Lymphoid Malignancies

Past research indicated a possible role for antigen in the pathogenesis of CLL and identified numerous interactions between tumor cells and the tissue microenvironment. I lead work in my group that identified the lymph node microenvironment as a key site of CLL cell activation through B-cell receptor and NF-ĸB signaling. These studies provide a framework for targeted therapy in CLL. Extending these observations we have employed single cell sequencing to identify a distinct minor subpopulation of activated and proliferating CLL cells in the lymph node microenvironment that interface with the immune milieu, and promote disease progression. Using exome sequencing of matched samples from blood and lymph nodes, we found that clonal evolution primarily occurred in the lymph node and associated with a suppressed T-cell inflammatory response. In turn, CLL cells actively modulate their surroundings giving rise to a complex, bidirectional crosstalk between tumor and host cells.

Beyond these microenvironmental dependencies, a diverse set of putative pathogenic mutations have been identified through large-scale sequencing efforts. Heterogeneity in the genetic architecture of CLL has been closely linked to disease aggressiveness and its capacity to evolve under the selective pressures of treatment. Over the last decade an increasing number of driver mutations have been identified revealing marked heterogeneity in the genetic composition of CLL. Together with many colleagues we contributed to an aggregate “CLL map” build on genomic, transcriptomic, and epigenomic data from over 1,000 patients.

Among the first forays into targeted therapy of CLL, we conducted a single center study with ibrutinib in CLL patients. We focused the study on patients with deletion of chromosomal arm 17p (del17p) and/or mutations in TP53. These patients generally had poor outcomes with chemotherapy. Virtually all patients responded to ibrutinib and achieved long-lasting benefit.

Figure 1. Durable benefit of treatment with ibrutinib for patients with CLL and TP53 aberration (del17p and/or TP53 mutation). From Ahn, Tian, Wiestner. NEJM 2020.

We also reported on safety, especially regarding cardiac events and demonstrated on-target effects of ibrutinib on BCR and NF-ĸB signaling in blood and lymph node, and identified improvements in both humoral and cellular immunity on treatment. Investigating mechanisms of resistance to ibrutinib, we identified mutations in BTK and PLCG2 in most patients with CLL progressing on ibrutinib, showed that these mutations are often detectable long before the diagnosis of clinical relapse, and identified emergence of multiple subclones carrying different mutations. Based on our study results, we developed and validated a 4-factor prognostic model for patients being treated with BTK inhibitors. We also identified expression of CD49d on CLL cells as a predictor of outcome with BTKi therapy, and showed that CD49d could independently add to the prognostic information of the 4-factor model.

In our phase 2 study of acalabrutinib, a more BTK selective inhibitor than ibrutinib, we found that 
twice daily dosing maintained near complete occupancy of BTK in blood and tissues and more profoundly inhibited oncogenic signaling than once daily dosing.

Figure 2. Visual abstract summarizing key aspects of our phase 2 investigator-initiated study of acalabrutinib in patients with relapsed/refractory or high-risk treatment-naïve CLL. From Sun et al, Blood 2020

Novel therapies are needed for high-risk patients previously treated with BTKi and venetoclax. In the laboratory, NRX-0492, a targeted protein degrader, induced rapid and sustained degradation of wild type and C481 mutant BTK at sub-nanomolar concentrations in primary CLL samples and demonstrated in vivo anti-CLL activity in patient-derived xenografts and the CLL-PDX model. This led us to join a multi-center first in human study further investigating BTK degradation with the clinical compound, NX-2127, with the same pharmacological mechanism as NRX-0492.

Figure 3. NRX-0492 effectively degrades wildtype and mutant BTK in vitro and in vivo. (A) NRX-0492 consists of noncovalent BTK-binding “hook” linked to a “harness” that recruits cereblon to target BTK for ubiquitination and proteasomal degradation. (B) NRX-492 degrades BTK in U-CLL and M-CLL samples at low nanomolar concentrations. (C) CLL cell transcriptome assessed by RNA sequencing of CLL cells from 4 patients treated with DMSO, hook, 1 µM ibrutinib or 2 nM NRX-0492 for 18 hours followed by 20 µg/ml anti-IgM stimulation for 6 hours. Median centered heatmap depicts expression of BCR target genes. (D) In vivo activity in the CLL PDX model. BTK degradation and inhibition of cell proliferation assessed by flow cytometry.

In our clinical and laboratory efforts we investigate strategies to strengthen the immune system in patients with CLL. Response to vaccines in CLL patients are often inadequate and can be further reduced in patients undergoing treatment. Vaccines for herpes zoster (RZV, Shingrix) and hepatitis B (HepB-CpG, Heplisav) were approved in 2017 but were untested in CLL patients. These vaccines uniquely trigger a recall immune response, and de novo immune response, respectively. The antibody and cellular response rates to Shingrix, respectively, were 76.8% and 70.0% in treatment-naïve patients and 40% and 41.3% in patients receiving a BTKi (P =.0002). Response to HepB-CpG was nearly absent in CLL patients on BTKi and severely impaired in treatment-naïve patients. During the Covid pandemic, we found that interrupting BTKi therapy can improve response to SARS-CoV2 boosters. We now expand on these observations in a comprehensive vaccine study which provides a mechanism to investigate response to many vaccines in different treatment settings. In these studies, we strive to identify strategies that can improve vaccine responses.

T-cell engaging bispecific antibodies targeting CD3 (on T cells) and CD19 (CD3×CD19) or CD20 (Epcoritamab) on tumor cells. The bispecific antibody recruits autologous (the patient’s own) T-cell cells to kill the CLL cells. In the laboratory, using CLL cells from patients for in vitro experiments, we found that this strategy works well against CLL cells from all patients but was most effective when patients were being treated with a BTKi. We are working on incorporating bispecific antibodies into clinical studies in our program.

Figure 4. Humoral and cellular response to recombinant zoster vaccine (RZV) in treatment-naïve and patients treated with a Bruton Tyrosine Kinase inhibitor.

A New Oral BTK Protein Degrader with Dr. Adrian Wiestner - CLL Society

ASH 2020: Dr. Adrian Wiestner on Early Detection of BTK Mutations Preceding Ibrutinib Relapse in Chronic Lymphocytic Leukemia (CLL) - CLL Society

ASH 2020: Dr. Adrian Wiestner Discusses the Importance of Test Before Treat™ Predictive and Prognostic Biomarker Testing in Patients with Chronic Lymphocytic Leukemia (CLL) - CLL Society

ASH 2018: Dr. Wiestner on the Immune System in CLL (chronic lymphocytic leukemia) - CLL Society

Improve the Response to COVID-19 Vaccination - CLL Society

ASH 2022: Dr. Clare Sun Explains Her Research on Sudden Death Among a Small Number of Chronic Lymphocytic Leukemia (CLL) Patients Using Ibrutinib - CLL Society

Clonal Evolution While on Acalabrutinib for CLL - CLL Society

Observational Study of Cardiac Arrhythmia in Subjects Treated With BTK Inhibitors - CLL Society

Vaccinations in CLL - CLL Society

ASH 2020: Dr. Christopher Pleyer Discusses the Effect of Bruton Tyrosine Kinase Inhibitors on the Efficacy of Adjuvanted Recombinant Hepatitis B and Zoster Vaccines in Patients with Chronic Lymphocytic Leukemia (CLL) - CLL Society

ASH 2021: Effect of Bruton Tyrosine Kinase Inhibitor on Serologic and Cellular Immune Responses to Recombinant Herpes Zoster Vaccine - CLL Society