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Purpose: Despite approval of B-cell lymphoma (BCL)-2 inhibitor venetoclax for certain hematologic malignancies, its broader clinical benefit is curtailed by resistance. Our study aimed to determine if treatment with novel anticancer agents targeting BCL-2 and mouse double minute 2 (MDM2) could overcome venetoclax resistance in preclinical models. Experimental Design: Venetoclax-sensitive and venetoclax-resistant acute myeloid leukemia (AML) and acute lymphoblastic leukemia cells and xenograft models were used to evaluate antitumor effects and underlying mechanisms associated with combined BCL-2 inhibitor lisaftoclax (APG-2575) and MDM2 inhibitor alrizomadlin (APG-115). Results: The combination exhibited synergistic antiproliferative and apoptogenic activities in TP53 wild-type AML cell lines in vitro. This synergy was further exemplified by deep antitumor responses and prolonged survival in AML cell line–derived and patient-derived xenograft models. Interestingly, the combination treatment resensitized (to apoptosis) venetoclax-resistant cellular and mouse models established via chronic drug exposure or genetically engineered with clinically relevant BCL-2 gene mutations. Synergistic effects in reducing cellular viability and proliferation were also demonstrated in primary samples of patients with venetoclax-resistant AML treated with lisaftoclax and alrizomadlin ex vivo. Mechanistically, alrizomadlin likely primes cancer cells to BCL-2 inhibition-induced cellular apoptosis by downregulating expression of antiapoptotic proteins myeloid cell leukemia-1 and BCL–extra-large and upregulating pro-death BCL-2–associated X protein. Conclusions: Lisaftoclax in combination with alrizomadlin overcomes venetoclax resistance mediated by various mechanisms, including BCL-2 mutations. In addition, we posit further, putative molecular mechanisms. Our data rationalize clinical development of this treatment combination in patients with diseases that are insensitive or resistant to venetoclax.

Supplementary Figure and Table legends

Supplementary Table 1. Clinical characteristics of patients with relapsed or refractory acute myeloid leukemia.

Purpose:Despite approval of B-cell lymphoma (BCL)-2 inhibitor venetoclax for certain hematologic malignancies, its broader clinical benefit is curtailed by resistance. Our study aimed to determine if treatment with novel anticancer agents targeting BCL-2 and mouse double minute 2 (MDM2) could overcome venetoclax resistance in preclinical models.Experimental Design:Venetoclax-sensitive and venetoclax-resistant acute myeloid leukemia (AML) and acute lymphoblastic leukemia cells and xenograft models were used to evaluate antitumor effects and underlying mechanisms associated with combined BCL-2 inhibitor lisaftoclax (APG-2575) and MDM2 inhibitor alrizomadlin (APG-115).Results:The combination exhibited synergistic antiproliferative and apoptogenic activities in TP53 wild-type AML cell lines in vitro. This synergy was further exemplified by deep antitumor responses and prolonged survival in AML cell line–derived and patient-derived xenograft models. Interestingly, the combination treatment resensitized (to apoptosis) venetoclax-resistant cellular and mouse models established via chronic drug exposure or genetically engineered with clinically relevant BCL-2 gene mutations. Synergistic effects in reducing cellular viability and proliferation were also demonstrated in primary samples of patients with venetoclax-resistant AML treated with lisaftoclax and alrizomadlin ex vivo. Mechanistically, alrizomadlin likely primes cancer cells to BCL-2 inhibition-induced cellular apoptosis by downregulating expression of antiapoptotic proteins myeloid cell leukemia-1 and BCL–extra-large and upregulating pro-death BCL-2–associated X protein.Conclusions:Lisaftoclax in combination with alrizomadlin overcomes venetoclax resistance mediated by various mechanisms, including BCL-2 mutations. In addition, we posit further, putative molecular mechanisms. Our data rationalize clinical development of this treatment combination in patients with diseases that are insensitive or resistant to venetoclax.

Supplementary Figure 1. Synergistic antitumor activity of lisaftoclax combined with alrizomadlin in AML CDX mouse models. Supplementary Figure 2. Mean plasma and tumor drug concentrations in a subcutaneous OCI-AML-3 CDX model in mice. Supplementary Figure 3. Upregulation of P53 and BAX and downregulation of MCL-1 and BCL-xL by lisaftoclax combined with alrizomadlin in RS4;11cells expressing BCL-2G101V mutation. Supplementary Figure 4. Characterization of RS4;11 cells expressing BCL-2 resistant mutants. Supplementary Figure 5. Synergistic antiproliferative activity of combined treatment with lisaftoclax and alrizomadlin in parental RS4;11 cells, RS4;11 cells with WT BCL-2, and venetoclax-resistant RS4;11 cells expressing BCL-2 mutants.

This work established a finite element analysis (FEA) model of an inserted tooth-type slip assembly under bear setting load and axial load, calculated the differences between the inserted teeth of the sidetracking packer slip-formed furrow shapes on the casing face, and analyzed the setting reliability of the inserted tooth slip sidetracking packer. The orthogonal optimization analysis of the structural parameters of the sidetracking packer was carried out on the basis of the furrow effect. Finally, the setting experiment was conducted with the inserted tooth slip sidetracking packer developed to verify correctness of the FEA model and the simulation results. The results show that in the FEA and calculation of the setting process of the inserted tooth-type slip, it is not only necessary to consider the furrow friction coefficient, but also the effect of the ridge on the furrow friction coefficient. The corresponding furrow friction coefficient varies according to the different furrow effects occurring on the casing surface caused by the various types of teeth inserted on the packer slips. The furrow effect is related to the sharpness of the tooth tips of the slips. The sharper the tooth tips, the more obvious the furrow effect is. Under the dual effects of the furrow effect and the adhesion effect, the carbide teeth of the slip feed into the casing wall to produce a uniform and distinct indentation on the premise of meeting the inserted tooth strength to ensure a reliable setting and hanging the inserted tooth slip sidetracking packer. The optimal combination of slip parameters was obtained by taking the optimal bite depth uniformity as the objective function: slip tooth installation spacing L = 10 mm, slip tooth installation angle α = 80°, slip tooth diameter d = 10 mm, and slip wedge angle β = 6°. The standard deviation of bite depth uniformity of the optimized slip teeth is 74.45% lower than that before optimization. The research results of this paper basically meet the requirements of engineering applications.