared with treat-to-progression, assuming each had the same efficacy. Table 1 reports patients' perceived benefits and drawbacks of fixed-duration versus treat-to-progression therapies. The DCE survey was completed by 229 adults with a self-reported diagnosis of CLL for at least 3 months. The mean age was 66 years, and nearly 60% of the sample identified as female (n = 136). About 60% of the sample (n = 138) was diagnosed 5 or more years ago, and 152 respondents (66%) had experience with treatment for CLL. The results of the preference analysis showed that respondents preferred a fixed duration of either 6 months or 12 months versus treat-to-progression and, based on the MAR estimates, were willing to accept the following levels of risks to have a fixed-duration treatment versus treat to progression: more than a 3% increased risk of TLS, 6%-7% increased risk of atrial fibrillation, and 21%-26% increased risk of fatigue (Table 2). Past research has shown efficacy as the most important factor, yet qualitative interview participants also identified treatment duration as an important factor in their decision when choosing a CLL therapy. This finding was confirmed by the quantitative preference study, which revealed a preference for fixed-duration therapies over treat-to-progression regardless of the timeframe (6 or 12 months). Results from this study can help inform shared decision-making when considering alternative therapies for CLL.

Counting Chromosome Abnormalities in Chronic Lymphocytic Leukemia: Comparison of Iscn 2020 and MDS International Working Group Guidelines for Establishing Karyotype Complexity

Karyotype complexity is established by enumerating the number of chromosome abnormalities present in a karyotype. In chronic lymphocytic leukemia (CLL), karyotype complexity, defined as ≥3 chromosome abnormalities, is an important prognostic indicator associated with poor outcomes. Additionally, stratifying patients (pts) by highly complex karyotypes (≥5 chromosome abnormalities) has shown significantly poorer outcomes compared to pts with 3-4 abnormalities (Baliakas, Blood, 2019). In principle, counting chromosome abnormalities is straightforward; however, in practice it can be nuanced, and the number of abnormalities counted for the same karyotype can differ across laboratories. A lack of standardized rules has resulted in variability in establishing complexity across CLL studies. We and others have proposed adapting the international working group on myelodysplastic syndrome (MDS) framework for use in CLL (Chun, Leuk Res, 2010; Jondreville, Am J Hematol, 2020). Additionally, a separate set of guidelines were published in the 2020 International System for Human Cytogenomic Nomenclature (ISCN). To investigate how different guidelines may impact patient categorization in correlative studies, we evaluated karyotype complexity using both guidelines in a previously reported, well-characterized, dataset with long term follow-up of patients with CLL treated with ibrutinib (Kittai, Blood, 2021). 456 patient karyotypes underwent expert review by two cytogeneticists. Discordant scores were discussed to reach consensus. Pts were categorized as not complex (0-2 abnormalities), complex (3-4), or highly complex (≥5) for each set of guidelines. The weighted kappa was used to evaluate agreement between the two guidelines. Progression free survival (PFS) from ibrutinib start and overall survival (OS) were evaluated by the Kaplan-Meier method. Univariable Cox regression model discrimination power for PFS and OS were evaluated by Uno's concordance statistic. MDS guidelines categorized 50.2% as not complex, 19.7% complex and 30.1% as highly complex; similarly, ISCN categorized 50.2% as not complex, 20.2% complex and 29.6% as highly complex. Complexity categories assigned by the two guidelines had a weighted kappa agreement of 0.88 (95% CI 0.84-0.91), with 11% of pts placed in discordant complexity categories. Albeit numbers of pts were small, evaluation of these discordant cases compared to the consensus concordant cases showed outcomes varied. Discordant cases categorized by the MDS guidelines as complex and by ISCN as highly complex showed longer OS compared to the concordant complex and highly complex cases, suggesting ISCN overrated complexity (Fig 1). Conversely, discordant cases categorized by MDS guidelines as highly complex while ISCN categorized as complex showed decreased OS compared to the concordant complex and highly complex cases, suggesting ISCN underrated complexity in these cases (Fig 2). PFS trended similarly for these groups. Both the MDS and the ISCN guidelines showed increased karyotype complexity was significantly associated with shorter PFS (both with p<0.0001) and OS (both with p<0.0001). Comparing the classifiers not complex, complex and highly complex for the entire cohort found no significant difference between the prediction power of the MDS and ISCN guidelines for PFS. For OS, the MDS guidelines had a statistically stronger prediction power compared to ISCN (p=0.0235). Further, when the poorest prognostic group (highly complex) was compared to all others, the MDS guidelines showed significantly stronger prediction power for both PFS (p=0.0359) and OS (p=0.0039) compared to ISCN. Although both sets of guidelines were prognostically significant, in this predominately relapsed/refractory CLL patient population receiving ibrutinib, the MDS guidelines performed more robustly when used as a categorical variable. Clinical trials incorporating karyotype complexity for prognostication, or for treatment stratification, should specify a set of guidelines to apply consistently across the trial. Given the importance of karyotype complexity, particularly with targeted therapies, further studies are needed to evaluate these two sets of guidelines across varying CLL populations to establish recommendations for assigning karyotype complexity uniformly in CLL.

Outcomes and Treatment Sequences of Therapies with BCL2- and BTK Inhibitors in Chronic Lymphocytic Leukemia (CLL): An Analysis of Patient Data within the German CLL Study Group (GCLLSG) Registry

Introduction: Targeted agents, in particular BCL2-inhibitors (BCL2i) and BTK-inhibitors (BTKi), have profoundly improved the outcome of patients (pts) with chronic lymphocytic leukemia (CLL) and are considered standard treatment. However, the transition from treatments evaluated within clinical trials into routine clinical practice can be complex. We here present real-world data from the German CLL Study Group (GCLLSG) registry and report outcomes and treatment sequences of pts treated with either BCL2i (venetoclax) or BTKi (acalabrutinib, ibrutinib, spebrutinib, tirabrutinib, or zanubrutinib) as first administered targeted agent. Methods: Pts from multiple German sites with confirmed diagnosis of CLL and with at least one documented first-line CLL treatment between July 1 st 2014 and January 30 th 2023 were included in this analysis. Pts participating in an active clinical trial were excluded. Treatments were categorized according to the backbone of therapy and pts were allocated to either the venetoclax (Ven) or the BTKi cohort depending on their first treatment with one of these substances. Time to next treatment (TTNT), event-free survival (EFS) and overall survival (OS) were analyzed using Kaplan-Meier methods. Results: A total of 274 pts were treated with a Ven-based regimen while 915 pts were treated with a BTKi as first therapy. Median observation time from the first documented treatment (irrespective if given as first-line or in a later therapy line) was 26 months (range 0 - 296) in the Ven cohort and 78 months (range 0-266) in the BTKi cohort. For the Ven cohort, median age at the time of first Ven treatment was 71 years (range 32 - 87) with 172 pts (62.8%) being male. Median CIRS score was 2 (range 0-13) and 12 pts (7.5%) had an ECOG performance status > 1. Among 57 pts with known molecular genetics, 32 pts (56.1%) had an unmutated IGHV status and 9 out of 51 evaluable pts (17.6%) had a deletion and/or mutation TP53. Applying the CLL-IPI, 11 out of 58 evaluable pts (19.0%) were in the very high-risk group. In the BTKi cohort, median age at time of first BTKi treatment was 72 (range 32 - 92), with 628 pts (68.6%) being male. Median CIRS score was 3 (range 0-17) and 45 pts (9.5%) had an ECOG performance status > 1. Among 162 pts with available data, 108 pts (66.7%) had an unmutated IGHV status and 59 out of 130 evaluable pts (45.4%) had a deletion and/or mutation TP53. Using the CLL-IPI, 46 out of 145 evaluable pts (31.7%) were in the very high-risk group. A total of 152 of 274 pts (55.5%) in the Ven cohort received Ven as first-line treatment (median number of prior treatments: 0, range 0-9) and 352 of 915 pts (38.5%) in the BTKi cohort received a BTKi as first-line treatment (median number of prior treatments: 1, range 0-12). Among all 222 administered treatments prior to first Ven and 1090 treatments administered prior to first BTKi, chemoimmunotherapy (70.3% and 68.3%, respectively) and chemotherapy (10.4% and 17.8%, respectively) were the most frequent. For subsequent therapy following the first documented Ven or BTKi treatment, most pts were treated with targeted agent regimens. In the Ven cohort, 37 subsequent treatments were documented in 26 pts, of which 10 (27.0%) were a Ven-containing regimen and 13 (35.1%) a BTKi-containing regimen. Within the BTKi cohort, 393 subsequent treatments were administered in 264 pts, of which 146 (37.2%) included Ven and 106 (27.0%) a BTKi Treatment sequences were visualized with a sankey-diagram ( Figure 1). Median EFS from the start of the first documented Ven treatment was 31.8 months, with an estimated 2-year EFS rate of 66.6%. Median TTNT was 63.7 months and median OS was 96.5 months with an estimated 2-year OS rate of 89.5%. When Ven was given as first-line treatment, the estimated 2-year OS rate was 93.6% versus 86.5% when given in a later therapy line for the first time. For the BTKi cohort, median EFS from start of first documented BTKi treatment was 23.2 months, with an estimated 2-year EFS rate of 48.6%. Median TTNT was 68.4 months and median OS was 85.9 months with an estimated 2-year OS rate of 84.9%. When BTKi were given as first-line treatment, the estimated 2-year OS rate was 92.3% versus 81.3% when given in a later therapy line for the first time. Conclusion: EFS, TTNT and OS were comparable following Ven versus BTKi-based therapy in pts documented in the GCLLSG registry between 2014 and 2023.

Endpoint Surrogacy in Chronic Lymphocytic Leukemia: A Pooled Analysis of the German CLL Study Group

Introduction Overall survival (OS) is generally considered the most patient-relevant endpoint in oncology trials, however, its implementation can entail several challenges in the context of chronic lymphocytic leukemia (CLL): 1) The competing survival risk due to comorbidities in the mostly elderly CLL patient population; 2) the high efficacy of targeted therapies warrant longer observation periods or extensive patient recruitment, which can be operationally challenging; 3) the interference with increasingly potent relapse therapies, which can neutralize differences of efficacy of first line regimen. Hence, surrogate endpoints, such as progression-free survival (PFS), overall response or complete response rate (ORR, CR) or minimal residual disease (MRD), are commonly used in CLL research. However, data supporting a correlation between surrogate endpoints and OS in the treatment for CLL is scarce. Here, we present a large, systematic analysis of over 4000 patients (pts) treated in studies of the German CLL Study Group (GCLLSG) to determine the adequacy of surrogate endpoints in clinical trials for pts with CLL. Methods We analysed twelve phase II and phase III trials of the GCLLSG spanning from 1999 to 2022. Only pts receiving first-line therapy were included in the analysis. To determine correlation, the Kaplan-Meier survival function was estimated for both OS and PFS for each study separately. From these functions, the probability of survival (OS and PFS) was estimated at different time points. These pairs of time points were then correlated across the studies, with each pair of probabilities weighted by the number of pts in each corresponding study using Spearman's correlation coefficients. Additionally, we compared OS according to MRD and iwCLL response status at end-of-treatment (for fixed-duration regimens) or end-of-induction-treatment (for MRD-guided treatments) (EO[I]T) using Kaplan-Meier functions calculated from the time point of MRD assessment in peripheral blood. EO(I)T ranged from month 9 to month 15 after treatment initiation, depending on the study protocol. Results The full analysis set comprised 4237 pts. The median observation time was 67 months. Median age was 64 years (range 27 - 90). 45.1% of pts were older than 65 years. 3159 pts (74.6%) had received chemo/chemoimmunotherapy (C/CIT), whereas 1078 (25.4%) were treated with targeted therapies. There were 2114 progressive disease (PD) events and 1211 deaths captured within the trial data. The Spearman's Rho of the estimated survival probabilities for the time points 12/24, 24/36, 36/48 and 60/72 months (PFS/OS) was 0.95, 0.96, 0.94 and 0.92, respectively ( Panel A). Focussing on pts with confirmed OS event (n=1211), the Spearman's Rho between PFS and OS in months was 0.66 for pts receiving chemo/chemoimmunotherapy and 0.88 for pts receiving targeted treatment. ORR and CR rate was 89.8% and 31.5%, respectively. Pts with a CR had a significantly longer PFS (HR 0.38, 95% CI 0.34-0.42, p <0.001) and OS (HR 0.39, 95% CI 0.33- 0.46, p <0.001) compared to pts with non-CR. MRD measurements at the EO(I)T were available for 2521 pts (59.5%), of which 39% were treated with targeted agents. Rates of undetectable (<10 -4), intermediate (≥10 -4 and <10 -2) and high MRD (≥10 -2) were 59.1%, 21.7%, 19.3%, respectively, across treatment groups. The median OS for undetectable and intermediate MRD was not reached and 60.7 months for high MRD status ( Panel B). The estimated 60-month OS was 84.6%, 71.3% and 51.1%, respectively. Accordingly, median PFS for uMRD was 61.8 months and 60-month PFS was 52.6%. In pts ≤65 years of age, median OS was not reached with undetectable or intermediate MRD and 70.5 months in pts with high MRD status. In pts >65 years, median OS was not reached for uMRD and was 75.2 and 58.8 months, respectively, for pts with intermediate or high MRD. The corresponding hazard ratios for high versus uMRD status at EO(I)T were 3.33 (95% CI 2.57 - 4.3) in older and 5.89 (4.15 - 8.35) in younger pts. Conclusion In this large analysis of 12 prospective CLL trials, we found a robust correlation between PFS and OS, thereby supporting the use of PFS as a surrogate endpoint in clinical studies. More importantly, the MRD status at EO(I)T was associated with OS across all treatment modalities, demonstrating its utility in identifying pts with potentially dismal outcomes and providing a rationale for MRD-guided treatment strategies.

Tislelizumab Plus Zanubrutinib in Patients with Richter Transformation: Primary Endpoint Analysis of the Prospective, Multi-Center, Phase-II RT1 Trial of the German CLL Study Group

Background Richter transformation (RT) describes the development of an aggressive lymphoma, in most cases a diffuse large B cell lymphoma (DLBCL) or Hodgkin's lymphoma (HL), in patients (pts) with chronic lymphocytic leukemia (CLL). Pts with RT have a dismal prognosis with chemoimmunotherapy (CIT) like R-CHOP, with overall response rates (ORR) of <40% and median overall survival (OS) of 6-8 months. Targeted therapies have the potential to improve outcomes, but few prospective studies have been run in this entity. Here we present the first data of the international phase-II RT1 trial, in which the PD1 inhibitor tislelizumab is combined with the next-generation BTK inhibitor zanubrutinib to treat pts with RT. Methods Pts with histologically confirmed RT (DLBCL or HL [HL only when not eligible for standard therapy]) and up to one prior line of RT-directed therapy were eligible. Each cycle (of 21 days) consisted of one infusion of 200 mg tislelizumab at day 1 and 160 mg zanubrutinib twice daily orally. Six cycles of induction were followed by 6 cycles of consolidation; pts with complete response (CR), partial response (PR) or stable disease (SD) continued maintenance until disease progression/non-tolerance. The primary endpoint was ORR after induction therapy for pts receiving ≥ 3 cycles according to the refined Lugano Classification with the aim to test the null hypothesis of ≤0.40. Secondary endpoints include duration of response (DOR), progression-free survival (PFS), OS and time to next treatment (TTNT). Results Of 59 enrolled pts, 48 pts received at least 3 cycles of treatment and comprised the analysis population according to the study protocol. Eleven pts stopped earlier due to withdrawal, death, PD or toxicity. Median observation time was 13.9 months (interquartile range 1.7-35.9). Median age was 67 years (range 45 - 82). 22 (45.8%) pts had ECOG 1 or higher. Median LDH at enrollment was 335 U/l. Sixteen (34.8%) pts had del(17p)/ TP53mut, 29 (70.7%) pts had unmutated IGHV. 25 (64.1%) pts had high or very high-risk CLL according to CLL-IPI, 11 (28.2%) intermediate and 3 (7.7%) low risk. Complex karyotype (≥3 aberrations) was detected in 16 (42.1%) pts. 46 (95.8%) pts had DLBCL-RT and 2 (4.2%) pts had HL-RT. In 26 (54.2%) pts, the RT was reported as clonally related to the CLL (22 [45.8%] unknown). Overall, 36 (75.0%) pts had received prior CLL-directed therapy, including CIT (25 pts) and targeted agents (32 pts) as well as prior allogeneic stem cell transplant (SCT) in 3 pts. 12 (25.0%) pts had treatment-naive CLL. Thirteen pts (27.1%) had received prior RT-directed therapy, including CHOP-like regimens and BTK inhibitors. 35 (72.8%) pts had not received prior RT-directed therapy. Twenty-eight of 48 pts responded to induction therapy resulting in an ORR of 58.3% (95% CI, 0.43-0.72), including 9 (18.8%) CR and 19 (39.6%) PR, meeting the study's primary endpoint (p=0.008) (Fig A). SD was reported in 6 (12.5%) pts, 11 (22.9%) pts had progressive disease (PD), three had missing assessment due to PD. The median DOR was not reached, the 6-month DOR rate was 70.6%. Median PFS was 10 months (95% CI 3.8 -16.3) with a 12-month rate of 46.9% (Fig B). Median OS was not reached (12-month OS rate 74.7%). Median TTNT was not reached (12-month TTNT rate 56.2%). Post-protocol treatment included CIT in 21 cases (48.8%), radiation in 1 case (2.3%), BTK/BCL2 inhibition in 7 (16.3%) cases and 8 (18.6%) allogeneic SCT. SCT was conducted as consolidation in two pts with PR, and as salvage treatment for 5 pts with SD/PD (one missing response). Overall, 606 adverse events (AEs) were reported, of which 173 (28.5%) were ≥ grade 3 and 129 (21.3%) were serious AE. Of the 29 pts who discontinued study treatment, three were due to toxicity, the rest for PD and/or start of next line of therapy. The most frequent AEs were infections and infestations (17.7%), including Covid-19, pneumonia and urinary tract infections, gastrointestinal disorders (12.7%), including diarrhea and nausea, and blood and lymphatic system disorders (11.2%), including anemia, neutropenia and thrombocytopenia. Conclusions Combined checkpoint and BTK inhibition by tislelizumab plus zanubrutinib is an effective and well-tolerated treatment strategy for pts with RT. Responses are durable and overall survival in the RT1 study is encouraging given the otherwise poor prognosis of RT. Translational studies are ongoing to identify predictors of response to checkpoint inhibition in RT.

Treatment with idelalisib in patients with chronic lymphocytic leukemia – real world data from the registry of the German CLL Study Group

Idelalisib in combination with rituximab is an efficacious treatment for patients suffering from chronic lymphocytic leukemia (CLL) with known limitations due to toxicities. However, the benefit after prior Bruton tyrosine kinase inhibitor (BTKi) therapy remains unclear. For this analysis, 81 patients included in a non-interventional registry study of the German CLL study group (registered at www.clinicaltrials.gov as # NCT02863692) meeting the predefined criteria of a confirmed diagnosis of CLL and being treated with idelalisib containing regimens outside clinical trials were considered. 11 patients were treatment naïve (13.6%) and 70 patients (86.4%) pretreated. Patients had median of one prior therapy line (range 0–11). Median treatment duration with idelalisib was 5.1 months (range 0–55.0 months). Of 58 patients with documented treatment outcome, 39 responded to idelalisib containing therapy (67.2%). Patients treated with the BTKi ibrutinib as last prior treatment prior to idelalisib responded in 71.4% compared to a response rate of 61.9% in patients without prior ibrutinib. Median event free survival (EFS) was 15.9 months with a 16 versus 14 months EFS in patients with ibrutinib as last prior treatment or not, respectively. Median overall survival was 46.6 months. In conclusion, treatment with idelalisib appears to have a valuable impact in patients being refractory to prior ibrutinib therapy even though there are limitations in our analysis due to the low number of patients included.

COMORBIDITY & CLL DIFFERENT SCORES – DIFFERENT RESULTS

Introduction: Due to the implementation of the outpatient centres, our network provides care for 50% of the patients with hematological diseases in Vienna. Here we analyse the influence of our own CIRS Score on survival of CLL patients. Additionally we focus on renal comorbidities. Methods: 1150 CLL patients who were diagnosed, observed and treated between 2011 and 2021 within our haematological network were analysed. Study data was generated by reviewing electronic and paper medical records. Comorbidities were assessed with the conventional CIRS Score including utilizing our own Version where we did not focus on severity to detect more patients. We differentiated between 2 groups: low risk pts. without any or with just one existing co-morbidity and high-risk patients with 2 or more organ systems involved, regardless of the severity. Moreover we demonstrate a Composite Comorbidity Score “Severe4” with <3 or >3 points, validated in 4 Organ Systems as shown on ASH 2022 by Shouse et al. and the cut-off Score of 6 points as used by the German CLL Study Group. Furthermore the Cockcroft-Gault formula (GFR). Results: 55% of the patients were male. The median age at diagnosis was 68 years. IGHV was mutated in 53% of patients, del(17p) was present in 7,4% of patients. 27% of our 1150 patients received CLL specific treatment. As shown in Figure 1 our own interpretation of the CIRS-Score was able to identify 625 pts. classified as high risk with a significant lower OS. Regardless of the Severity, the involvement of 2 or more organ systems was more relevant than just one. Similarly to the Severe4 Score we could also show a significant OS difference in 167 pts. using a cut-off Score of 3 Points in at least one category. A highly significant OS difference can also be shown in 245 pts when using the established cut-off Score of 6 points. Emphasizing the weakness of the CIRS Score in detecting renal comorbidity due to the high threshold concerning the Serum Creatinine of 1,5 mg/dl we were able to demonstrate that 144 of our pts. had a GFR of <60 mL/min at time of diagnosis, which reflects Stage 3 CKD, resulting in a significant lower OS. When using the CIRS Score alone only 31 pts. would have been classified as renal impaired. Keyword: chronic lymphocytic leukemia (CLL) No conflicts of interests pertinent to the abstract.

A History of Targeted Therapy Development and Progress in Novel-Novel Combinations for Chronic Lymphocytic Leukemia (CLL).

Over the last 10 years, the traditional treatment paradigms for CLL have been upended as the use of traditional chemoimmunotherapy regimens has declined in favor of novel targeted therapies. Targeted therapies have become the new standard of care in CLL given their superior progression-free survival (and overall survival, in some cases) when compared with chemoimmunotherapy, as well as their improved toxicity profiles. Targeted agents are FDA approved for the treatment of CLL including ibrutinib, acalabrutinib, zanubrutinib, and venetoclax. Importantly, as opposed to traditional chemotherapy regimens, the benefits of these targeted therapies appear to be consistent regardless of high-risk mutational status. In this review, we discuss the pivotal CLL studies of the last decade and the data supporting doublet and triplet novel-novel combinations. We explore the use of new surrogate end points for PFS/OS in targeted therapies such as undetectable minimal residual disease (uMRD) and their potential role in minimizing toxicity by permitting earlier treatment discontinuation. We also highlight areas that warrant further exploration and future studies that may help address some of these key questions.

Proteogenomic Analysis of SF3B1mut CLL Reveals Altered Protein Signaling Cascades and Metabolomic Pathways

Although mutation in the U2 snRNP spliceosome component SF3B1 has been appreciated as a common driver of chronic lymphocytic leukemia (CLL) for over a decade, we still know little of its mechanism of action. Prior transcriptome-based characterizations have demonstrated widespread changes in alternative splicing in SF3B1mut CLL. We hypothesized that protein level characterization could potentially yield fresh insights on alternative expression of proteins and on the impact of altered signaling pathways in patients bearing this mutation. To this end, we assembled CLL specimens from a cohort of 105 patients enrolled on the German CLL Study Group (GCLLSG) trials CLL8 and CLL11, collected prior to therapy. Of these, 52 patients expressed SF3B1wt and 53 patients harbored clonal SF3B1mut (CCF>0.8); 25 of these were the K700E mutation. Additionally, we characterized these samples by WES (available for 97 patients) and RNA-Seq (available for 96 patients). Proteomes and phosphoproteomes from all 105 samples were analyzed using a multiplexed isobaric-tagged (TMT) high resolution mass spectrometry (MS) workflow. Briefly, cells were lysed and proteins were digested into peptides with LysC and trypsin. Protein yield per sample ranged from 20 μg to 300 μg. Peptides were labeled with TMT-11 reagent and combined into 11 multiplexes while accounting for input material availability and randomizing for SF3B1 mutation, IGHV status and ATM/TP53 mutation. To enable cross TMT-plex comparison, we constructed a common reference consisting of 40 samples with highest protein amount and an equal representation between SF3B1wt and SF3B1mut samples. Each plex was fractionated into 24 fractions by high pH reversed phase separation and 500 ng protein per fraction were analyzed by MS (Orbitrap Exploris, Thermo Fisher Scientific). For phosphoproteome analysis, peptides were pooled into 12 fractions and enriched by IMAC prior to MS analysis. Spectra were analyzed by Spectrum Mill v7.1 (Broad Institute) and searched against a human reference proteome database supplemented with non-canonical ORFs supported by Riboseq evidence of translation (nuORFdb v1.0). Patient specific proteins containing single amino variants, indels, and splice isoforms, derived from WES and RNA-Seq, were included in the search database. We obtained a total of 10,419 proteins, with a median of 8,882 proteins per TMT-plex and 6,969 proteins that were quantifiable across all samples. The phosphoproteome analysis identified 32,410 sites; while each TMT-plex yielded a median of 19,380 phosphosites per plex, 5,810 were quantifiable across all patients. We integrated proteome data with patient specific mutation calls, RNA-Seq and copy number alteration (CNA) data using Panoply, a cloud-based platform for proteogenomic data analysis (Broad Institute). We confirmed the impact of chr11 and chr13 deletion as well as chr12 amplification on protein expression. We readily identified peptides spanning the SF3B1mut amino acid regions in corresponding samples. By unsupervised clustering, samples were grouped by SF3B1 status on the CNA and proteome level, a trend not observed based on RNA or phosphoproteome analysis. Gene set enrichment analysis revealed enrichment of TNFA and PI3K signaling in the SF3B1wt clusters, while the SF3B1mut associated clusters showed a strong association with metabolism changes. This analysis was highly concordant with a parallel interrogation comparing proteins expressed in SF3B1wt CLL versus SF3B1mut CLL. Here 1,955 proteins and 1,737 phosphosites were significantly altered (5% FDR). Proteins involved in B cell receptor and MAP kinase signaling were highly expressed in SF3B1wt samples, while again amino acid metabolism pathways as well as oxidative phosphorylation were upregulated in SF3B1mut samples. To more deeply investigate the role of metabolism pathways in SF3B1mut CLL, we subjected 6 independent CLL samples to MS-based metabolomics analysis. Inosine (P=0.029) and guanosine (P=0.047) were lower in the SF3B1mut extracts (P=0.001), while acylcarnitines (P=0.0004) were increased, indicating fatty acid oxidation as a preferred energy source in SF3B1mut CLL. Altogether, our study reveals fresh evidence of altered energy metabolism in SF3B1mut CLL. Ongoing studies focus on defining the mechanism by which SF3B1mut exerts these metabolic changes and novel therapeutic opportunities afforded by these findings.

Characteristics of Rare Germline ATM Variants in Chronic Lymphocytic Leukemia (CLL)

Background The use of next-generation sequencing in the clinic has increasingly uncovered new variants of unknown significance (VUS), which are often ignored clinically. Ataxia telangiectasia mutated (ATM) is a tumor-suppressor gene that has a critical role in DNA double-strand break repair and has been implicated in the pathogenesis of CLL. In this study, we assessed rare germline ATM variants including VUS using a novel variant calling algorithm, evaluated their impact on the clinical characteristics of CLL and functionally assessed their effect on ATM kinase activity. Methods We evaluated the entire ATM coding region via germline whole exome sequencing from 885 CLL patients representing three different previously reported cohorts (327 DFCI; 276 German CLL Study Group, and 282 ICGC), along with a cohort of ancestry matched controls. We utilized DeepVariant (Poplin et al Nature Biotech 2018) to detect germline variants, followed by a standardized quality control workflow (AlDubayan et al JAMA 2020). All individuals harbored ATM germline variants. Variants were annotated using dbSNP, Clinvar and Varsome, and variants with GnomAD population allele frequency (AF) < 1% were defined as rare. For clinical characterization of CLL patients with rare germline ATM missense variants, we focused on the DFCI cohort. For functional analysis, we evaluated ATM kinase activity by ATM and KAP1 phosphorylation in response to 1 Gy gamma-irradiation using flow cytometry in primary CLL samples with/without rare germline ATM missense variants. We also generated CRISPR/Cas9 engineered cell lines to evaluate the function of ATM p.L2307F which was frequent in our CLL cohort and which we previously demonstrated as enriched in CLL, yet generally annotated as a VUS. Results We identified a total of 130 ATM germline variants among the CLL cases, including 87 missense, and 9 splice site alterations. 80 of 87 missense variants were rare and 59 were annotated as VUS. The top 5 most frequent rare germline missense variants with high odds ratio for association with CLL were ATM p.L2332P (AF cases vs controls, 0.328% vs 0.0188%, OR 17.46), p.L2307F (OR 16.88), p.F763L (OR 18.89), p.Y1442H (OR 35.39) and p.S99G (OR 16.28). In 63 DFCI patients with rare missense variants, the median age at diagnosis was 55 (31-82), 21.7% had a familial history of CLL and 29% had del(11q). Phosphorylation of ATM and KAP1 (pATM, pKAP1) in response to gamma-irradiation was reduced in 12 patients assessed (8 rare and 4 non-rare) with germline ATM variants (ATM variant alone group) compared to those without any somatic or germline ATM aberrancy (WT group) (P<0.01). Similar results were seen when the analysis was restricted to the 8 CLL patients with rare ATM variants. Phosphorylation levels were also significantly lower in 17 patients with rare germline ATM variants with concurrent del(11q) (del 11q + ATM variant group) compared to 13 of those with del(11q) alone (del 11q group) (P<0.01) (Figure 1). Altogether, these results suggest that germline ATM variants have a functional impact on DNA damage response in primary CLL cells. We previously identified the VUS variant ATM p.L2307F as particularly enriched among CLL patients (Tiao G et al., Leukemia. 2017). We therefore generated ATM L2307F knock-in NALM-6 cell lines via CRISPR/Cas9. We generated 7 cell lines including ATM wild type (+/+), ATM monoallelic mutated (+/L2307F), ATM monoallelic knocked out (+/-), ATM monoallelic mutated with knock out of the other allele (-/L2307F), and ATM biallelic knockout (-/-). The induction of both pATM and pKAP1 levels by gamma irradiation was reduced in proportion to ATM mutational status as shown in Figure 2. Conclusions We demonstrate that rare ATM germline variants are common in CLL and affect the DNA damage response in primary CLL cells when controlling for somatic 11q deletion. We further characterize the p.L2307F variant using CRISPR and demonstrate that this allele represents a functional hypomorph in cell-based assays. Further studies are needed to assess the impact of each individual rare germline variant on the function of ATM, CLL pathogenesis and the inherited risk of CLL. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Fludarabine, Cyclophosphamide and Rituximab (FCR) As First Line Treatment in Patients with Chronic Lymphocytic Leukemia (CLL): A Long-Term Analysis of the German CLL Study Group (GCLLSG) Registry

Introduction: Large randomized trials investigating targeted agents have resulted in a markedly improved progression-free survival (PFS) when compared to standard chemoimmunotherapy. However, for small subgroups with a favorable risk profile, overall survival was not consistently superior for targeted agents. Long-term follow up on FCR suggested that favorable risk patients (pts) benefit significantly from this therapy and can expect an exceptionally long PFS. In addition, targeted first line (1L) therapies for favorable risk pts are reimbursed only in a minority of countries. Here, we report long-term efficacy data of FCR treatment obtained from pts documented within the GCLLSG registry. Methods: Documentation within the GCLLSG registry started in 2013 and included both pts from prospective trials (29.7%), most importantly of the CLL8 trial (22.6%) and CLL10 trial (62.1%) as well as retrospective data. Pts with confirmed diagnosis of CLL who received FCR as 1L treatment were included. Patient characteristics were reported in median, interquartile ranges or frequencies and percentages. Response was reported as response or non-response. Event-free survival (EFS), which was defined as the time from start of (first) FCR treatment to disease progression, initiation of subsequent treatment for CLL or death. EFS was chosen as an endpoint for this report due to concerns that PFS might give unrealistically good results due to lack of documentation. However, PFS and TTNT data will be presented at the meeting. Overall survival was calculated from initiation of 1L treatment for all pts who received 1L treatment after 2013. Hazard ratios (HR) and 95% confidence intervals (CI) were calculated by Cox proportional hazards regression. Variables that were significantly associated with EFS in univariate analyses (significance level was set at 5%) were considered as candidates for the multivariate modelling to identify independent prognostic risk factors. Results: As of September 2021, a total of 5922 pts were enrolled in the registry of which 417 received FCR as 1L treatment between 2003 and 2021. 223 pts (53.5%) received treatment before 2013 and 194 (46.5%) after 2013. Median observation time from 1L treatment was 95.8 (interquartile range 58.7-126.8) months. Patient characteristics showed a median age of 58 (range 24-83) years. Unmutated IGHV (U-CLL) was present in 61.4% of the pts. Regarding genomic aberrations based on the hierarchical model, del(17p) was found in 1.9%, del(11q) in 15.2%, trisomy12 in 15.5%, mono-allelic del(13q) in 34.3% and none of these changes in 33.0% of the pts. 88.8% of the pts responded to first line therapy, 11.2% were documented as non-responder. Median EFS was 71.1 month, for 188 pts no event was documented at the time point of data cut-off. Age group (≤ 65 years, >65 years), del(11q), IGHV mutational status, TP53 status, and serum thymidine kinase (TK) were associated with EFS in univariate analyses. Overall, 116 pts with 67 events were included in the final multivariable model for EFS. U-CLL (HR 1.949 (95%CI: 1.170-3.247; p=0.010) and TK >10 U/l (HR 2.107 (95%CI: 1.153-3.849; p=0.015) were independent adverse prognostic risk factors for EFS. When low TK and M-CLL were combined, the median EFS was not reached, with a 5-year survival rate of 95.8%. In contrast, median EFS was 84.1 months for pts with U-CLL and high TK. In U-CLL pts, median EFS was 87.0 months for pts with TK ≤ 10 U/l, and 58.9 months for pts with TK > 10 U/l. Median overall survival was not reached with a 5-year survival rate of 92.7%. Conclusion: Within this cohort of pts with a long follow up in the GCLLSG registry, FCR treatment was associated with a long EFS especially in pts with a low risk profile exhibiting a mutated IGHV status und low TK. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Abstract IA26: Understanding CLL and Richter’s syndrome biology through mouse models of human genetics

Abstract Large-scale human next generation sequencing studies have progressively defined the complex genetic landscape of chronic lymphocytic leukemia (CLL) and Richter’s syndrome (RS), yet the difficulties in rapid manipulation of primary CLL cells and the limited availability of cell lines have challenged studies aimed at interrogating the function of the so-called disease “drivers”, defined in human studies through statistical inference. Mouse models represent powerful tools to study mechanisms of normal and malignant B cell biology. Conditional knock-in/knock-out systems have allowed B-cell restricted modeling of CLL genetic lesions, with MDR mice being the first example of a genetically-faithful CLL model, recapitulating the most common genomic aberration of human disease, del(13q). Since then, a series of models generated by our group and others have allowed to uncover key functional changes underlying disease onset and transformation, including in the context of combined mutation in the splicing factor Sf3b1 and the DNA damage response gene Atm, sole mutation in the B cell developmental factor Ikzf3, the nuclear export receptor Xpo1, the ribosomal protein Rps15, and the DNA methyltransferase Dnmt3a (the latter not a CLL lesion per se, but recapitulating disordered methylation patterns typical of CLL). The CRISPR-Cas9 gene editing technology has further enabled B-cell restricted introduction of multiple combinatorial loss-of-function lesions, allowing the generation of faithful models not only of CLL but, importantly, of transformation into RS. In this presentation, I will describe the application of mouse models to studies of CLL and RS pathogenesis and treatment, with a focus on key aspects uncovered through the characterization of genetically- inspired mouse lines of recurrent disease drivers. I will provide a perspective on how these novel models combined with new technology allowed the dissection of mechanisms of disease evolution and response to therapy with greater depth than previously possible, representing valuable platforms for functional genomic analyses. Citation Format: Elisa ten Hacken. Understanding CLL and Richter’s syndrome biology through mouse models of human genetics [abstract]. In: Proceedings of the Third AACR International Meeting: Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2022 Jun 23-26; Boston, MA. Philadelphia (PA): AACR; Blood Cancer Discov 2022;3(5_Suppl):Abstract nr IA26.

P667: LONG-TERM EFFICACY OF ACALABRUTINIB-BASED REGIMENS IN PATIENTS WITH CHRONIC LYMPHOCYTIC LEUKEMIA AND HIGHER-RISK GENOMIC FEATURES: POOLED ANALYSIS OF CLINICAL TRIAL DATA

Background: Chronic lymphocytic leukemia (CLL) has a highly variable disease course; patients (pts) with higher-risk genomic features typically have poor response to chemoimmunotherapy, and pts with del(17p) also have short, suboptimal response to venetoclax + obinutuzumab. Thus, novel agents such as Bruton tyrosine kinase inhibitors (BTKi), including the highly selective covalent BTKi acalabrutinib (A), are preferred treatments in this higher-risk population based on demonstrated effectiveness in higher-risk subgroups in individual studies. Aims: To assess long-term efficacy of A-based regimens in pts with treatment-naive (TN) or relapsed/refractory (R/R) CLL and higher-risk genomic features. Methods: Data were pooled from CLL pts with higher-risk genomic features treated with A ± obinutuzumab (O) in 3 clinical studies in TN CLL (ELEVATE-TN, CL-001 [TN cohort], CL-003) and 3 in R/R CLL (ASCEND, ELEVATE-RR, CL-001 [R/R cohort]). Higher risk was defined as del(17p) and/or TP53 mutation [del(17p)/TP53m], unmutated IGHV (uIGHV), or complex karyotype (CK, ≥3 chromosomal abnormalities). Efficacy analyses (progression-free survival [PFS], overall survival [OS], response rates) focus on del(17p)/TP53m and uIGHV; safety analyses also include pts with CK. Results: 801 pts (TN 313; R/R 488, median 2 prior therapy lines [range 1–10]) were included; 64 (20%) TN and 219 (45%) R/R pts had del(17p)/TP53m, among whom 44 (69%) and 170 (78%) also had uIGHV. Overall, 288 (92%) TN and 425 (87%) R/R pts had uIGHV and 47 (15%) and 160 (33%) had CK. Median pt age was 68 (TN) and 66 (R/R) y. For A-based regimens combined (A and A+O), overall response rate (ORR) was 91% (n=58/64; complete response [CR]: 20% [n=13/64]) in TN pts with del(17p)/TP53m and 96% (n=276/288; CR 16% [n=47/288) in TN pts with uIGHV. At 47.3 mo median follow-up (range 1.0–82.0), median PFS was not reached (NR) in TN pts with del(17p)/TP53m with A-based regimens; PFS rates at 48 mo suggest similar efficacy with A and A+O in TN pts with del(17p)/TP53m (76% and 77%, respectively) (Fig 1A). Median PFS was NR among TN pts with uIGHV with both A and A+O; PFS rates at 48 mo with A and A+O were 84% and 86%, respectively (Fig 1B). Median OS was NR with A and A+O in TN pts with del(17p)/TP53m or uIGHV; OS rates at 48 mo were similar for A and A+O in pts with del(17p)/TP53m (89% for both) and in pts with uIGHV (92% and 95%). In the R/R cohort (A monotherapy only), ORR was 86% (n=184/214; CR 5% [n=11/214]) in pts with del(17p)/TP53m and 87% (n=356/408; CR 7% [n=29/408]) in pts with uIGHV. At 44.0 mo median follow-up (range 0–88.0), median PFS was 38.6 mo and 46.9 mo and PFS rates at 36 mo were 54% and 65% in pts with del(17p)/TP53m and uIGHV, respectively (Fig 1C). Median OS was 60.6 mo and NR in pts with del(17p)/TP53m and uIGHV, respectively; OS rates at 36 mo were 73% and 82%. Among all higher-risk pts, incidences of grade ≥3 treatment-related AEs were 30% (TN) and 35% (R/R). Discontinuations due to treatment-related AEs occurred in 4% (TN) and 6% (R/R) of pts; 60% and 27%, respectively, remained on treatment at data cutoff. Image:Summary/Conclusion: In this pooled analysis of clinical trial data in 801 CLL pts with higher-risk genomic features, efficacy of A-based regimens led to high PFS and OS rates at a median follow-up of nearly 4 y. The safety profile in this analysis was similar to the overall safety profile of acalabrutinib. Our results demonstrate the long-term benefit of A-based regimens in CLL pts with higher-risk genomic features, regardless of line of therapy.