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1850 Impact of Docirbrutinib (AS-1763) Treatment in CLL: Preclinical Data and Early Clinical Biomarkers

Program: Oral and Poster Abstracts
Session: 641. Chronic Lymphocytic Leukemia: Basic and Translational: Poster I
Hematology Disease Topics & Pathways:
Research, Lymphoid Leukemias, Apoptosis, Translational Research, CLL, Drug development, Diseases, Treatment Considerations, Lymphoid Malignancies, Biological Processes
Saturday, December 7, 2024, 5:30 PM-7:30 PM

Natalia Timofeeva, MD1, Breana Herrera1*, Shady I. Tantawy, MD1, Hitomi Fujiwara, PhD2*, Lizbeth Loza1*, Mariko Hatakeyama2*, Tokiko Asami2*, Hiroshi Ohmoto2*, Kyoko Miyamoto, MD, PhD3*, Yu Nishioka2*, Akinori Arimura3*, Masaaki Sawa, PhD2*, Nitin Jain, MD4 and Varsha Gandhi, PhD1,4

1Department of Experimental Therapeutics, MD Anderson Cancer Center, Houston, TX
2Carna Biosciences, Inc., Kobe, Japan
3CarnaBio USA, Inc., South San Francisco, CA
4Department of Leukemia, MD Anderson Cancer Center, Houston, TX

Background: Non-covalent BTK inhibitors (ncBTKi), such as pirtobrutinib, have emerged as critical therapeutic options for patients (pts) with chronic lymphocytic leukemia (CLL). These ncBTKi are highly relevant as they provide an alternative mechanism of action, bypassing the common resistance associated with covalent BTK inhibitors (cBTKi) that target the C481 residue. However, non-C481 mutations have been identified as contributors to pirtobrutinib resistance, further emphasizing the need for effective ncBTKi therapies. At ASH 2023 our group presented data introducing a potent, highly selective, orally available ncBTKi, AS-1763 (docirbrutinib). Building on these foundational results, our current work expands the study by preclinical and clinical investigations.

Study Design and Methods: We used cell free assay systems to evaluate selectivity and potency of docirbrutinib against 15 BTK-mutants including predicted potential double mutants. We further validated potency of docirbrutinib in cells by measuring inhibition of BTK autophosphorylation in BTK-mutant-transfected HEK293 cell lines. Biological and biochemical effects were tested in treatment-naïve (TN with BTK-wild type) and relapsed/refractory (R/R with BTK and/or Bcl-2 mutant) CLL cells during in vitro investigations with 0.01, 0.1, and 1 µM docirbrutinib alone or with Bcl-2i (venetoclax) or Mcl-1i (AZD5991). Finally, we investigated biomarkers in longitudinal samples obtained from pts during dose-escalation docirbrutinib clinical trial in R/R CLL (NCT05602363 Clinical Trials.gov).

Results: We have generated a total of 15 recombinant BTK mutant proteins (C481, T474, L528, T316 variants and double mutants). Docirbrutinib showed potent inhibitory activities for those BTK mutants while inhibitory potencies of ibrutinib or pirtobrutinib were diminished against some BTK mutants such as C481 or T474 and L528, respectively. Docirbrutinib inhibited BTK autophosphorylation (pY223) in HEK293 cells transfected with those BTK mutants. In proliferation assays, docirbrutinib inhibited cell growth of c/ncBTKi-resistant OCI-Ly10 cells harboring mutant BTKs (C481S, T474I or L528V). In TN CLL lymphocytes from 11 CLL pts, 72-hr incubation with docirbrutinib induced modest yet significant apoptosis which was comparable to ibrutinib or pirtobrutinib. In parallel, B-cell activation, as measured by surface expression of CD86 in cells (n=14 CLL pts), was inhibited with 10 and 100 nM docirbrutinib (p=<0.01). Docirbrutinib effectively inhibited calcium release (n= 10 pts) even at 10 and 100 nM (p=<0.0001). At 1 µM, inhibition was similar to pirtobrutinib (p=0.47) but lower than ibrutinib (p=0.016). In concert, spontaneous (p=0.04) and CXCL12-induced (p=0.007) migration (n=9 pts) was lessened by docirbrutinib yielding results comparable to ibrutinib and greater than pirtobrutinib. Docirbrutinib mitigated phosphorylation of BTKTyr223 and PLCγ2Tyr1217 while reducing levels of MCL-1 and BCL-XL proteins. In R/R patient samples, docirbrutinib inhibited BCR pathway and inhibited cell viability in CLL lymphocytes containing C481S, C481R, L528W, or T474I mutants. Notably, in vitro incubation of R/R CLL cells (n=5) harboring either BTK and/or BCL-2 (G101V, D103E, A113G) mutations were sensitized by docirbrutinib to venetoclax- and AZD5991-induced cell death. Cell death was caspase-dependent and was blocked by QVD. Longitudinal analysis of samples from 3 pts enrolled in the docirbrutinib clinical trial revealed a decrease in plasma concentrations of CCL3/CCL4 during treatment, as well as reduced BTKTyr223 and PLCγ2Tyr1217 phosphorylation and decreased MCL-1 protein levels (Figure 1). One patient exhibited disease progression, accompanied by a resurgence of CCL3/CCL4 levels.

Conclusions: Docirbrutinib, pan-mutant ncBTKi demonstrated equipotent activity against both wild-type and multiple BTK mutants in biochemical and cellular assays, as well as its capacity to disrupt BCR signaling pathway in BTK mutant cell lines. In both TN and R/R primary CLL cells, docirbrutinib effectively blocked BCR pathway signaling and increased sensitivity to Bcl-2 and MCL-1 inhibitors even with diverse BTK-mutant and Bcl-2 mutant backgrounds. In the dose-escalation trial, docirbrutinib treatment showed decreased BCR pathway biomarkers such as CCL3/CCL4 and phospho-BTK and PLCγ2. Updated data will be presented.

Disclosures: Fujiwara: Carna Biosciences: Current Employment. Hatakeyama: : Current employment: Current Employment. Asami: Carna Biosciences: Current Employment, Patents & Royalties. Ohmoto: Carna Biosciences: Current Employment. Miyamoto: CarnaBio USA: Current Employment. Nishioka: Carna Biosciences: Current Employment. Arimura: Carna Biosciences: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees. Sawa: Carna Biosciences: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Jain: Precision Biosciences: Consultancy, Honoraria, Other: Travel Support, Research Funding; Pharmacyclics: Consultancy, Honoraria, Other: Travel Support, Research Funding; MEI Pharma: Consultancy, Honoraria, Other: Travel Support; Aprea Therapeutics: Research Funding; Dialectic Therapeutics: Research Funding; Incyte: Research Funding; Servier: Research Funding; NovalGen: Research Funding; MingSight: Honoraria, Research Funding; Medisix: Research Funding; Takeda: Research Funding; Janssen: Consultancy, Honoraria, Other: Travel Support; Kite, a Gilead Company: Consultancy, Honoraria, Other: Travel Support, Research Funding; Newave: Research Funding; TransThera Sciences: Research Funding; Loxo Oncology: Research Funding; Fate Therapeutics: Research Funding; Pfizer: Research Funding; ADC Therapeutics: Research Funding; Ipsen: Consultancy, Honoraria, Other: Travel Support; Genentech: Consultancy, Honoraria, Other: Travel Support, Research Funding; Cellectis: Consultancy, Honoraria, Other: Travel Support, Research Funding; CareDx: Consultancy, Honoraria, Other: Travel Support; Bristol Myers Squibb: Consultancy, Honoraria, Other: Travel Support, Research Funding; BeiGene: Consultancy, Honoraria, Other: Travel Support; AstraZeneca: Consultancy, Honoraria, Other: Travel Support, Research Funding; Adaptive Biotechnologies: Consultancy, Honoraria, Other: Travel Support, Research Funding; AbbVie: Consultancy, Honoraria, Other: Travel Support, Research Funding; TG Therapeutics: Consultancy, Honoraria, Other: Travel Support. Gandhi: AstraZeneca: Honoraria, Other: travel support; AbbVie: Consultancy, Honoraria, Research Funding; Pharmacyclics: Research Funding; Clear Creek Bio: Consultancy, Research Funding; LOXO/Lilly: Research Funding; Dava Oncology: Honoraria; Sunesis: Honoraria, Research Funding.

*signifies non-member of ASH