Antibody-drug Conjugate Targeting Research Articles

Abstract 5736: AZD9592: An EGFR-cMET bispecific antibody-drug conjugate (ADC) targeting key oncogenic drivers in non-small-cell lung cancer (NSCLC) and beyond

Abstract De novo and acquired drug resistance can limit the long-term efficacy of targeted cancer therapies such as tyrosine kinase inhibitors targeting key oncogenic drivers like EGFR and cMET. Mechanisms of resistance include secondary mutations of EGFR and cMET and other downstream oncogenic pathways such as KRAS and amplification of alternate growth factor receptors. MET amplification or protein overexpression has been established as the most common mechanism of clinical resistance to EGFR inhibitors such as osimertinib. AZD9592 is a first-in-class bispecific ADC designed to target EGFR and cMET, while overcoming pathway-mediated resistance mechanisms that limit other targeted agents. Here we describe the generation, characterization and preclinical evaluation of AZD9592. The ADC was constructed on the backbone of the clinically validated DuetMab monovalent bispecific IgG platform and was engineered with higher affinity for cMET compared to EGFR (>15 fold), with the aim of reducing EGFR-driven toxicity in normal tissues. The antibody is conjugated via a cleavable linker to a proprietary topoisomerase 1 inhibitor (TOP1i) payload (AZ14170132). The internalization and in vitro cytotoxicity (IC50 in the low nM range) of AZD9592 were found to be optimal when both EGFR and cMET were engaged. When EGFR alone was engaged, cytotoxicity was significantly reduced, consistent with the lower affinity for EGFR. Treatment of cells with AZD9592 induced multiple DNA damage response pathway markers (like ATM, ATR, γΗ2ΑX), consistent with the proposed primary mechanism of action (MOA) of direct tumor-cell killing caused by double strand DNA breaks. AZD9592 monotherapy showed activity in vivo in patient-derived xenograft (PDX) models representing multiple EGFR and cMET expressing tumor types, including both EGFR mutant (m) and wild-type NSCLC and head and neck squamous cell carcinoma. Responses (≥30% regression from baseline tumor volume) were observed across a wide range of clinically relevant dose levels, including a 41% response rate in EGFRm NSCLC tumors treated at the lowest tested dose of 2 mg/kg. AZD9592 combined with osimertinib also showed benefit in PDX models derived from patients who progressed on osimertinib alone, as well as models representing primary resistance (EGFR ex20ins). AZD9592 was well tolerated in cynomolgus monkeys over a 6-week period (dosing every 3 weeks). The key safety findings were limited hematological effects, consistent with the MOA of the TOP1i payload. Plasma pharmacokinetics in cynomolgus monkeys showed an acceptable profile at tolerated doses, in line with other EGFR and cMET directed antibodies. These results demonstrate that AZD9592 has a promising efficacy and safety profile in preclinical models representing diverse opportunities in multiple clinical settings. Citation Format: Frank Comer, Yariv Mazor, Elaine Hurt, Chunning Yang, Ryan Fleming, Harini Shandilya, Balakumar Vijayakrishnan, Meghan Sterba, Ruoyan Chen, Edward Rosfjord, Nicolas Floch, Anton I. Rosenbaum, Yue Huang, Jiaqi Yuan, Kevin Beaumont, Lisa Godfrey, Lara McGrath, Fernanda Arnaldez, Puja Sapra. AZD9592: An EGFR-cMET bispecific antibody-drug conjugate (ADC) targeting key oncogenic drivers in non-small-cell lung cancer (NSCLC) and beyond. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5736.

Abstract 6307: A novel pegylated bispecific antibody-drug conjugate (P-BsADCpb-adc) targeting cancers co-expressing PD-L1 and CD47

Abstract ADCs have demonstrated improved efficacy and target selectivity comparing to the non-specific small molecule cytotoxicity drugs in cancer treatment. Yet, in solid tumor therapies, effort for further improvement of efficacy and safety has been hindered by the poor tumor penetration of conventional IgG based ADC, low internalization efficiency, undesired efflux of ADC from tumor cells, narrow therapeutic window due to the on-target/off tumor and Fc induced toxicity, etc. To address those issues, we previously reported that the compound JY201, a Polyethylene Glycol (PEG)-based bispecific ADC (P-BsADC) targeting two epitopes of Her2, demonstrated advantages in tumor penetration, internalization efficiency, lysosome trafficking, no Fc related toxicity, and better efficacy in tumor inhibition than transtuzumab deruxtecan (Ds-8201). In this study, we reveal another novel pegylated P-BsADC (JY207) formed by site-specific conjugation of a bispecific single chain fusion protein targeting PD-L1 and CD47 with PEG-MMAE (a pegylated cytotoxic payload MMAE). As expected, the compound JY207 retains all the advantages that JY201 possesses. Furthermore, JY207 does not bind to human red blood cells, but preferentially binds to CD47/PD-L1 double positive tumor cells, thus reduces the possibility of on-target toxicities. In vitro cytotoxicity studies showed that JY207 has strong potencies in CD47/PD-L1 double positive tumor cells, while showing almost no killing effect to CD47 or PD-L1 single positive tumor cells. In CDX models and a PDX model of transplanted tumor tissues from lung cancer patients, the compound demonstrated excellent tumor inhibition at low doses. In an in vitro plasma stability test, JY207 displays high stability in cynomolgus monkey and human serums. Preliminary repeated-dosing toxicological study has found the maximum tolerated dose of JY207 in CD47/PD-L1 double transgenic mice is 50mg/kg. In vivo pharmacokinetics and toxicological studies of JY207 are being conducted in cynomolgus monkeys and are expected to show desirable results. All those findings in this study warrant JY207 as a promising candidate for the clinical development for patients with CD47/PD-L1 double positive cancers. Citation Format: Shumin Liu, Weidong Lyu, Shuqiang Yin, Yang Lei, Qiudong Zhuo, Liling Zheng, Bin Sun, Shuangyu Tan, Lidong Jiang, Teng Zhang, Bo Gao, Rui Xu, Dechang Huang, Yong Li, Zibin Wu, David Wu, Yvonne (Yu) Wen. A novel pegylated bispecific antibody-drug conjugate (P-BsADCpb-adc) targeting cancers co-expressing PD-L1 and CD47. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6307.

Abstract 4000: A novel pegylated bispecific antibody-drug conjugate (P-BsADC) targeting Her2+ cancers with improved efficacy and therapeutic window

Abstract Despite the fact that ADCs improve the efficacy and target selectivity comparing to the non-specific small molecule cytotoxicity drugs in cancer treatment, traditional ADCs still suffer from many issues which include low tumor penetration and accumulation, inefficient internalization, undesired efflux of ADC from tumor cells, significant on-target off-tumor toxicity, Fc mediated uptake that results in off-target toxicity, limited extravasation across capillary walls due to big molecular size, poor diffusion into the tumor masses due to increased tumor interstitial fluid pressure, and the binding-site-barrier. To address these issues, we previously reported that the compound JY201, a Polyethylene Glycol (PEG)-based bispecific ADC (P-BsADC) targeting two epitopes of Her2, demonstrated advantages in tumor penetration, internalization efficiency, lysosome trafficking effectiveness, no Fc related toxicity, and better efficacy in tumor inhibition than transtuzumab deruxtecan (Ds-8201). Continuing from our previous study, here we further reveal that JY201 can penetrate the tumor deeply and distribute more homogeneously in entire tumor masses while Ds-8201 limits its diffusion to the regions very close to the blood vessels in the tumor. Furthermore, JY201 shows better efficacy than Ds-8201 in inhibiting tumors with low expression of Her2 in pdx (patient derived xenograft) and cdx models. In addition, JY201 can effectively inhibit tumors resistant to Ds-8201. In an in-vitro plasma stability test, JY201 demonstrated high stability in cynomolgus monkey and human serums. JY201 also has a biodistribution profile advocating better safety than Ds-8201 in tumor bearing mice. In the repeated-dosing toxicological study in Her2 transgenic mice, JY201 with the dose of 50mg/kg was well tolerated and did not induce any tissue/organ damage to the animals. Due to much shorter half-life (5 times shorter) in mice for PEGylated proteins than in primates, we expect JY201 will have much higher tolerated dose than the 50mg/kg in primates. In summary, the findings from this study provide solid preclinical evidence for JY201 to be developed further as an efficacious and safe clinical treatment for patients with Her2 positive cancers. Citation Format: Yu (Yvonne) Wen, Shuqiang Yin, Weidong Lyu, Yang Lei, Qiudong Zhuo, Zibin Wu, Bin Sun, Shuangyu Tan, Lidong Jiang, Teng Zhang, Bo Gao, Rui Xu, Yong Li, Liling Zheng, Shumin Liu, David (Dechun) Wu. A novel pegylated bispecific antibody-drug conjugate (P-BsADC) targeting Her2+ cancers with improved efficacy and therapeutic window. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4000.

Abstract 6297: Rationale for the development of a differentiated Trop2 ADC

Abstract Background: Trophoblast antigen 2 (Trop2) or Tumor-Associated Calcium Signal Transducer 2 (TACSTD2) is a transmembrane glycoprotein that is overexpressed in several solid tumor cancers. Antibody drug conjugate (ADC) targeting of Trop2 has been clinically validated. The Trop2 ADC Trodelvy® (also known as Sacituzumab govitecan/IMMU-132) is approved for therapy of TNBC and urothelial cancer, and Datopotamab DXd (DS-1062) is currently being tested in NSCLC. Peak Bio’s Thailanstatin suite of immunomodulatory linker-toxins (L-Ts) is a set of 7 related molecules with distinct ADC features that have been extensively characterized in vitro and in vivo as Her2 ADCs and is being used as a platform to generate a pipeline of potentially differentiated ADCs (AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1832). Results: M2.8 is a fully humanized Peak Bio antibody (Ab) that targets human Trop2 (hTrop2) with high binding affinity [Kd (M)= 8.97-09]. M2.8 was optimized for binding affinity and humanization relative to its parent M2.1 [Kd (M)= 1.78-08], and, both M2.1 and M2.8, display species-specificity for the human and cynomolgus monkey Trop2 proteins. When bound to hTrop2-positive cell lines, M2.1 and M2.8 can rapidly internalize and deliver an ADC payload. To identify the optimal release for therapeutic efficacy, we conjugated our family of L-Ts with hRS7 at a comparable drug-to-antibody ratio (DAR) of 4. Maximal tumor growth inhibition (TGI) was associated with the Trop2 ADCs that were conjugated via amine coupling to lysine residues using non-cleavable L-T L22, later renamed PH1. To determine the effect of antibody on TGI efficacy, different doses, and DARs of M2.1 PH1 ADC were compared to PH1 ADCs made from other Trop2 Abs. At low dose levels, M2.1 PH1 ADCs exhibited comparable TGI with hRS7 PH1, TINA PH1, and T6-16 PH1 ADCs, but exhibited significantly greater TGI than hRS7 SN38 (IMMU-132), suggesting that payload potency, rather than epitope, was important in targeting Trop2. The results also suggested that a PH1 DAR of 4 was sufficient to obtain 90% TGI with multiple anti-Trop2 Abs. M2.1 PH1 ADC was safely tolerated by cynomolgus monkeys at 3 and 6 mg/kg Q3W x 3 doses with mild-to-moderate changes in liver enzymes and platelets that reversed to baseline within 7-10 days of administration of each dose. The optimized M2.8 PH1 DAR4 ADCs killed Trop2-positive cell lines with single-digit nanomolar potency, while exhibiting low off-target cytotoxicity vs normal human skin fibroblasts and Trop2-negative cell lines in vitro. M2.8 PH1 ADC regressed all 200mm3 sized NCI-N87 tumors and 50% of these tumors exhibited long-term regression for 5 months. Conclusion: The target validation studies suggest that Trop2 PH1 ADCs were sufficiently differentiated in areas of preclinical efficacy and well tolerated in a toxicologically relevant host. Citation Format: Satyajit K. Mitra, William Monteith, Mary Do, Greg Tuffy, Scott Savage, Jeffrey Kang, Mastewal Abuhay, Teodora Losic, Sanjeevani Ghone, William E. Haskins, Vasu Jammalamadaka, Sanjeev Satyal. Rationale for the development of a differentiated Trop2 ADC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6297.

Abstract 6320: CUSP06/AMT-707, a new CDH6-targeting antibody-drug conjugate, demonstrates potent antitumor activity in preclinical models

Abstract Cadherin-6 (CDH6), also known as K-cadherin, is a type II classic cadherin molecule that plays an important role in the embryonic development of the kidney, but has very limited expression in adult tissues. It is overexpressed in several human malignancies, primarily in ovarian, renal, but also (to a lesser degree) gastric, thyroid, cholangiocarcinoma and other tumor types. The characteristics of limited expression in normal tissues, high expression in tumor tissues, and rapid internalization upon antibody binding makes CDH6 a well-suited Antibody-drug Conjugate (ADC) target. We developed a novel CDH6-targeting ADC, CUSP06, consisting of a proprietary humanized antibody selective for CDH6, a protease cleavable linker, and an exatecan payload, with a drug-to-antibody ratio (DAR) of 8. CUSP06 selectively bound to cell surface CDH6 and was efficiently internalized into CDH6 positive ovarian and renal cancer cells. CUSP06 exhibited strong antiproliferative activity against several CDH6 positive cancer cell lines in vitro. Furthermore, compared to a DXd-based ADC, a exatecan-based ADC demonstrated improved bystander effect. Treatment with CUSP06 resulted in tumor regression in several CDH6 positive cell line derived xenograft (CDX) models, including PA-1, OVCAR3, and 786-O. In addition, CUSP06 demonstrated potent antitumor activity with tumor regression observed in two CDH6 low and high expressing patient derived xenograft (PDX) models. The preclinical activity of CUSP06 against CDH6-expressing tumors provide compelling support for the clinical development of CUSP06 in CDH6-expressing human cancers. CUSP06 is currently in IND-enabling studies. CUSP06 showed an expected toxicity profile consistent with it's exatecan payload in the ongoing pilot toxicology studies. We plan to initiate a Phase 1 first-in-human clinical trial in the 2nd half of 2023. Citation Format: Wei Lu, Jing Shi, Shu-Hui Liu, Nicole Covino, Xun Meng, Eric D. Slosberg. CUSP06/AMT-707, a new CDH6-targeting antibody-drug conjugate, demonstrates potent antitumor activity in preclinical models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6320.

Abstract 2923: JAB-X1800: A potent immunostimulatory antibody-drug conjugate (iADC) targeting CD73

Abstract Background: STING emerges as a key immune sensor and activated STING pathway plays a key role in innate and adaptive immune response. STING agonist has shown promising potency to stimulate anti-tumor immunity. Considering immune-related toxicity due to systemic administration, as well as difficulty in intra-tumor administration, targeted delivery of STING agonist by iADC is warranted. We developed CD73-STING iADC JAB-X1800, which uses anti-CD73 mAb for targeted delivery of payload JAB-27670, a highly potent non-cyclic dinucleotide STING agonist. Method: Binding assay and HEK293-STING-luciferase reporter assay were performed to test affinity of JAB-27670. Tumor cells with CD73 expression and THP-1 cells were co-cultured in the presences of iADC JAB-X1800 to assess the IFN-β production. The cell killing activity of iADC JAB-X1800 was evaluated when cancer cells co-cultured with human PBMCs. Furthermore, human and mouse plasma stability of iADC JAB-X1800 was assessed. Finally, the anti-tumor effects of iADC JAB-X1800 alone or in combination with anti-PD-1 were evaluated in animal models. Results: JAB-27670 is a highly potent STING agonist with sub-nanomolar EC50 in binding and luciferase reporter assays. In CD73-expressing cancer cells and THP-1 co-culture experiments, iADC JAB-X1800 induced IFN-β production with significant improvement compared with the free payload. JAB-X1800 also induced potent killing in vitro. JAB-X1800 is stable in human and mouse plasma. A single dose of JAB-X1800 induced tumor regression in MDA-MB-231 xenograft model and hCD73-MC38 syngeneic model. JAB-X1800 conferred immunologic memory when tumor free mice were re-challenged. JAB-X1800 showed synergistic anti-tumor activity with anti-PD-1 in hCD73-MC38 syngeneic model. Conclusions: The CD73-targeted STING agonist iADC JAB-X1800 demonstrated high stability in plasma, robust activity, and good tolerability in animal models, supporting clinical development of JAB-X1800 alone and in combination with anti-PD-1 for patients with CD73-expressing tumors. Citation Format: Yang Ye, Guiqun Yang, Hongling Wang, Xin Sun, Yanping Wang, Fashun Yan, Lei Wang, Wei Long. JAB-X1800: A potent immunostimulatory antibody-drug conjugate (iADC) targeting CD73 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2923.

Abstract 4396: TROP2 expression in non-small cell lung cancer

Abstract Introduction: Non-small cell lung cancer (NSCLC) is a major cause of cancer-related deaths globally with high unmet need. A variety of novel therapeutic strategies are being explored to improve patient outcomes including the use of antibody drug conjugates (ADCs) to deliver highly potent, cytotoxic payloads to tumors. Trophoblast cell-surface antigen 2 (TROP2), also known as tumor-associated calcium signal transducer 2 (TACSTD2), has emerged as an attractive target for ADCs. Sacituzumab govitecan-hziy is a novel ADC composed of a TROP2 antibody coupled to SN-38 and is being evaluated in NSCLC. High TROP2 mRNA expression is observed in many tumor types, however, TROP2 protein expression in NSCLC is not well-established. Here we characterized TROP2 expression across three independent datasets to explore the TROP2 relationship to baseline characteristics, molecular features of interest, and its prognostic value in NSCLC. Methods: We analyzed three independent datasets: (1) The Cancer Genome Atlas (TCGA) NSCLC adenocarcinoma (LUAD, N=660) and squamous cell carcinoma (LUSC, N=484) mRNA expression data, (2) a NSCLC adenocarcinoma FFPE tumor sample set (“Translational data set”; N=107), (3) an independent clinical NSCLC adenocarcinoma (N=103) and squamous cell carcinoma (N=37) data set. TROP2 IHC was performed on tumor sample sets (2) and (3) using the SP295 antibody (Robust Prototype Assay, Ventana) with H-Score and assessment of the percentage of membrane-positive tumor cells as readouts. Wilcoxon rank-sum and Kruskal-Wallis tests (continuous variables), Kaplan-Meier method (survival probability curves) and log-rank test (time to event outcome) were used (GraphPad Prism 8.1.2 and R version 4.0.5). Results: TROP2 mRNA was highly expressed in NSCLC and expression was similar in adenocarcinoma and squamous cell carcinoma in the TCGA data set. TROP2 expression did not vary with patient’s age, gender, race or tumor stage. Furthermore, TROP2 expression was not associated with TP53, KRAS or driver mutation status (including EGFR, ALK, ROS, RET, MET). TROP2 expression in the Translational sample set was consistent with the findings from the TCGA data analysis: TROP2 mRNA and protein expression were high and independent of patient baseline demographics, tumor stage and TP53 and KRAS mutation status. Finally, in an independent clinical NSCLC dataset, TROP2 protein was highly expressed and there was no relationship between TROP2 and baseline demographics including gender and age. TROP2 membrane H-scores and percentage of membrane-positive tumor cells also highly correlated in the two independent datasets. Conclusions: Three large independent datasets confirmed high expression of TROP2 in NSCLC both, as mRNA and protein. TROP2 expression in tumors did not differ between histological subtypes, baseline characteristics or clinically relevant driver alterations. Based on the TCGA data set, TROP2 expression was not prognostic for survival. Citation Format: Peiwen Kuo, Emon Elboudwarej, Lauri Diehl, Jilpa Patel, Sabeen Mekan, Juliane M. Jürgensmeier. TROP2 expression in non-small cell lung cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4396.

Abstract 960: Overlapping expression landscape of antibody drug conjugate targets, trophoblast cell surface antigen 2 (Trop-2) & human epidermal growth factor receptor 2 (HER2), in breast cancer

Abstract Background Antibody drug conjugates (ADC) are novel drugs linking potent payloads to antibodies targeting antigen-expressing tumors. Sacituzumab govitecan (SG), targeting Trop-2, is approved for metastatic triple negative breast cancer (TNBC); and trastuzumab deruxtecan, targeting HER2, is approved for HER2-positive and HER2-low metastatic breast cancer. To understand the potentially overlapping clinical landscape of Trop-2 and HER2 antigens, we evaluated RNA expression data in breast cancer from The Cancer Genome Atlas (TCGA) project. Methods TCGA dataset was assessed for Trop-2 and HER2 expression via processed RNA sequencing (RNA-seq) data of the corresponding genes TACSTD2 and ERBB2. Medium/high gene expression was assessed as >100 transcripts per million (TPM). Samples were classified HER2-low per ASCO/CAP guidelines. Gene expression across clinical parameters was assessed via one-way ANOVA. Results 1076 patients with primary breast cancer were included. The majority (59%) had both high TACSTD2 expression (TACSTD2hi) and high ERBB2 expression (ERBB2hi) (see Table). Median TACSTD2 expression was 572 TPM (IQR 349-666 TPM); median ERBB2 expression was 122 TPM (IQR 73-192 TPM). No significant difference was observed in TACSTD2 or ERBB2 expression among invasive ductal carcinoma, invasive lobular carcinoma, mixed histology, or other (p = 0.07, 0.23). No significant difference in TACSTD2 expression was noted between HER2-low and HER2-negative subtypes (p=0.34). Conclusions While SG is approved in TNBC, TACSTD2 is expressed across all breast cancer subtypes, including HER2-low, suggesting a broader population may benefit from Trop-2-targeted ADCs. Furthermore, given that over half of breast cancers have high expression of both TACSTD2 and ERBB2, additional studies are needed to understand the optimal sequencing of ADC-based therapies for patients with breast cancer. Table Patient subsets(HR = hormone receptor) n TACSTD2hi & ERBB2hi TACSTD2hi & ERBB2low TACSTD2low & ERBB2hi TACSTD2low & ERBB2low All 1,076 632 (59%) 394 (36%) 19 (2%) 31 (3%) Histology 843 Invasive ductal carcinoma 507 267 (32%) 215 (25%) 10 (1%) 15 (2%) Invasive lobular carcinoma 130 103 (12%) 24 (3%) 1 (<1%) 2 (<1%) Mixed 91 63 (7%) 24 (3%) 2 (<1%) 2 (<1%) Other 115 65 (8%) 43 (5%) 2 (<1%) 5 (<1%) HER2 Status 345 HR+/HER2-low 254 170 (49%) 77 (22%) 6 (1%) 1 (<1%) TNBC/HER2-low 55 12 (3%) 39 (11%) 1 (<1%) 3 (<1%) HR+/HER2-negative 24 13 (4%) 11 (3%) 0 0 TNBC/HER2-negative 12 1 (<1%) 10 0 1 (<1%) Citation Format: Arielle J. Medford, Nicholas J. Haradhvala, Neelima Vidula, Rachel Abelman, Laura M. Spring, Leif W. Ellisen, Gad Getz, Aditya Bardia. Overlapping expression landscape of antibody drug conjugate targets, trophoblast cell surface antigen 2 (Trop-2) & human epidermal growth factor receptor 2 (HER2), in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 960.

Abstract 562: Anti-tumor efficacy of GMF-1A3, an MMAE-based antibody drug conjugate targeting cell surface cleaved Amphiregulin in breast cancer

Abstract The Epidermal Growth Factor Receptor ligand, Amphiregulin, is a key proliferative effector of estrogen receptor signaling in breast cancer and also plays a role in other malignancies. Amphiregulin is a single-pass transmembrane protein proteolytically processed by TACE/ADAM17 to release the soluble EGFR ligand, leaving a residual transmembrane stalk that is subsequently internalized. Here, we describe the development of an antibody drug conjugate, GMF-1A3-MMAE, targeting an AREG neo-epitope revealed following ADAM17-mediated cleavage. The antibody does not interact with uncleaved Amphiregulin, providing a novel means of targeting cells with high rates of Amphiregulin shedding. Using fluorescent dye conjugation, we demonstrated that the antibody is internalized by cancer cells in a manner dependent on the presence of cell surface cleaved Amphiregulin. Antibodies conjugated with monomethyl auristatin E (MMAE) were cytotoxic in vitro and induced rapid regression of established breast tumor xenografts in immunocompromised mice. We further demonstrate that these antibodies recognize the Amphiregulin neo-epitope in formalin fixed paraffin embedded tumor tissue, suggesting their utility as a companion diagnostic for patient selection. A pan-cancer tissue microarray analysis indicates that the target is commonly detected (> 50% of cases) in breast, prostate, liver and lung cancer. Citation Format: Kristopher A. Lofgren, Sreeja Sreekumar, Paraic A. Kenny. Anti-tumor efficacy of GMF-1A3, an MMAE-based antibody drug conjugate targeting cell surface cleaved Amphiregulin in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 562.

Abstract 2331: Loss of LRRC15 in osteosarcoma cells disrupts extracellular matrix integration, intercellular communication, and differentiation

Abstract Background: Leucine rich repeat containing 15 (LRRC15) is a 581 amino acid type 1 transmembrane protein that contains 15 leucine rich repeats. Cysteine clusters flank the LRR domain that functions as the collagen binding domain. LRRC15 has been found to be upregulated during osteogenic differentiation of mesenchymal stem cells, induced by TGFβ, and highly expressed in multiple solid tumors, notably in osteosarcoma (1). Antibody drug conjugates targeting LRRC15 have shown promising activity in pre-clinical studies (2). Results: To investigate the function of LRRC15 in the osteosarcoma microenvironment, we generated inducible shRNA knockdown (KD) derivatives targeting 3’UTR and ORF regions of LRRC15 of four osteosarcoma cell lines. Using these models, we characterized the biological properties of LRRC15 knockdown under 2-D and 3-D growth conditions. We found that LRRC15 KD in 2-D culture resulted in reduced adhesion to extracellular matrix (ECM) components and impaired cell migration. In 3-D spheroid culture, LRRC15 KD severely impacted scaffold-free spheroid compaction and invasion of spheroids embedded in Matrigel. Dysregulation of multiple ECM, cell adhesion and communication pathways were revealed by transcriptomic analysis from samples collected at time points during the LRRC15 KD. Of interest, several key transcription factors including RUNX2, CTNNB1, and SP7 were downregulated in LRRC15 deficient cells. Conclusion: Our comprehensive results establish the importance of LRRC15 as a critical cell-adhesion molecule essential for the maintenance of the osteogenic intercellular communication, invasion and differentiation revealing novel aspects of osteosarcoma cell-biology. Reference:1.Purcell JW, Tanlimco SG, Hickson J, Fox M, Sho M, Durkin L, et al. LRRC15 Is a Novel Mesenchymal Protein and Stromal Target for Antibody-Drug Conjugates. Cancer Res 2018;78:4059-4072.2.Slemmons KK, Mukherjee S, Meltzer P, Purcell JW, Helman LJ. LRRC15 antibody-drug conjugates show promise as osteosarcoma therapeutics in preclinical studies. Pediatr Blood Cancer 2020:e28771. Citation Format: Sanjit Mukherjee, Ankit Jajoo, Jack Zhu, Marbin Pineda, Robert Walker, James Purcell, Lee J Helman, Paul S. Meltzer. Loss of LRRC15 in osteosarcoma cells disrupts extracellular matrix integration, intercellular communication, and differentiation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2331.

Abstract 559: Expression of molecular targets for antibody-drug conjugates in patients with NSCLC and RET fusions

Abstract Background RET fusions are found in 1-2% of patients (pts) with advanced non-small cell lung cancer (NSCLC). RET inhibitors are highly effective, but resistance eventually occurs. Antibody-drug conjugates (ADC) are promising treatments in NSCLC, that could be employed at time of resistance. Data on the membranous expression of ADC targets, such as EGFR/HER2/HER3 and the possible impact of novel therapeutic agents in RET+ NSCLC are lacking. Methods We conducted a single-center observational study, collecting tissue specimens from 19 patients (pts) affected by RET+ NSCLC. The expression of EGFR/HER2/HER3 proteins on the cellular membrane was evaluated by immunohistochemistry (IHC). EGFR expression was assessed by using the DAKO EGFR pharmDx kit (Dako, Glostrup, Denmark); HER2 staining (4B5 clone by Roche clone) was evaluated using ASCO/CAP guidelines in scale from 0 to 3+; HER3 staining (D22C5 clone by Cell Signaling) was categorized by intensity as 0, 1+, 2+, and 3+, according to the same criteria used for HER2 evaluation. All IHC analyses were performed at Gustave Roussy. Results Out of 19 pts, 18 were evaluable for EGFR expression, 18 for HER2, 17 for HER3. All analyses were conducted on archival tissue samples from tumor biopsies performed predominantly in treatment-naïve pts (17/19). Pts had adenocarcinoma in 5/19 cases, KIF5B fusion partner in 13/19 cases. Median PD-L1 expression in evaluable pts was 15% (IQR 1- 60.00). EGFR expression was high, moderate, and low in 4 (22%), 2 (11%) and 5 (27%) cases, respectively. HER3 expression was 2+ in 4 (24%) cases, 1+ in 5 (29%) pts and 0 in the other 47%. None had HER2 membranous expression. EGFR expression was more frequently found in pts with uncommon fusion partners (6/11 vs 0/7, p=0.038) and non-adenocarcinoma histology (5/11 vs 0/7, p=0.10). HER3 expression evaluated at 2+ were identified in 3 cases with KIF5B-RET fusion and adenocarcinoma and 1 case with ANKRD26-RET fusion and undifferentiated histology. PD-L1 expression did not show significant difference between pts with positive vs negative EGFR or HER3 staining (p=1). Conclusions EGFR and HER3 had variable expression in RET+ NSCLC, deserving further exploration for novel treatment strategies. HER2 expression was not found in our RET cohort. Citation Format: Arianna Marinello, Maria Rosa Ghigna, Anas Gazzah, Karine Godefroy, Ludovic Lacroix, Damien Vasseur, Jordi Remon-Masip, Fabrice Barlesi, David Planchard, Jean-Yves Scoazec, Benjamin Besse, Mihaela Aldea. Expression of molecular targets for antibody-drug conjugates in patients with NSCLC and RET fusions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 559.

Abstract 4894: The anti-ROR1 ADC STRO-003 demonstrates immune-modulating properties that may enhance checkpoint blockade

Abstract There is growing evidence that tumor-targeted cytotoxins can also enhance anti-tumor immunity by inducing immunogenic cell death (ICD) in tumor cells and promoting recruitment of immune effector cells. We sought to investigate the immune stimulating potential of STRO-003, an antibody drug conjugate (ADC) composed of an anti-Receptor Orphan Receptor Kinase 1 (ROR1) antibody conjugated to a topoisomerase I targeted exatecan warhead via a cleavable linker. ROR1 is an oncofetal transmembrane receptor with restricted expression in normal tissues. Its overexpression in several cancer indications has been described, including in ovarian, non-small cell lung carcinoma (NSCLC), triple negative breast cancer (TNBC), and hematological malignancies, thus making it an ideal ADC target. We have previously demonstrated potent in vivo activity of STRO-003 in a panel of ROR1-expressing lung cancer patient-derived xenograft (PDX) models. Here we show that the exatecan warhead, SC3386, and STRO-003 ADC induced ICD in vitro as evidenced by presentation of cell-surface calreticulin and release of HMGB1. Consistent with this finding, STRO-003-treated cells elicited monocyte activation in a PBMC co-culture assay. To determine if these immunogenic properties could improve therapeutic efficacy, we evaluated STRO-003 in combination with an immune checkpoint inhibitor in a mouse syngeneic model expressing ROR1. We have demonstrated that STRO-003 in combination with checkpoint inhibition significantly enhances efficacy in a syngeneic mouse model and supports durable anti-tumor immunity. Follow-up vaccination studies were performed to further explore the significance of STRO-003-induced ICD and protective immunity in vivo. These results demonstrate that tumor cells pre-treated with STRO-003 or SC3386 undergo potent immunogenic cell damage which can, in turn, mount protective immunity in vivo. Citation Format: Alice Yam, Helena Kiefel, Andrew McGeehan, Robert Yuan, Sihong Zhou, Dayson Moreira, Indrani Dutta, Xiaofan Li, Cuong Tran, Gang Yin, Kristin Bedard, Trevor Hallam. The anti-ROR1 ADC STRO-003 demonstrates immune-modulating properties that may enhance checkpoint blockade. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4894.

Abstract 4673: Therapeutic potential of polatuzumab vedotin for the treatment of diffuse large B-cell lymphoma

Abstract Background Diffuse large B-cell lymphoma (DLBCL) is typically treated with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP). Although most patients can be cured with R-CHOP, up to one-third of them relapses with a dismal outcome in most cases. Numerous approaches have been attempted to improve the treatment outcomes with R-CHOP. Polatuzumab vedotin is an antibody-drug conjugate targeting CD79b, which is ubiquitously expressed on over 90% of B-cell NHL malignancies, including DLBCL. To determine the potential of polatuzumab vedotin as a therapeutic agent, we evaluate its potency across a series of DLBCL patient derived xenograft (PDX) models. Method We established and characterized a series of DLBCL lymphoma PDX models. Eight DLBCL PDX models were selected for polatuzumab vedotin efficacy study, of which three are GCB DLBCL. Gene expression analysis of these lymphoma PDX models was performed using an Illumina NovaSeq 6000 system following Illumina-provided protocols for 2 × 150 paired-end sequencing. The expression status of CD79b on these DLBCL PDX models was also evaluated by IHC. Conclusion Our data showed that a single dose of polatuzumab vedotin at just 2 mg/kg could reached a comparable effect with R-CHOP and no significant body weight loss was observed. Our result showed that there is no apparent correlation between polatuzumab vedotin responses (TGI) and the CD79b expression (IHC Score). Although polatuzumab vedotin demonstrated encouraging activity in the treatment of DLBCL, some of the PDX models are also resistant to polatuzumab vedotin treatment. To further investigate the molecular parameters of the relative sensitivity of polatuzumab vedotin. Gene expression profiling of those PDX models and their sensitivity to polatuzumab vedotin were conducted, we found that the expression level of BCL-XL was correlated with reduced sensitivity to polatuzumab vedotin. Citation Format: Hui Qi, Fuyang Wang, Bingrui Han, Xiaomin Wang, Xiangnan Qiang, Zhixiang Zhang, Qiangyang Gu. Therapeutic potential of polatuzumab vedotin for the treatment of diffuse large B-cell lymphoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4673.

Abstract 1876: NAV-001, a high-efficacy antibody-drug conjugate targeting mesothelin with improved delivery of potent payload by counteracting MUC16/CA125 effects

Abstract Subsets of tumor-produced proteins, referred to as Humoral Immuno-Oncology (HIO) factors, can bind to IgG1 antibodies and suppress their immune-effector activities. Antibody-drug conjugates (ADCs) targeting tumor cell surface antigens can internalize and kill target cells upon liberation of their cytotoxic payload. Binding of the antibody component of an ADC by a HIO factor may potentially hamper the efficacy of a bound ADC. To assess any suppression of ADC’s activity by HIO factors, we evaluated the efficacy of a HIO-refractory ADC, NAV-001, and a HIO-bound ADC, SS1, both targeting mesothelin. The HIO factor MUC16/CA125 protein, which binds to SS1 ADC, was shown to have a negative effect on its internalization and tumor cell killing. The MUC16/CA125 refractory NAV-001 ADC was shown to have robust killing of tumor cells regardless of MUC16/CA125 expression in vitro (EC50 ~20 pM) as well as in vivo at single, sub-mg/kg dosing. Moreover, NAV-001-PNU, which contains the PNU-159682 topoisomerase II inhibitor as a payload, demonstrated good stability in vitro and in vivo as well as robust bystander activity against tumor and stromal cells not expressing the target antigen mesothelin, while maintaining a tolerable safety profile in vivo. Single-dose NAV-001-PNU demonstrated robust tumor regression of a number of patient-derived xenograft models from different tumor types regardless of MUC16/CA125 expression. These findings suggest that identification of HIO-refractory antibodies to be used in ADC format may improve therapeutic efficacy as observed for NAV-001 and warrants NAV-001-PNU’s advancement to human clinical trials as a monotherapy to treat mesothelin-positive cancers. Citation Format: Luigi Grasso, J. Bradford Kline, Nicholas C. Nicolaides. NAV-001, a high-efficacy antibody-drug conjugate targeting mesothelin with improved delivery of potent payload by counteracting MUC16/CA125 effects [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1876.

Abstract 3995: Trastuzumab deruxtecan, antibody-drug conjugate targeting HER2, effectively inhibits growth of patient-derived xenograft model with CIC-rearranged sarcoma

Abstract Background: Capicua-double homeobox 4 (CIC-DUX4)-rearranged sarcomas (CDS) are extremely rare and highly aggressive sarcomas. There is no standard therapy for the patients with advanced CDS, and therapeutic development is needed. We evaluated preclinical efficacy of trastuzumab deruxtecan (T-DXd), a humanized monoclonal HER2-targeting antibody conjugated to a topoisomerase 1 inhibitor, DXd, in patient-derived xenograft (PDX) models with CDS. Methods: Patient-derived tumor tissue was transplanted into subcutaneous around the flank of female NOG mice, which were treated with vehicle or T-DXd (3 mg/kg, intravenous, Day0) when the mean tumor volume reached 200 mm3. Tumor volume was assessed twice weekly for 3 weeks to assess the efficacy of T-DXd. Results: This study included 3 CDS-derived PDXs. HER2 expression was low in one PDX and not expressed in two PDXs. One PDX was established from a specimen at initial diagnosis, two were established from specimens after prior-chemotherapy. T-DXd demonstrated significant tumor growth delay compared to vehicle in all PDX models investigated. One PDX with no efficacy was HER2-negative and had a treatment history of topoisomerase I inhibitor. In contrast, PDX of HER2-negative topoisomerase I inhibitor-resistant Ewing sarcoma was also administered T-DXd under the same conditions and was found to be refractory. Conclusion: The present study showed that a therapeutic potential of T-DXd in CDS patients, including HER2-negative. Citation Format: Yuki Kojima, Shigehiro Yagishita, Kazuki Sudo, Tatsunori Shimoi, Shintaro Iwata, Shun-ichi Watanabe, Chitose Ogawa, Akihiko Yoshida, Yasushi Yatabe, Kan Yonemori, Akinobu Hamada. Trastuzumab deruxtecan, antibody-drug conjugate targeting HER2, effectively inhibits growth of patient-derived xenograft model with CIC-rearranged sarcoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3995.

Abstract 1544: MBK-103, a potent novel conjugation platform-based antibody-drug conjugate, changing therapeutic options in folate receptor alpha positive cancer patients

Abstract Folate receptor alpha (FOLR1) is a clinically validated target, that is overexpressed extracellularly in numerous solid malignancies with a high unmet medical need. Ovarian, non-small cell lung and breast adenocarcinomas are among indications with the highest frequency of FOLR1-positive patients with one out of two patients showing upregulated topoisomerase I expression. MBK-103, an antibody-drug conjugate targeting FOLR1, is based on Mablink’s novel proprietary hydrophilic drug-linker platform. It is comprised of: (i) an Fc-attenuated humanized IgG1 monoclonal antibody, that selectively binds to FOLR1; (ii) a polysarcosine hydrophobicity masking entity allowing for a high drug-to-antibody ratio (DAR) of 8, while improving the pharmacokinetics and tolerability of the drug; (iii) a proprietary dipeptide cleavable unit and iv) exatecan, a potent topoisomerase I inhibitor. MBK-103 showed excellent ex vivo stability in human plasma enabling a prolonged half-life, similar to that of the unconjugated monoclonal antibody. This led to an increased drug exposure, resulting in a potent antitumor efficacy observed in a collection of more than ten in vivo mouse tumor models with a diverse FOLR1 expression, already at doses as low as 1-3 mg/kg. In addition, the distinct mode of action of exatecan along with the unique characteristics of the proprietary drug-linker platform resulted in the in vivo antitumor potency obtained also in colorectal cancer models, in which tubulin-inhibiting agents exhibit only limited therapeutic effects. The compound was also very well tolerated in cynomolgus monkeys with HNSTD at 50 mg/kg and even when administered repeatedly. High conjugation yields along with 100% homogeneity are major drivers of the successful ongoing manufacturing process with planned IND submission in Q4/2023. Citation Format: Warren Viricel, Louise Conilh, Edouard Leroy, Jean-Guillaume Lafay, Frederique Brune, Lenka Kyrych Sadilkova, Charles Dumontet. MBK-103, a potent novel conjugation platform-based antibody-drug conjugate, changing therapeutic options in folate receptor alpha positive cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1544.

Abstract 1604: Preclinical development of ADCT-211, a novel pyrrolobenzodiazepine dimer-based antibody-drug conjugate targeting solid tumors expressing IL13RA2

Abstract Interleukin 13 receptor subunit alpha 2 (IL13RA2) is one of the two major receptors for the cytokine interleukin 13 (IL-13). While IL13RA1 has low affinity for IL-13 and it is expressed ubiquitously in humans, IL13RA2 has high binding affinity to IL-13 and its expression in normal tissues is mainly restricted to the testes. IL13RA2 was initially regarded as a decoy receptor because it has a short cytoplasmic tail with no known signaling motifs, and its binding to IL-13 does not lead to activation of the JAK/STAT6 pathway. However, recent studies demonstrated that IL-13-mediated IL13RA2 signaling occurs via STAT6-independent pathways. IL13RA2 is reported to be expressed at a high frequency in glioblastoma multiforme (GBM) as well as in other solid tumors including melanoma, renal cell carcinoma and adrenocortical carcinoma and is correlated with poor prognosis. ADCT-211 is an antibody-drug conjugate composed of HuCl47, a humanized IgG1 antibody directed against human IL13RA2, site-specifically conjugated using GlycoConnectTM technology to PL1801, which contains HydraspaceTM, a valine-alanine cleavable linker and the PBD dimer cytotoxin SG2000 (drug to antibody ratio ~ 1.8). The purpose of this study was to characterize the in vitro and in vivo anti-tumor activity of ADCT-211 in human cancer cell lines and xenograft models, to determine its safety, tolerability and pharmacokinetic (PK) in the rat and to measure IL13RA2 expression by immunohistochemistry (IHC) in human tumor specimens. HuCl47 showed specific binding to recombinant IL13RA2, while it did not bind to IL13RA1. In vitro, ADCT-211 had highly potent and targeted cytotoxicity against various IL13RA2-expressing solid cancer cell lines. In vivo, a single dose of ADCT-211 at 10 mg/kg was able to completely eradicate tumors in the subcutaneous (sc) A375 human melanoma xenograft model and resulted in 8/8 tumor-free survivors (TFS) at the end of the study on day 57. In the sc U251 human glioblastoma xenograft model, a single dose of ADCT-211 at 3.75 mg/kg showed potent and durable antitumor activity and it resulted in 5/10 partial responders and 5/10 complete responders, 2 of which were TFS at the end of the study on day 51. In an MTD study in male rats, ADCT-211 was stable and tolerated up to 24 mg/kg single dose, with exposure data being indicative of a linear PK profile with a half-life of 12-19 days. Expression of membranous IL13RA2 was confirmed by IHC in a panel of primary and refractory GBM samples and malignant melanoma, highlighting them as potential indications for the clinical development of ADCT-211. In conclusion, ADCT-211 demonstrated potent and specific in vitro and in vivo anti-tumor activity and it was stable and well tolerated in the rat, warranting further development of ADCT-211 into the clinic in IL13RA2-expressing cancers. Citation Format: Francesca Zammarchi, Karin Havenith, Ben Leatherdale, Britanny Roberts, Lara Montolio, Afroze Patel, Narinder Janghra, Pedro Alves, Kristina Zaitseva, Cecile Oblette, Ian Kirby, Lolke de Haan, Patrick H van Berkel. Preclinical development of ADCT-211, a novel pyrrolobenzodiazepine dimer-based antibody-drug conjugate targeting solid tumors expressing IL13RA2 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1604.

Abstract 4890: A novel topoisomerase I inhibitor antibody-drug conjugate targeting CEACAM5 has potent anti-tumor activity in colorectal cancer models

Abstract Carcinoembryonic antigen cell adhesion molecule 5, CEACAM5, is a glycosylphosphatidylinositol-anchored glycoprotein highly expressed on the cell surface of several epithelial tumors. CEACAM5 is expressed in virtually all colorectal cancer, ~90% of which to high levels while normal tissue expression is limited. The high prevalence of CEACAM5 expression in colorectal tumor cells prompted us to develop an investigational anti-CEACAM5 antibody-drug conjugate (ADC) for the potential treatment of CRC patients. We developed a novel ADC, by conjugating an anti-CEACAM5 antibody with a drug linker to a topoisomerase I inhibitor payload. The anti-CEACAM5 antibody was chosen based on its high selectivity for CEACAM5 and its potential to direct cytotoxic payloads to tumor. The topoisomerase I payload was optimized for potency, reduced PGP efflux and enhanced bystander activity. The novel anti-CEACAM5 topoisomerase I inhibitor ADC binds to CEACAM5 at nanomolar (nM) concentrations and kills CEACAM5-positive colon tumor cells with varying levels of CEACAM5 at sub-nM concentrations with no or very low cytotoxicity towards CEACAM5-negative cells. Mechanistically, the potent anti-tumor activity of the ADC is both mediated by direct internalization, processing, and release of the cytotoxic payload within the CEACAM5-expressing tumor cells, and by a bystander effect mediated by diffusion of the payload to the neighboring CEACAM5-negative tumor cells. The novel anti-CEACAM5 topoisomerase I inhibitor ADC is well tolerated in rats after repeated administration of 30 and 50 mg/kg/day, Q1W x 4. In vivo efficacy of this ADC at 1, 3 and 10 mg/kg (single administration) was evaluated in four CRC patient-derived xenografts (PDXs) models. The conjugate elicits potent and specific and dose dependent antitumor activity with complete regression (CR) at 10 mg/kg in the 4 models. This robust anti-tumor activity was further confirmed in a Single Mouse Trial of 16 CRC PDX models, consisting in the use of one animal per PDX model per treatment arm and for which the evaluation of efficacy was based on the criteria RECIST (Response Evaluation Criteria In Solid Tumors) used in clinic. In these criteria, the overall response rate includes complete response (CR) and partial response (PR) and the a disease control rate includes CR, PR and stable disease (SD). The ADC induces a disease control rate of 95% and an overall response rate of 50% following a single dose of 10 mg/kg. The outstanding anti-tumor activity across CRC PDX models and its favorable safety profile in rats support further evaluation of this investigational novel topoisomerase I ADC in CRC patients. Citation Format: Yves Baudat, Haley Neff-LaFord, Celine Nicolazzi, Dave Meyer, Johann Petur Sigurjonsson, Ryan Lyski, Valeria Fantin, Marie-Priscille Brun, Marielle Chiron, Stephanie Decary. A novel topoisomerase I inhibitor antibody-drug conjugate targeting CEACAM5 has potent anti-tumor activity in colorectal cancer models. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4890.

Abstract 2967: Preclinical development of a B7-H3-targeting ADC with a novel DNA topoisomerase I inhibitor for solid tumors

Abstract Introduction: B7-H3 (CD276) is an immunomodulatory protein of the B7 superfamily molecules, it is overexpressed in a wide range of solid tumors. B7-H3 overexpression often correlates with poor survival and higher tumor grade, it has become an attractive target for the development of novel cancer therapies recently. B7-H3 targeting antibody-drug conjugates (ADCs), such as MGC018 and DS7300a, have shown preliminary clinical efficacy. DB-1311 is a B7-H3 targeting ADC comprised of a humanized B7-H3 antibody conjugated to a novel DNA topoisomerase I inhibitor P1021 via a cleavable linker. We herein report preclinical efficacy and safety data of DB-1311 and its differentiation from DS7300a. Methods: The binding of anti-B7-H3 to other B7 family members were evaluated via ELISA and SPR. Anti-B7-H3 antibody was conjugated to P1021 with different Drug-Antibody-Ratio (DAR) values. The efficacy of B7-H3 ADCs with different DAR values was tested in vivo, pharmacokinetics and safety profile were evaluated in cynomolgus monkeys. Based on potency and toxicity analysis, DB-1311, an ADC with DAR6, was selected for preclinical development. In vitro and in vivo studies were then conducted with DB-1311 to further investigate its pharmacological effects. Results: B7-H3 antibody with DAR6 demonstrated higher efficacy than its DAR4 counterpart in lung cancer (Calu6), glioma(U87MG) and prostate cancer (PC-3) models. In monkeys, the DAR6 ADC showed good stability and dose-proportionally increased exposure. Additionally, it was well tolerated with repeat dose administration up to 80 mg/kg and didn’t show significantly worse toxicities than DAR4 counterpart. In preclinical study, DB-1311 demonstrated specific, dose-dependent cytotoxicity toward B7-H3-positive tumor cell lines in vitro and potent antitumor activity in vivo but minimum tumor growth inhibition activity in a B7-H3 negative xenograft model. Moreover, DB-1311 showed more potent antitumor activity than DS7300a analogue in vitro and in vivo. Conclusions: DB-1311, a preclinical B7-H3 ADC candidate, with a DAR value of approximately 6, exhibited potent cytotoxicity in vitro and antitumor activity in vivo toward a range of B7-H3-expressing tumor cell lines representing several cancer types and was well tolerated in cynomolgus monkeys with promising pharmacokinetics characteristics. The preclinical antitumor activity and safety profile warrants further development of DB-1311 to evaluate its efficacy and safety in clinical studies. Citation Format: Chenggang Li, Jun Yao, Chen Qu, Bing Li, Yu Zhang, Rong Shi, Yang Qiu, Haiqing Hua. Preclinical development of a B7-H3-targeting ADC with a novel DNA topoisomerase I inhibitor for solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2967.

Abstract 1884: DB-1310, a novel Her3 targeting antibody-drug conjugate, exhibits therapeutic efficacy for solid tumors

Abstract Background: HER3 (ErbB3), a member of the human epidermal growth factor receptor family, is frequently overexpressed in various cancers. Its high expression is associated with poor prognosis in several solid tumors. Many anti-Her3 antibodies have been developed for cancer therapy. To date, only patritumab deruxtecan (U3-1402) showed clinical efficacy in both NSCLC and breast cancer among all Her3-targeting therapies. DB-1310 is a HER3 ADC composed of a novel humanized anti-Her3 immunoglobulin G1 (IgG1) monoclonal antibody, covalently linked to a proprietary DNA topoisomerase I inhibitor payload via a maleimide tetrapeptide-based cleavable linker with a drug antibody ratio (DAR) of approximately eight. In the present study, we aim to evaluate the efficacy and safety of DB-1310 in preclinical models. Methods: The binding of DB-1310 to Her3 and other HER families was measured by ELISA and SPR. The competition of binding epitope for DB-1310 and patritumab was tested by FACS. The sensitivity of breast cancer and lung cancer cell lines to DB-1310 was evaluated by in vitro cell killing assay. In vivo growth inhibition study evaluated the sensitivity of DB-1310 to Her3-positive breast cancer and lung cancer xenograft models. In addition, the efficacy of DB-1310 was compared to that of patritumab deruxtecan in tumor xenograft models. The pharmacokinetics and safety profile were also measured in cynomolgus monkey. Results: In this study, DB-1310 showed high affinity to both Human and Cyno Her3 without cross reacting with Her2, Her4 or EGFR. DB-1310 binds Her3 using a different epitope from patritumab. DB-1310 has high internalization capability and translocates into lysosomes after internalization. In vitro, DB-1310 exhibited cytotoxicity in a panel of Her3- expression human breast cancer and lung cancer cell lines. It also showed dose-dependent antitumor killing activity in a HER3-positive breast cancer HCC1569 xenograft model and a lung cancer NCI-H441 xenograft model. Tumor regression was observed in a NSCLC patient-derived xenograft model with EGFR mutation under DB-1310 treatment. Moreover, DB-1310 showed stronger tumor growth inhibition than that of patritumab deruxtecan at the same dose. The systemic exposure AUC0-last and Cmax of DB-1310 and total antibody increased dose proportionally from 1 mg/kg to 10 mg/kg with low released payload in serum. The DB-1310 also showed good safety profile with highest non-severely toxic dose (HNSTD) above 30mg/kg. Conclusions: DB-1310 exhibited its efficacy in tumor models both in vitro and in vivo. These studies suggest that DB-1310 as a novel HER3-targeting ADC showed antitumor efficacy for HER3 positive tumors with good safety profile. Citation Format: Xi Li, Jun Yao, Chen Qu, Luo Lan, Bing Li, Yu Zhang, Rong Shi, Haiqing Hua, Yang Qiu. DB-1310, a novel Her3 targeting antibody-drug conjugate, exhibits therapeutic efficacy for solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1884.