Shanafelt, Tait

Abstract Introduction: Chronic lymphocytic leukemia (CLL) is marked by impaired T cell responses that limit the efficacy of immunotherapies. CD19-directed chimeric antigen receptor T cell therapy (CART19) received FDA approval to treat CLL in 2024. However, itsefficacy in CLL patients remains low, with durable remission rate of ~20%. Ibrutinib, a Bruton tyrosine kinase inhibitor, has shown synergy with CART19 in preclinical and early clinical studies. We aimed to study the impact of ibrutinib on CART19 utilizing clinical samples from the pivotal clinical trial E1912 (NCT02048813) of ibrutinib/rituximab (IR) vs fludarabine/cyclophosphamide/rituximab (FCR). This trial led to the FDA approval of first line ibrutinib in CLL. Methods: We generated CART19 incorporating either CD28 (28ζ) or 41BB (BBζ) costimulatory domains from T cells isolated from patients with CLL treated with IR or FCR on the E1912 trial. IR samples were collected at baseline (pretreatment), 6- or 12-months during treatment, or at progression. FCR samples were collected at baseline, 15-18 months during treatment, or at progression. For animal experiments, NOD-SCID-γ-/- (NSG) mice were intravenously (i.v.) engrafted with 1×10⁶ luciferase⁺ CD19⁺ JeKo-1 cells. Upon tumor establishment (~10⁸ photons/sec by bioluminescence imaging), mice were randomized and treated i.v. with 1×10⁶ 28ζ or BBζ derived from longitudinal IR- or FCR-treated patient samples. To further assess the impact of prolonged ibrutinib exposure on CART19 function, we chronically stimulated healthy donor-derived 28ζ or BBζ in vitro for 22 days with JeKo-1 cells and 100nM ibrutinib or vehicle control and then tested CART19 functions. Finally, we performed single cell RNA sequencing (scRNAseq) of longitudinal patient T cell samples treated with IR. Results: Ex vivo T cell expansion was significantly enhanced following IR compared to FCR. BBζ generated from IR-treated patients after 6 months of ibrutinib exhibited enhanced in vitro expansion (p=0.016) and antigen-specific proliferation (p=0.038) in the presence of JeKo-1 cells. This corresponded with significantly improved antitumor efficacy and prolonged survival in vivo compared to cells generated at disease progression (p=0.017). In contrast, 28ζ showed no significant differences in efficacy across IR studied treatment timepoints. BBζ from FCR-treated patients did not show differences in tumor control based on treatment timepoints but demonstrated shorter survival compared to BBζ from IR-treated patients. 28ζ from FCR-treated patients generated at disease progression demonstrated significantly reduced tumor control and shorter survival compared to 28ζ produced at baseline (p=0.0002), indicating detrimental effects of FCR treatment on the therapeutic potential of CART products. To further investigate the impact of prolonged ibrutinib exposure, we utilized in vitro models of CART exhaustion. We found that, after repeated antigen stimulation with JeKo-1 cells, ibrutinib-treated BBζ had significantly increased antigen-specific degranulation (p=0.005), and cytokine production (IL-2, p=0.045; TNF-α, p=0.003; IFN-γ, p=0.001), compared to BBζ treated with vehicle control. In contrast, 28ζ did not show improved degranulation or cytokine production in cocultures with ibrutinib. Prolonged ibrutinib exposure led to downregulation of FOXO3, a transcription factor associated with T cell exhaustion which has been shown to cause dysfunction specifically in BBζ. These results further indicate that ibrutinib may be more beneficial to BBζ. Indeed, scRNAseq revealed that after ≥ 6 months of ibrutinib, T cells showed downregulation of inflammatory, oxidative stress, and apoptosis pathways, paralleled by modest activation of MYC and E2F targets. However, at disease progression, T cells under ibrutinib treatment displayed marked upregulation of interferon signaling, inflammation responses, and stress-related programs, indicating a shift toward immune reactivation or dysfunction.Conclusion: Our study shows that BBζ, but not 28ζ, generated 6 months during ibrutinib treatment has superior antitumor activity and persistence compared to similar later timepoints or FCR-treated counterparts. Prolonged ibrutinib exposure enhances BBζ function by modulating stress and apoptotic pathways. These findings highlight the importance of pre-CART treatment timing and support ibrutinib as a potential enhancer of CART cell fitness in CLL.

Abstract Background: The ECOG-ACRIN randomized phase 3 clinical trial E1912 established ibrutinib-rituximab (IR) as the standard of care for CLL patients <= 70 years in the frontline setting by improving the progression free (PFS) and overall survival (OS) compared to previous standard chemo-immunotherapy with fludarabine, cyclophosphamide and rituximab (FCR) (Shanafelt et al. NEJM 2019). Quality of life (QOL) was an important secondary endpoint on this trial comparing continuous (IR) vs time-limited (FCR) therapy due to the differing profile of treatment-related toxicities over the short and long-term. E1912 patient-reported outcomes (PROs) results are presented here. Methods: Patients enrolled on E1912 completed the Functional Assessment of Cancer Therapy-General (FACT-G) and Leukemia subscale at randomization (baseline), 3, 6, and 12 months post-randomization, and every 6 months for 2 years regardless of disease status. The primary outcome was the 31-item FACT-Leukemia Trial Outcome Index (FACT-Leu TOI), calculated by summing items from the FACT-G physical wellbeing (PWB), functional well-being (FWB), and leukemia subscales (score range: 0-124). The primary endpoint was defined as the difference in FACT-Leu TOI change scores from randomization to 12 months (at response evaluation) between patients treated with continuous therapy Arm A (IR) vs time-limited therapy Arm B (FCR). Mean change scores from baseline to each time point were calculated using all cases with data at baseline and the corresponding time point. Comparisons between treatment arms were performed using two-sample t tests. Linear mixed effects models were used to estimate the trajectories of PRO scores. Results: PRO data was extracted on 3/8/2021. PRO data was provided at baseline and 12 months for 233/354 (65.8%) patients on the IR arm and 118/175 (67.4%) patients on the FCR arm. At enrollment, there were no significant differences in the baseline FACT-Leu TOI scores (mean ± SE) between the two arms: IR (93.27 ± 1.03) vs FCR (92.68 ± 1.38; p=0.73). The FACT-Leu TOI score improved from baseline to 12 months in both arms (Table 1 and Figure 1). The change scores from baseline to 12 months, the primary outcome, were not significantly different between IR (7.59 ± 1.09) vs FCR (8.22 ± 1.44; p=0.73). Change in FACT-Leu TOI from baseline to 3 months was 5.77 ± 0.77 and 4.06 ± 1.18 (p=0.22); and from baseline to 6 months was 6.87 ± 0.87 and 8.01 ± 1.44 (p=0.50) in the IR and FCR arms, respectively. After the first 6 months of treatment, the improvement in FACT-Leu TOI scores was maintained in both treatment arms. There was no significant difference in total FACT-Leu TOI score in the continuous therapy (IR) vs time-limited therapy (FCR) arms over the first 36 months post registration. Analysis of FACT subscales showed PWB improved in the IR arm (0.37 ± 0.22) and declined in the FCR arm (-0.92 ± 0.39) from baseline to 3 months (p=0.004 for difference in PWB change between arms) but there was no significant difference between the two arms for change in PWB scores from baseline to 6 and 12 months. There were no significant differences between the two arms for change in FWB and FACT-Leu subscales from baseline to 3, 6 and 12 months. Conclusions: QOL improves in CLL patients age 70 and younger when treated in the frontline setting with IR or FCR. The improvement in QOL is maintained during continuous therapy with ibrutinib and there is no significant decline in QOL over time. Given the improvement in PFS and OS with IR over FCR seen in E1912, the results of this QOL analysis support the use of frontline ibrutinib in previously untreated younger CLL patients. Acknowledgement: This study was conducted by the ECOG-ACRIN Cancer Research Group (Peter J. O'Dwyer, MD and Mitchell D. Schnall, MD, PhD, Group Co-Chairs) and supported by the National Cancer Institute of the National Institutes of Health under award numbers: U10CA180794, U10CA180820, UG1CA189863, UG1CA190140, UG1CA232760, UG1CA233180, UG1CA233230, UG1CA233253, UG1CA233277, UG1CA233290, UG1CA233339, UG1CA189859, UG1CA233332. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. Figure 1 Figure 1. Disclosures Kay: Oncotracker: Membership on an entity's Board of Directors or advisory committees; CytomX Therapeutics: Membership on an entity's Board of Directors or advisory committees; Pharmacyclics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Tolero Pharmaceuticals: Research Funding; Acerta Pharma: Research Funding; Behring: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees, Research Funding; TG Therapeutics: Research Funding; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Juno Therapeutics: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Agios Pharm: Membership on an entity's Board of Directors or advisory committees; Rigel: Membership on an entity's Board of Directors or advisory committees; Dava Oncology: Membership on an entity's Board of Directors or advisory committees; Morpho-sys: Membership on an entity's Board of Directors or advisory committees; Targeted Oncology: Membership on an entity's Board of Directors or advisory committees; MEI Pharma: Research Funding; Sunesis: Research Funding; Genentech: Research Funding; Bristol Meyer Squib: Membership on an entity's Board of Directors or advisory committees, Research Funding. O'Brien: Amgen, Astellas, Celgene, GlaxoSmithKline, Janssen Oncology, Aptose Biosciences Inc., Vaniam Group LLC, AbbVie, Alexion, Verastem, Juno Therapeutics, Vida Ventures, Autolus, Johnson and Johnson, Merck, Bristol Myers Squibb, NOVA Research Company, Eli Lill: Consultancy; Kite, Regeneron, Acerta, Caribou, Gilead, Pharmacyclics, TG Therapeutics, Pfizer, Sunesis: Research Funding. Coutre: Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; AstraZeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees; Beigene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Data Safety Monitoring Committee, Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; Acerta: Other: Data Safety Monitoring Committee, Research Funding. Barr: AstraZeneca: Consultancy; Gilead: Consultancy; Janssen: Consultancy; Genentech: Consultancy; Beigene: Consultancy; Bristol Meyers Squibb: Consultancy; Seattle Genetics: Consultancy; TG Therapeutics: Consultancy; Morphosys: Consultancy; Abbvie/Pharmacyclics: Consultancy. Cashen: Secura Bio, ADC Therapeutics: Consultancy. Mato: Adaptive Biotechnologies: Consultancy, Research Funding; DTRM BioPharma: Consultancy, Research Funding; Johnson and Johnson: Consultancy, Research Funding; Acerta/AstraZeneca: Consultancy, Research Funding; Sunesis: Consultancy, Research Funding; AstraZeneca: Consultancy; BeiGene: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; AbbVie: Consultancy, Research Funding; LOXO: Consultancy, Research Funding; MSKCC: Current Employment; Pharmacyclics LLC, an AbbVie Company: Consultancy, Research Funding; TG Therapeutics: Consultancy, Other: DSMB, Research Funding; Nurix: Research Funding; Janssen: Consultancy, Research Funding; Genmab: Research Funding. Erba: AbbVie Inc; Agios Pharmaceuticals Inc; Astellas; Bristol Myers Squibb; Celgene, a Bristol Myers Squibb company; Daiichi Sankyo Inc; Genentech, a member of the Roche Group; GlycoMimetics Inc; Incyte Corporation; Jazz Pharmaceuticals Inc; Kura Oncology; Nov: Other: Advisory Committee; AbbVie Inc: Other: Independent review committee; AbbVie Inc; Agios Pharmaceuticals Inc; ALX Oncology; Amgen Inc; Daiichi Sankyo Inc; FORMA Therapeutics; Forty Seven Inc; Gilead Sciences Inc; GlycoMimetics Inc; ImmunoGen Inc; Jazz Pharmaceuticals Inc; MacroGenics Inc; Novartis; PTC Therapeutics: Research Funding; AbbVie Inc; Agios Pharmaceuticals Inc; Bristol Myers Squibb; Celgene, a Bristol Myers Squibb company; Incyte Corporation; Jazz Pharmaceuticals Inc; Novartis: Speakers Bureau. Stone: Onconova: Consultancy; AbbVie: Consultancy; Boston Pharmaceuticals: Consultancy; Bristol Myers Squibb: Consultancy; Gemoab: Membership on an entity's Board of Directors or advisory committees; Elevate Bio: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Research Funding; Jazz: Consultancy; Syros: Membership on an entity's Board of Directors or advisory committees; Syndax: Membership on an entity's Board of Directors or advisory committees; Agios: Consultancy, Research Funding; Takeda: Consultancy; BerGen Bio: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Foghorn Therapeutics: Consultancy; Innate: Consultancy; Janssen: Consultancy; GlaxoSmithKline: Consultancy; Amgen: Membership on an entity's Board of Directors or advisory committees; Arog: Consultancy, Research Funding; Actinium: Membership on an entity's Board of Directors or advisory committees; Syntrix/ACI: Membership on an entity's Board of Directors or advisory committees; Aprea: Consultancy; Celgene: Consultancy; Macrogenics: Consultancy. Litzow: Actinium: Research Funding; Amgen: Research Funding; AbbVie: Research Funding; Pluristem: Research Funding; Jazz: Other: Advisory Board; Omeros: Other: Advisory Board; Astellas: Research Funding; Biosight: Other: Data monitoring committee. Tallman: NYU Grand Rounds: Honoraria; Kura: Membership on an entity's Board of Directors or advisory committees; Syros: Membership on an entity's Board of Directors or advisory committees; Innate Pharma: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Biosight: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Oncolyze: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; Orsenix: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Rafael Pharmaceuticals: Research Funding; Glycomimetics: Research Funding; Biosight: Research Funding; Orsenix: Research Funding; Abbvie: Research Funding; Mayo Clinic: Honoraria; UC DAVIS: Honoraria; Northwell Grand Rounds: Honoraria; NYU Grand Rounds: Honoraria; Danbury Hospital Tumor Board: Honoraria; Acute Leukemia Forum: Honoraria; Miami Leukemia Symposium: Honoraria; New Orleans Cancer Symposium: Honoraria; ASH: Honoraria; NCCN: Honoraria. Shanafelt: Genentech, Pharmacyclics: Research Funding. Wagner: Eli Lilly, Johnson & Johnson: Other: Spouse, previously held individual stocks;; Celgene-BMS: Consultancy, Membership on an entity's Board of Directors or advisory committees; Athenex Inc: Consultancy.

Background MBL is a precursor to chronic lymphocytic leukemia (CLL) and is subclassified into low-count (LC) MBL (absolute B-cell count<0.5x109/L) and high-count (HC) MBL (absolute B-cell count between 0.5 and 5x109/L). We previously reported that a polygenic risk score (PRS) based on a weighted average of 41 CLL-susceptibility variants was associated with risk of both MBL and CLL among a cohort of individuals from CLL families. Here we evaluate this PRS in an independent cohort of MBL and CLL individuals of European ancestry (EA), all of whom were ascertained agnostic to CLL family-history status. We also evaluate the PRS by MBL subtype (LC/HC), and in African American (AA) CLL cases and controls. Methods We genotyped 535 EA MBLs (139 HC-MBL, 396 LC-MBLs), 735 CLLs (640 EA, 95 AA), and 2,866 controls (2,631 EA, 235 AA) from the Mayo Clinic CLL Resource, Duke University, and Weill Cornell Medical College. We computed the CLL-PRS for each individual and used logistic regression to estimate odds ratios (OR) and 95% confidence intervals, adjusting for age and sex. To assess discriminatory accuracy, we computed the c-statistic. Among EA individuals, we calculated a trend test among LC-MBL, HC-MBL, and CLL risk using the P-value for heterogeneity from a polytomous logistic regression analysis. Moreover, we plotted a boxplot for the PRS among controls, LC-MBL, HC-MBL, and EA CLL, as well as for AA CLL cases and controls, and tested the statistical difference using the Kruskal Wallis test and Mann-Whitney test, respectively. Results We found a significant association of PRS with overall MBL risk (OR=1.87, P=1.1x10-28) with good discrimination (c-statistic=0.72). Significant associations were also found for LC-MBL (OR=1.75, P=7.5x10-19, c-statistic=0.72), HC-MBL (OR=2.22, P=1.4x10-17, c-statistic=0.74), and CLL of EA (OR=2.60, P=1.2x10-62, c-statistic=0.78), with a significant difference among these cohorts (Figure 1.A) and a significant positive trend across these cohorts (Pheterogeneity=8.4x10-6). Although we observed a 33% increased risk of CLL in AA (c-statistic=0.57), the PRS was borderline significant (P=0.07, Figure 1.B). Conclusion The CLL-PRS is a strong prediction-tool for risk of CLL and MBL among individuals of EA. Future studies are needed to improve the PRS for AAs including performing GWAS of AA in order to identify CLL-susceptibility SNPs that are more representative within known CLL loci and to discover novel CLL loci that are unique for AAs. Disclosures Parikh: GlaxoSmithKline: Honoraria; Janssen: Honoraria, Research Funding; Ascentage Pharma: Research Funding; AbbVie: Honoraria, Research Funding; Merck: Research Funding; TG Therapeutics: Research Funding; Genentech: Honoraria; Pharmacyclics: Honoraria, Research Funding; MorphoSys: Research Funding; AstraZeneca: Honoraria, Research Funding; Verastem Oncology: Honoraria. Braggio:DASA: Consultancy; Bayer: Other: Stock Owner; Acerta Pharma: Research Funding. Brander:Genentech: Consultancy, Honoraria, Other, Research Funding; Juno/Celgene/BMS: Other, Research Funding; MEI Pharma: Other, Research Funding; Ascentage: Other, Research Funding; ArQule: Consultancy, Other, Research Funding; NCCN: Other; Teva: Consultancy, Honoraria; Tolero: Research Funding; AstraZeneca: Consultancy, Honoraria, Other, Research Funding; Pharmacyclics LLC, an AbbVie Company: Consultancy, Honoraria, Other, Research Funding; Pfizer: Consultancy, Other; TG Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding; Novartis: Consultancy, Other; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding; Tolero: Research Funding; Teva: Consultancy, Honoraria; DTRM: Other, Research Funding; BeiGene: Other, Research Funding; Novartis: Consultancy, Other; NCCN: Other; Verastem: Consultancy, Honoraria, Other, Research Funding. Cerhan:NanoString: Research Funding; BMS/Celgene: Research Funding. Kay:Astra Zeneca: Membership on an entity's Board of Directors or advisory committees; Agios Pharma: Membership on an entity's Board of Directors or advisory committees; Sunesis: Research Funding; Pharmacyclics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Acerta Pharma: Research Funding; Juno Theraputics: Membership on an entity's Board of Directors or advisory committees; Morpho-sys: Membership on an entity's Board of Directors or advisory committees; Rigel: Membership on an entity's Board of Directors or advisory committees; Cytomx: Membership on an entity's Board of Directors or advisory committees; Tolero Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol Meyer Squib: Membership on an entity's Board of Directors or advisory committees, Research Funding; MEI Pharma: Research Funding; Abbvie: Research Funding; Oncotracker: Membership on an entity's Board of Directors or advisory committees; Dava Oncology: Membership on an entity's Board of Directors or advisory committees. Furman:Acerta: Consultancy; AstraZeneca: Consultancy, Research Funding; Beigene: Consultancy; Abbvie: Consultancy; Pharmacyclics: Consultancy; Sunesis: Consultancy; TG Therapeutics: Consultancy, Research Funding; Verastem: Consultancy; Incyte: Consultancy; Genentech: Consultancy; Janssen: Consultancy, Speakers Bureau; Loxo Oncology: Consultancy; Oncotarget: Consultancy. Shanafelt:Mayo Clinic: Patents & Royalties: and other intellectual property; Genentech, Pharmacyclics LLC, an AbbVie Company, AbbVie, GlaxoSmithKline, and Merck: Research Funding.

Background : The presence of TP53 aberrations (defined as del(17p) or TP53 gene mutation) is a strong negative predictor of survival in patients (pts) with CLL. First-line chemoimmunotherapy is suboptimal in pts with CLL bearing TP53 aberrations, with 3-year progression-free survival (PFS) and overall survival (OS) rates of only 18% and 38%, respectively, with fludarabine, cyclophosphamide, and rituximab (Hallek, Lancet 2010). Ibrutinib is the only once-daily Bruton's tyrosine kinase (BTK) inhibitor with significant PFS and OS benefit demonstrated in multiple randomized phase 3 studies (eg, RESONATE-2, ECOG1912) in the first-line treatment of CLL/small lymphocytic lymphoma (SLL). Previous reports of single-agent ibrutinib or ibrutinib-based combination treatments have further demonstrated favorable PFS benefit in pts with TP53 aberrations in both the first-line and relapsed/refractory settings. Despite these study-specific subgroup analyses, there are limited data on long-term outcomes in pts with TP53 aberrations treated with first-line BTK inhibitors. We performed a pooled analysis of data across 4 studies to evaluate the long-term efficacy and safety of first-line ibrutinib-based therapy in pts with CLL bearing TP53 aberrations. Methods : Data for first-line ibrutinib treatment in pts with TP53 aberrations were pooled across 4 clinical trials in CLL/SLL: PCYC-1122e (NCT01500733; single-agent ibrutinib; n=34), PCYC-1130 (NCT02264574; ibrutinib + obinutuzumab; n=18), ECOG1912 (NCT02048813; ibrutinib + rituximab; n=26), and RESONATE-2 (NCT01722487; single-agent ibrutinib; n=11). ECOG1912 and RESONATE-2 excluded pts with del(17p) but did not exclude pts with TP53 mutations. Long-term PFS (assessed by investigator), OS, and safety are reported. Results : Eighty-nine pts with TP53 aberrations receiving first-line ibrutinib treatment were included in this pooled analysis. Median age was 65 years (range 33-87), and 69% of pts were male. At baseline, 53% had Rai stage III/IV, 38% had bulky disease (lymph nodes ≥5 cm), and 69% (of 87 evaluable) had unmutated IGHV. All patients had either del(17p) or TP53 mutation; 53% (of 89 evaluable) had del(17p) and 91% (of 58 evaluable) had TP53 mutation. Among 16 pts with del(17p) who had TP53 sequencing results available, 11 (69%) had both del(17p) and TP53 mutation. Forty-five pts received ibrutinib as a single agent and 44 received ibrutinib in combination with an anti-CD20 agent. With a median follow-up of 50 months (range 0.1 to 95.9 months), median PFS was not reached (95% CI: 67 months to not estimable; Figure 1A). At 48 months, the PFS rate was 79% and the OS rate was 88% (Figure 1B). Median duration of ibrutinib treatment was 46 months (range 0.1 to 95.5 months). Reasons for treatment discontinuation were progressive disease (20%), study closure (12%), adverse event (10%), withdrawal by pt (7%), death (3%), and other (physician decision due to scheduled pt surgery; 1%). Grade ≥3 adverse events of clinical interest with up to 8 years of treatment with ibrutinib were infection (22%; most commonly pneumonia in 7%), hypertension (13%), atrial fibrillation (12%), and major hemorrhage (7%). With the current follow-up, 46% of pts with TP53 aberrations remained on ibrutinib treatment. Conclusions : With a median follow-up of 4 years, first-line ibrutinib-based treatment resulted in sustained efficacy with high PFS and OS rates in CLL pts with TP53 aberrations, a population with historically poor outcomes. Although pts with TP53 aberrations remain at risk for progression, first-line treatment with ibrutinib has partially overcome the poor prognosis in this high-risk population with 4-year PFS and OS rates of 79% and 88%, respectively. No new safety signals were identified in this analysis. These results from a large, pooled, multi-study dataset demonstrated the long-term benefit of first-line ibrutinib-based treatment in pts with TP53 aberrations. Disclosures Allan: Janssen, AbbVie, and AstraZeneca: Other: Travel/accommodations/expenses; Celgene, Genentech, AstraZeneca, TG Therapeutics, and Janssen: Research Funding; AstraZeneca, Pharmacyclics LLC, an AbbVie Company, Genentech, AbbVie, Ascentage, and Cellectar: Consultancy; Janssen, AstraZeneca, and AbbVie: Honoraria. Shanafelt:Genentech, Pharmacyclics LLC, an AbbVie Company, AbbVie, GlaxoSmithKline, and Merck: Research Funding; Mayo Clinic: Patents & Royalties: and other intellectual property. Wiestner:Pharmacyclics LLC, an AbbVie Company; Acerta, Merck, Nurix, Verastem, and Genmab: Research Funding; National Institutes of Health: Patents & Royalties: and other intellectual property. Moreno:Janssen, AbbVie, Sunesis, and AstraZeneca: Consultancy; AbbVie and Janssen: Research Funding; Janssen: Speakers Bureau. O'Brien:Eisai: Consultancy; Juno Therapeutics: Consultancy; Aptose Biosciences: Consultancy; GlaxoSmithKline: Consultancy; Sunesis: Research Funding; Acerta: Research Funding; Amgen: Consultancy; Celgene: Consultancy; Pfizer: Research Funding; Pharmacyclics: Research Funding; Astellas: Consultancy; Gilead: Consultancy; TG Therapeutics: Research Funding; Vida Ventures: Consultancy; KITE: Research Funding; Vaniam Group LL: Consultancy; Alexion: Consultancy; Regeneron: Research Funding; AbbVie: Consultancy; Verastem: Consultancy; Janssen Oncology: Consultancy. Braggio:DASA: Consultancy; Bayer: Other: Stock Owner; Acerta Pharma: Research Funding. Liu:AbbVie: Current equity holder in publicly-traded company; Pharmacyclics LLC, an AbbVie Company: Current Employment. Dean:AbbVie: Current equity holder in publicly-traded company; Pharmacyclics LLC, an AbbVie Company: Current Employment. Lai:Pharmacyclics LLC, an AbbVie Company: Current Employment; AbbVie and Bristol-Myers Squibb: Current equity holder in publicly-traded company; AbbVie: Other: Travel/accommodations/expenses.

INTRODUCTION Nearly 20% of patients (pts) are ≥ 80 years (yrs) of age at the time of CLL diagnosis. Increased medical comorbidities, as well as declining functional status and organ function, often accompany advanced age. However, clinical trials often enroll a disproportionally low number of people from this age demographic due to stringent eligibility parameters, which provides limited evidence to guide decisions. Here, we report clinical characteristics and outcomes of pts ≥ 80 yrs of age at the time of CLL diagnosis. METHODS Between 1/1995 and 3/2020, we identified previously untreated CLL pts from the Mayo Clinic CLL Database who were ≥ 80 yrs of age at the time of CLL diagnosis, and seen within 3 yrs of diagnosis. Baseline characteristics, treatments administered, and time to first treatment (TFT) were analyzed for all pts. Overall survival (OS) was measured from diagnosis date in all pts and separately from treatment date for treated pts. Time to next treatment (TTNT) was measured in treated pts from first treatment date to date of second treatment or last known treatment. TFT and TTNT were analyzed accounting for competing risk of death. Standardized mortality ratio (SMR) compared observed survival to survival in a Minnesota population. Cox multivariable regression models were used to determine which factors were associated with OS (treatment as a time-dependent covariate) and TTNT (accounting for competing risk of death). RESULTS Among the 213 pts identified, the median age was 83 yrs (range 80-97); baseline characteristics are summarized in Table 1. The median number of medical comorbidities among all pts was 4 (range 0-8). At least 1 comorbidity was present in 97% of pts, most commonly hypertension (57%), genitourinary (n=48%), rheumatological (46%), and dyslipidemia (n=43%). Median follow-up was 3.8 yrs. A total of 56 pts received treatment and the median TFT was 6.7 yrs. Over this 25 year interval, the treatment approaches included monoclonal antibody alone (n=17), chemotherapy alone (n=17), or chemoimmunotherapy (n=14), novel agents (n=3), and multi-agent Richter's transformation regimens (n=5). Chlorambucil-based treatments (chlorambucil n=13, chlorambucil, prednisone n=3) were the most frequently used chemotherapy regimens. The most common chemoimmunotherapy regimens included rituximab, cyclophosphamide, prednisone (RCP) with (n=3) or without vincristine (n=6), and chlorambucil with an anti-CD20 monoclonal antibody (n=4; obinutuzumab n=2, rituximab n=2). Among 17 pts who received monoclonal antibody alone, treatments included rituximab alone (n=11), rituximab plus methylprednisolone (n=2), obinutuzumab (n=2), and rituximab with alemtuzumab (n=2). Frontline novel agent treatments included ibrutinib alone (n=1), acalabrutinib alone (n=1), and ibrutinib, obinutuzumab, venetoclax (n=1). Five pts with Richter's transformation (diagnosed concurrent with CLL n=4) received multi-agent chemotherapy. Among the 56 treated pts, 22 pts received second line therapy after a median TTNT of 2.8 yrs from start of first therapy. The most common second line treatments were chlorambucil-based (n=6) and rituximab-based (n=6) regimens. Age, sex, absolute lymphocyte count, del17p, and unmutated IGHV were not predictive of TTNT. The median OS among all 213 pts was 4.6 yrs. SMR was 1.2 (p=0.008) when excluding pts with Richter's transformation. In univariable analyses, receipt of therapy (HR 3.92, 95%CI 2.11-7.28; p<0.001), age (per 5 year increase; HR 1.84, 95%CI 1.43-2.38; p<0.001) and unmutated IGHV (HR 2.03, 95%CI 1.12-3.67; p=0.02) were predictors of shorter OS. The OS among pts who received treatment (Figure 1) was similar when compared across time periods of treatment start year grouped as 1995-2001 (n=10), 2002-2013 (n=32), and 2014-2020 (n=14). Treatment categories by time period are shown in Table 2. CONCLUSIONS In this study of 213 pts ≥ 80 yrs of age at time of CLL diagnosis, the median time to first therapy was ~7 yrs. Although survival outcomes did not differ by era across the 25-year span of this study, pts treated with novel agents were underrepresented and are a patient population which warrants additional evaluation. The finding that pts with CLL in this cohort had a 20% higher risk of death compared to an age- and sex-matched population emphasize the need for ongoing efforts to improve clinical outcomes for CLL pts in this demographic category. Disclosures Kenderian: Torque: Consultancy; Mettaforge: Patents & Royalties; Humanigen: Consultancy, Patents & Royalties, Research Funding; Novartis: Patents & Royalties, Research Funding; Kite: Research Funding; Gilead: Research Funding; Juno: Research Funding; BMS: Research Funding; Tolero: Research Funding; Sunesis: Research Funding; MorphoSys: Research Funding; Lentigen: Research Funding. Wang:Incyte: Research Funding; Innocare: Research Funding; Novartis: Research Funding. Shanafelt:Genentech, Pharmacyclics LLC, an AbbVie Company, AbbVie, GlaxoSmithKline, and Merck: Research Funding; Mayo Clinic: Patents & Royalties: and other intellectual property. Braggio:DASA: Consultancy; Bayer: Other: Stock Owner; Acerta Pharma: Research Funding. Kay:Morpho-sys: Membership on an entity's Board of Directors or advisory committees; Bristol Meyer Squib: Membership on an entity's Board of Directors or advisory committees, Research Funding; Tolero Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Research Funding; Oncotracker: Membership on an entity's Board of Directors or advisory committees; Juno Theraputics: Membership on an entity's Board of Directors or advisory committees; Rigel: Membership on an entity's Board of Directors or advisory committees; Cytomx: Membership on an entity's Board of Directors or advisory committees; Sunesis: Research Funding; MEI Pharma: Research Funding; Astra Zeneca: Membership on an entity's Board of Directors or advisory committees; Agios Pharma: Membership on an entity's Board of Directors or advisory committees; Abbvie: Research Funding; Pharmacyclics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Acerta Pharma: Research Funding; Dava Oncology: Membership on an entity's Board of Directors or advisory committees. Ding:alexion: Membership on an entity's Board of Directors or advisory committees; MEI Pharma: Membership on an entity's Board of Directors or advisory committees; Octapharma: Membership on an entity's Board of Directors or advisory committees; Abbvie: Research Funding; Astra Zeneca: Research Funding; Beigene: Membership on an entity's Board of Directors or advisory committees; DTRM: Research Funding; Merck: Membership on an entity's Board of Directors or advisory committees, Research Funding. Parikh:Merck: Research Funding; AstraZeneca: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; MorphoSys: Research Funding; Pharmacyclics: Honoraria, Research Funding; AbbVie: Honoraria, Research Funding; Ascentage Pharma: Research Funding; Genentech: Honoraria; Verastem Oncology: Honoraria; GlaxoSmithKline: Honoraria; TG Therapeutics: Research Funding.

Abstract CLL/SLL and other CLPD are associated with an increased risk of autoimmune hemolytic anemia (AIHA), immune thrombocytopenic purpura (ITP), pure red blood cell aplasia (PRBCA), and autoimmune agranulocytosis (AIAG). CLL patients with AIHA and ITP have advanced stage disease according to the Rai and Binet clinical classifications, implying a poor prognosis. However, previous community based studies suggest that patients with CLL and immune cytopenias could have a similar prognosis to other patients with CLL. To further define the prognostic significance of autoimmune cytopenia, we studied a cohort of patients seen at the Mayo Clinic Rochester (MCR). Methods The MCR Division of Hematology Chronic Lymphoproliferative Disease Database was used to identify patients with CLPD and autoimmune complications (AID) over a 10 year period (1 January 1995 to 30 December 2004). The clinical records of all patients with AID were thoroughly reviewed and collated with additional biological data. Results Forty-one (6%) of 665 patients with CLPD had AID. Of the patients with CLPD and AID, 21 (51%) had AIHA, 19 (46%) had ITP, 8 (20%) had PRBCA and 2 (5%) had AIAG. 17% of patients had 2 or more AID diagnoses. In 4 patients (10%) the diagnosis of AID preceded the diagnosis of CLPD, and in 7 patients (17%) the diagnoses were made concurrently. In the 32 patients with CLL, the median lymphocyte count at diagnosis was 24.8 x 109/L (5.18 – 121.8). Of 27 CLL patients who had interphase FISH analysis performed, 12 (44%) had abnormalities associated with poor prognosis CLL (17p13–11% 11q22–22%, 12+ 11%), and 15 (56%) had results associated with a more favorable prognosis (no detectable abnormalities or 13q14 deletion as the sole abnormality). CD38 was positive (> 30% of cells) in 38% of 21 tested patients. 39 patients required treatment for their AID. Treatment included corticosteroids (90%), splenectomy (31%), chemotherapy (31%), and rituximab (46%). Response rates following the first treatment for AID were: AIHA 53% complete response (CR) and 26% partial response (PR); ITP 56% CR and 22% PR; PRBCA 38% CR and 38% PR; AIAG no responses. Of the 41 patients seen over the 10 year period, there were 4 deaths (2 from CLPD). Conclusions This study of a large cohort of community and referred patients showed a prevalence of AID similar to that reported in community based studies. We found an equivalent prevalence of AIHA and ITP as well as a substantial number of patients with 2 or more AID complications. This could be due in part to detailed workup of all cytopenias. In addition, 27% of patients did not have a prior diagnosis of a B cell malignancy when presenting with AID. The measured biological risk factors (FISH anomalies and CD38) suggest that CLL patients with AID do not have a higher percentage of high risk CLL. The majority of patients responded well to AID treatment. Mortality appears to be lower than CLL patients with non-autoimmune cytopenia. These preliminary data suggest that AID is a common and often presenting complication of CLL/SLL and other CLPD. Early diagnosis and management of AID could lead to improved outcomes.

Abstract BACKGROUND: We have previously studied and reported that the combination of pentostatin (P, 2 mg/m2), cyclophosphamide (C 600 mg/m2) and rituximab (R 375 mg/m2) in previously untreated CLL is highly effective with an overall response (OR) rate of over 90% and a complete response rate (CR) of 41% (Blood109:405–411, 2007). We also found that this regimen can be effective even in older patients (>70 y), those with elevated beta-2 microglobulin levels, and patients with mildly reduced creatinine clearances (Cancer. 109:2291–2298, 2007). To determine whether similar benefit could be achieved without inclusion of an alkylating agent, we conducted a follow-up trial testing pentostatin and rituximab without cyclophosphamide and employing a higher pentostatin dose (4 mg/m2). METHODS: Eligible pts had documentation of active CLL by standard NCI-WG criteria, and were previously untreated. Treatment schema consisted of 6 cycles of pentostatin (4 mg/m2) and rituximab given every 21 days. Pentostatin was given on the first day of each cycle following infusion of rituximab. Rituximab was given at 100mg/m2 IV at day 1, then 375 mg/m2 IV on days 3 and 5 of the first treatment cycle. During cycles 2 to 6, Rituximab was given at 375 mg/m2 as a single IV infusion day 1 of week 4, 7, 10, 13 and 16. All patients were staged two months after completion of the 6 cycles of PR using the NCI-WG criteria. PATIENT CHARACTERISTICS: Overall, 33 patients were enrolled at Mayo Clinic and Ohio State University between July 2005 and February 2008. All 33 were eligible: 82% male, median age 65 (range: 45–81), with 9 (27%) being 70 years or older. 76% had a baseline ECOG PS of 0, and the rest were ECOG PS 1. Overall, 36% of patients had intermediate Rai risk (stage I–II) and 63% high Rai risk (stage 3–4) disease. Prognostic testing revealed that 36% were CD38+, 50% were Zap-70 +, and 39% had an unmutated IgVh status. Chromosome analysis by FISH found that 99% had detectable FISH panel defect, including 61%, 27% and 3% of patients with 1, 2, or 3 FISH detects, respectively. RESULTS: 28 of 33 patients (85%) completed therapy. While on treatment, 6 pts (18%) had a dose held or modified with 4 of these delays due to hematologic AE. For adverse events deemed at least possibly related to treatment, 4 (12%) pts experienced grade 3+ hematologic toxicity and 5 (15%) experienced grade 3+ non-hematologic toxicity. Out of all 33 enrolled patients, the overall response rate was 79% with 10 CR, 6 nPR, and 10 PR. At the time of this analysis, 29/33 patients are still alive with a median follow-up time of 14 months on surviving patients. To date 17/33 (52%) of patients have progressed with an estimated median time to progression of 12 months (95% CI: 8.5–21 months). 13/26 responders have progressed. Median duration of response is 12.5 months ((95% CI: 11–21 months). Finally, since eligibility were nearly identical and enrollment accrued at the same two academic centers, we compared the patient characteristics, response rates, and PFS of the 33 patients treated with PR to the 64 patients previously treated on our PCR trial. Patients in the two studies were generally similar with respect to demographic and prognostic characteristics, although patients in the PR trial had higher WBC and were less likely to be IgVH unmutated (Table). Although the differences in ORR and CR rate were not significantly different, the PFS appeared to be inferior in patients treated with PR as compared to PCR (12 months vs. 31 months; p=0.003). CONCLUSION: Although the PR regimen achieves a high OR response rate, the PFS appears inferior to PCR therapy. These findings suggest that increasing the purine nucleoside analogue dose does not eliminate the need to include cyclophosphamide in chemoimmunotherapy for patients with CLL. PCR Trial N=64 PR Trial N=33 P value Age, median(range) 63 years (38–80) 65 years (45–81) 0.34 ≥70 years(%) 28% 27% Male 77% 82% 0.61 Rai stage 0 5% 0 0.46 Rai stage I–II 42% 36% Rai stage III–IV 53% 64% White Cell Count, median(range) 79 × 109/L (11–519) 127 × 109/L (8–430) 0.04 <50 × 109/L 36% 28% 50–149 × 109/L 44% 25% >150 × 109/L 20% 47% Serum B2-microglobulin, median(range) 3.97 (1.8–13.5) 3.80 (2.0–8.2) 0.81 >2 × Upper Limit Normal(%) 57% 58% CD38 Positive 34% 36% 1.00 ZAP-70 Positive 36% 50% 0.26 IgVH Unmutated 71% 39% 0.004 FISH Normal, 11% 9% 13q- 35% 42% +12 21% 24% 6q- 2% 0 11q- 22% 18% 17p- 6% 3% other 3% 3% Overall Response Rate 91% 79% 0.12 Complete Response Rate 41% 30% 0.38 Median PFS 31 months 12 months 0.003

Abstract Myeloid cell factor-1 (Mcl-1) is an anti-apoptotic member of the Bcl-2 protein family that associates with tumor cell survival and drug resistance in chronic lymphocytic leukemia (CLL). Increased Mcl-1 expression is associated with failure to achieve remission after chemotherapy with fludarabine and chlorambucil in CLL patients. However, the influence of Mcl-1 expression has not been examined in CLL trials using treatment regimens that contain rituximab, which has shown substantially improved outcomes for a large subset of patients and have become a mainstay in the therapy of newly diagnosed CLL. We previously reported that combination chemoimmunotherapy with pentostatin, cyclophosphamide, and rituximab (PCR) has significant clinical activity with low accompanying toxicity in previously untreated CLL patients, and is especially well tolerated in older patients in whom the use of fludarabine may be associated with prohibitive toxicities. As part of this study, we prospectively investigated association of Mcl-1 protein expression with response and progression-free survival. Eligible patients received a regimen consisting of pentostatin (2 mg/m2), cyclophosphamide (600 mg/ m2), and rituximab (375 mg/m2) given intravenously on day 1 of a 21-day cycle for a maximum of 6 cycles. Responses were assessed according to NCI 1996 criteria and included bone marrow evaluation and two-color flow cytometry two months after completion of therapy. Flow cytometry-negative status was defined as less than or equal to 1% CD5+/CD19+ cells. Of the 64 patients evaluated in this trial, clinical responses were seen in 58 (91%), with 26 (41%) complete responses (CR), 14 (22%) nodular partial responses (nPR), and 18 (28%) partial responses (PR). Lysates from peripheral blood cells obtained pre-treatment were analyzed for Mcl-1 expression by immunoblot. Mcl-1 as a continuous variable was not found to be associated with other commonly described risk factors. However, Recursive Partitioning Analysis (RPA) was devised employing Mcl-1 expression as a continuous variable, which revealed an optimal cut point of 0.85. At this cutoff, flow cytometry-negative status was significantly different between the two groups (p=0.01) and median PFS was significantly higher (p=0.02) in patients with Mcl-1 levels of <0.85 (50.8 vs. 18.7 months). No difference by Mcl-1 expression was noted in other pre-treatment or response parameters. Our data describes the first prospective validation of Mcl-1 over-expression adversely influencing both the likelihood of attaining a flow cytometry-negative complete remission and extended progression free survival. Mcl-1 expression was also independent of other risk factors, indicating additional benefit for this measure in assessing patients. Mcl-1 expression may therefore predict poor response to chemoimmunotherapy, and these findings advocate pursuing therapeutic agents targeting this important anti-apoptotic protein.

Abstract Serum BLyS levels have been found to be elevated in a number of immune disease models and there is increasing evidence that this may correlate with pathogenesis of various B cell related disorders including B cell malignancies. While it is clear that BLyS expression is required for normal B cell development and homeostasis, the exact source of BLyS in these scenarios and definition of the mechanisms that control BLyS expression remain to be fully elucidated. Serum BLyS levels are elevated in a number of B cell malignancies known to have a familial incidence. Therefore, we sought to determine if there was any correlation between serum BLyS levels and family history of B cell cancers. In our initial studies we examined the serum BLyS levels in patients with sporadic vs. familial B-cell chronic lymphocytic leukemia (B-CLL; near relative with B-CLL, multiple myeloma, or non-Hodgkin lymphoma), and normal age-matched controls. In the normal controls (n=50), the mean serum BLyS level was 12.7 ng/ml, and we found that 4/50 (8%) had BLyS levels exceeding 20 ng/ml. In the sporadic CLL cohort (n=52), the mean serum BLyS level was 18.3 ng/ml, and we found that 5/52 (10%) had BLyS levels exceeding 20 ng/ml. In striking contrast, in the familial CLL cohort (n=24), the mean serum BLyS levels was 33.4 ng/ml, and we found that 11/24 (46%) had BLyS levels exceeding 20 ng/ml. The percentage of patients with elevated BLyS levels, as well as the mean BLyS levels in the familial CLL cohort compared to the normal controls, was significantly higher (mean, p=0.002) and suggests that elevated BLyS levels may correlate with familial CLL. Because of the correlation between BLyS levels and family history B cell malignancies, we next wanted to determine if there was a common underlying genetic event influencing BLyS expression in this group of patients. We began by sequencing the BLyS promoter in 19 patients with B-CLL and 11 normal controls. We identified 2 sites that were polymorphic, −661 A/G and −871 C/T. The −871 T/T genotype has been previously reported to be expressed at increased frequency in SLE patients and is associated with elevated BLyS mRNA levels. Using RFLP analysis we examined the presence of the two polymorphisms in our control, sporadic, and familial CLL cohorts. In the control group (n=50) we found that at −871, C/C (wildtype) was expressed in 24%, C/T in 54%, and T/T in 22% of patients. Similarily, in the sporadic CLL group (n=35) we found that at −871, C/C was expressed in 29%, C/T in 57%, and T/T in 14%. In the familial group (n=24) we found that at −871, C/C was expressed in only 8%, C/T in 67%, and T/T in 25%. These data suggest a decreased representation of −871 C/C in the familial group. No significant changes were observed between the three groups at the −661 A/G locus. Additionally, we found that none of the patients with BLyS levels exceeding 20 ng/ml expresses C/C at −871. In summary, our data suggest that BLyS levels are elevated in patients with familial CLL relative to both normal controls and individuals with sporadic CLL and that elevated BLyS levels may correlate with the presence of a T at −871 in the BLyS promoter.