2632 Background: BB-1701, a HER2-targeting antibody-drug conjugate (ADC) containing eribulin, has demonstrated promising antitumor activity in the clinical studies in breast cancer (BC) patients with HER2 low expression and non-small cell lung cancer (NSCLC) patients with HER2 mutation. Currently, there are no approved treatment options of ADC in combination with anti-PD-1 antibody for metastatic BC patients with HER2 low expression and NSCLC patients with HER2 mutation. We report the preliminary efficacy and safety results from the ongoing phase 2 study (including dose escalation and dose expansion) of BB-1701 in combination with Sintilimab (an anti-PD-1 antibody) in advanced or metastatic BC patients with HER2 low expression and NSCLC patients with HER2 mutation. Methods: Patients enrolled were ≥18 years of age; had confirmed locally advanced/metastatic HER2 low-expressing BC or HER2 mutated NSCLC, an ECOG PS <2 and measurable lesion(s) (per RECIST v1.1); and disease progression after ≥1 lines of prior standard therapies. HER2 expression was confirmed by IHC or NGS/PCR before patient enrollment. BB-1701 was administered at 1.2 mg/kg Q3W or 1.6 mg/kg Q3W, and sintilimab is administered at 200 mg Q3W. Results: As of 26 January 2026, a total of 12 patients with HER2 low-expressing BC or HER2 mutated NSCLC have been enrolled and treated, 6 patients at each dose level of BB-1701 during dose escalation. Median age is 63 years, 91.7%/8.3% patients were female/male, and 16.7%/83.3% patients have ECOG PS 0/1. The median number of prior systemic therapy lines was 2.0/1.0 for BC and NSCLC. All patients experienced at least one treatment-emergent adverse events (TEAEs). The most common (≥20%) reported all grade TEAEs are alanine aminotransferase increased, aspartate aminotransferase increased, γ-glutamyltransferase increased, hypercholesterolemia, peripheral neuropathy, anemia and Urinary tract infection. Three grade 3 TEAEs are γ-glutamyltransferase increased, pneumonia and open globe injury. There has been no grade 4 or grade 5 events as of data cut-off date. One treatment emergent serious adverse event is open globe injury. All patients were evaluable for efficacy. Among 5 BC patients, 4 patients achieved partial response (PR) with disease control rate (DCR) of 80.0%. Among 7 NSCLC patients, 2 patients achieved PR and 2 patients had stable disease (SD) with DCR of 57.1%. Details of data will be presented at the ASCO meeting. Conclusions: BB-1701 in combination with Sintilimab shows encouraging antitumor activity and a manageable safety profile in HER2 low-expressing BC patients and HER2 mutated NSCLC patients. The further dose expansion study is guaranteed based on the preliminary data from dose escalation study. Clinical trial information: CTR20241422.

Abstract Co-expression of EGFR and HER3, along with other tumor-associated antigen (TAA such as c-MET, HER2, B7H3, PD-L1, or others), is frequently observed across diverse human tumors. Targeting more than two TAAs simultaneously presents engineering challenges but offers distinct advantages: increased tumor cell avidity for ADCs, circumvention of antigen-loss-driven resistance, synergistic blockade of cross-talk between signaling pathways, mitigation of tumor heterogeneity, and broadened applicability across cancer types. A tri-specific ADC platform has been developed by combining one arm with both HER3 and EGFR affinities and another arm with affinity for a third TAA. Binding and internalization were assessed in cell lines with varying TAA expression levels to evaluate uptake efficiency. in vitro cytotoxicity was measured via dose-response assays across multiple lines with modest expression of each TAA. in vivo efficacy was tested in murine xenograft models representing diverse target antigen profiles, in comparison with benchmarks Izalonatamab brengitecan (BL-B01D1, EGFR/HER3 bispecific ADC) and an Amivantamab (EGFR/c-MET bispecific mAb )-derived ADC. Our results show that tri-specific ADCs exhibit significantly faster and more efficient internalization than the EGFR/HER3 bispecific ADC, particularly in cells with medium-to-low EGFR and HER3 expression. in vitro, tri-specific ADCs achieved lower IC50 values, indicating enhanced cytotoxic potency across a broad panel of tumor lines. In xenograft models, tri-specific ADCs delivered superior tumor growth inhibition relative to the bispecific benchmark, irrespective of heterogeneous target antigen expression. In conclusion, tri-specific ADCs offer a robust advantage in vitro and in vivo over EGFR/HER3 or c-MET bispecifics by enabling enhanced tumor cell binding, improved internalization, and more effective delivery of cytotoxic agents. This strategy holds promise for broader clinical application in tumors with varied receptor expressions and may address resistance in heterogeneous cancers. Citation Format: Yang Wang, Lixia Cao, Cui Feng, Fangdun Jiang, lixia Gu, Chen Li, Yifan Yang, Qi Zhang, Ming Zhou, Cancan Li, Wei Huang, Bonan Yan, Ziping Wei, Yuhong Zhou, . Tri-specific ADC platform offers potential therapeutic advantage over bi-specific ADC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 6939.

Abstract Targeting death receptor DR5 with agonist antibody could trigger extrinsic apoptosis via caspase activation in cancer cells while sparing normal tissues. Ozekibart (INBRX109), a tetra-valent anti-DR5 antibody, showed significant efficacy as a monotherapy in patients with advanced or metastatic, unresectable chondrosarcoma, with disease control rate (DCR) of 54%, (compared to 27.5% in the placebo group). Later in a study of advanced Ewing sarcoma, Ozekibart combined with irinotecan and temozolomide demonstrated improved clinical benefit with an ORR of 64% and a DCR of 92%. Encouraged with these results, we designed a DR5-targeting ADC with the aim to synergize the extrinsic apoptosis activity of DR5 engaging and superb cytotoxicity effect of Top-1 inhibitor. In this presentation we show systematic evaluation of this novel ADC including specific tumor cell engaging, anti-tumor efficacy, serum stability, and pharmacokinetics (PK) in target-humanized mice. First, the unique design of the tri-valent DR5-targeting antibody demonstrated superior activity over counterparts. Second,its resulting ADC form showed a robust tumor growth inhibition across diverse xenografts, potentially benefit from its dual mode of action design, surpassing clinically advanced control agents (INBRX109). Our ADC showed dose dependent in vivo antitumor activity without hook effect seen with agonist antibodies. Finally, this ADC showed better in vivo safety profile: lower on-target off-tumor apoptosis, longer half-life and better tolerability in comparison with that of INBRX109,suggesting a potentially wider therapeutic window. These results may support progression of this novel DR5-tageting ADC into clinical development as a potentially more potent and safer DR5-targeted therapy. Citation Format: Yang Wang, Lixia Cao, Cui Feng, Yifan Yang, Fangdun Jiang, Lixia Gu, Qi Zhang, Chen Li, Ming Zhou, Cancan Li, Wei Huang, Bonan Yan, Ziping Wei, Yuhong Zhou, . A DR5-targeting ADC exhibits superior anti-tumor activity and potentially better therapeutic window [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 2637.

Abstract Background: T-cell engagers (TCEs) represent a promising therapeutic modality, however their clinical translation to solid tumors is severely hampered by Cytokine Release Syndrome (CRS). This dose-limiting toxicity has contributed to the failure of numerous TCE trials. Previous conditional activation strategies, such as MMP-cleavable, pH-dependent, or polymer-masked pro-TCEs, were limited by tumor microenvironment heterogeneity, inconsistent activation, and immunogenicity. Here we report a first-in-class, β-glucuronidase-activatable pro-TCE platform featuring an optimal TCE-Mask ratio (TMR) designed to overcome these limitations through complete tumor-microenvironment activation. Methods: Utilizing site-specific conjugation, we generated a library of TMRx-pro-TCEs by attaching 2∼10 β-glucuronidase-cleavable masking peptides to various TAA/CD3 bispecific and trispecific antibodies and screened for optimal masking efficiency. in vitro TCE activity assessments included TAA-engaged T-cell activation and cytotoxicity against different types of tumor cells. In vivo efficacy and safety were evaluated in hPBMC-reconstituted tumor-bearing mice by monitoring tumor growth inhibition, plasma cytokine levels (IL-6, IFN-γ), body weight and survival. Results: Masking efficiency was precisely controlled by TMR, with optimal TMR values conferring the strongest suppression of nonspecific T-cell activation. Following β-glucuronidase cleavage of the masking peptides, pro-TCEs completely restored activities to the level of unmasked TCEs and demonstrated potent, target-specific cytotoxicity. Talatamab is a market approval TCE for DLL3-expressing small cell lung cancer treatment. As a POC example, an optimized TMR4-pro-Talatamab was constructed and demonstrated markedly attenuation of CRS in hPBMC-reconstituted mice. The same pro-Talatamab also inhibited tumor growth to the level equivalent to that of unmasked Talatamab, demonstrating full retention of TCE capability. Conclusions: This novel pro-TCE platform shows potential as a solution to the central challenge of CRS in TCE therapy. By enabling precise tumor-microenvironment activation, it preserves full antitumor efficacy while fundamentally improving systemic safety which may offer a transformative strategy for the effective translation of TCEs to solid tumor indications. Citation Format: Yang Wang, Wei Huang, Lixia Cao, Cui Feng, Lixia Gu, Fangdun Jiang, Yifan Yang, Qi Zhang, Chen Li, Ming Zhou, Cancan Li, Bonan Yan, Ziping Wei, Yuhong Zhou. A novel tumor-microenvironment-responsive pro-TCE therapy minimizes cytokine release for safer solid tumor treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(8_Suppl):Abstract nr LB259.

HER3 has emerged as a critical player in the resistance mechanisms against therapies targeting the epidermal growth factor receptor (EGFR). Its upregulation can activate alternative signaling pathways, enabling cancer cells to bypass the effects of EGFR inhibitors, which often results in treatment failure. To combat this issue, novel HER3-targeting therapies are under investigation, including antibody-drug conjugates (ADCs) such as Patritumab Deruxtecan (U3-1402), which have shown encouraging results in clinical trials, underscoring the potential of HER3 targeting to overcome resistance. The development of bispecific ADCs that target both HER3 and EGFR represents a pioneering approach that may enhance therapeutic efficacy through dual targeting. Izalonatamab brengitecan (BL-B01D1), an ADC designed for simultaneous targeting of EGFR and HER3, has demonstrated substantial clinical benefits in heavily pretreated patients with EGFR-mutant non-small cell lung cancer (NSCLC). Building on this strategy, we are advancing a novel bispecific ADC, BR-2302, which employs a human IgG1 format bi-specific antibody with high affinity for HER3 and moderate affinity for EGFR. This innovative design aims to maximize therapeutic effectiveness while minimizing on-target/off-tumor toxicity associated with the widespread expression of EGFR in normal tissues. Our comprehensive studies on BR-2302 have focused on its in vitro cytotoxicity and in vivo tumor inhibitory effects across various cancer cell lines and xenograft models expressing the target antigens. BR-2302 exhibited significant anti-tumor activity in vitro, achieving subnanomolar IC50 values against HER3-expressing cell lines regardless of their EGFR expression levels. in vivo experiments demonstrated remarkable tumor growth inhibition and sustained regression in human lung and colon cancer xenograft models, surpassing the tumor inhibitory effects of established benchmark ADCs. Collectively, our findings indicate that BR-2302 possesses promising anti-tumor properties. This innovative therapeutic approach has the potential to improve treatment outcomes for patients with solid tumors expressing both HER3 and EGFR, warranting further clinical exploration. Yang Wang, Bing Xia, Lixia Cao, Jianfeng Yang, Cui Feng, Fangdun Jiang, Chen Li, Lixia Gu, Yifan Yang, Jing Tian, Jiasheng Geng, Qi Zhang, Ziping Wei, Yuhong Zhou. Preclinical studies of BR-2302: A HER3xEGFR bi-specific Exd-containing ADC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3417.

CD73, also known as ecto-5'-nucleotidase, is a cell surface enzyme that catalyzes the conversion of AMP to adenosine. It is highly expressed in various types of cancers, including gastric, glioblastoma, non-small cell lung, and colorectal cancers. Elevated CD73 expression is highly correlated with poor prognosis, increased metastasis, and resistance to chemotherapy. In the tumor microenvironment, CD73-derived adenosine exerts multiple immunosuppressive effects, promoting tumor growth and metastasis. Currently, there are no approved antibody-drug conjugates (ADCs) targeting CD73. In this study, we introduce BB-1709, a CD73-targeting ADC that incorporates MMAE as its cytotoxic payload, demonstrating its antitumor activity through direct cytotoxicity against CD73-expressing tumor cells and its ability to inhibit CD73 enzyme activity. A comprehensive set of studies was conducted to assess its in vitro cytotoxicity, in vivo antitumor activity as well as its ability to inhibit CD73 enzyme activity. In vitro studies demonstrated that BB-1709 exhibited potent and selective cytotoxicity against various CD73-positive cancer cells and significantly inhibited CD73 enzyme activity, effectively blocking the conversion of AMP into adenosine. In vivo studies using xenograft models showed that a single dose of BB-1709 resulted in substantial antitumor activity. Pharmacokinetic and toxicology assessments in non-human primates revealed a favorable safety profile for BB-1709 at clinically relevant dosages and administration schedules. In summary, these results suggest that BB-1709 not only exhibits good preclinical efficacy and safety, but also has the potential to promote anti-tumor immune responses by inhibiting CD73 enzymatic activity. A phase I clinical trial of BB-1709 is currently underway to further explore its therapeutic potential in cancer treatment. Bing Xia, Lixia Cao, Jianfeng Yang, Cui Feng, Fangdun Jiang, Chen Li, Lixia Gu, Yifan Yang, Jing Tian, Jiasheng Geng, Ziping Wei, Yuhong Zhou. Preclinical studies of BB-1709, a CD73-targeting ADC potentially suppresses tumor growth through dual mechanisms. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2243.

1093 Background: BB-1701 is an HER2-targeting antibody-drug conjugate (ADC) containing eribulin. In phase I study, BB-1701 had shown promising antitumor activity in breast cancer (BC) patients with HER2 high/low expression and manageable safety profile. Currently, there are no approved treatment options for metastatic BC patients with high/low expression who have received HER2-ADC containing TOP-I inhibitor, especially for trastuzumab deruxtecan. We report the preliminary efficacy and safety results from the ongoing phase 2 study of BB-1701 in advanced or metastatic breast cancer patients with HER2 expression previously treated with HER2-ADC containing TOP-I inhibitor. Methods: Patients enrolled were ≥18 years of age; had confirmed locally advanced/metastatic HER2 expressing breast cancer; disease progression after previous HER2-targeting ADC (containing TOP-I inhibitor) therapy; an ECOG PS < 2; and measurable lesion(s) (per RECIST v1.1). HER2 expression was confirmed by IHC before patient enrollment. BB-1701 is administered at 1.6 mg/kg Q3W. Results: As of 28 January 2025, 23 patients with HER2 high/low-expressing breast cancer have been enrolled and treated. Median age is 51 years, all patients are female, and 26.1%/73.9% patients have ECOG PS 0/1. The median number of prior systemic therapy lines was 4.0, 21.7%/78.3% HER2 status are high expression/low expression. All patients experienced at least one treatment-emergent adverse events (TEAEs). The most common (≥10%) all grade TEAES are neutrophil count decreased, platelet count decreased, Aspartate aminotransferase increased, and white blood cell count decreased. One grade 3 TEAEs is peripheral neuropathy, and another grade 3 TEAE is neutrophil count decreased. There has been no grade 4 or grade 5 events as of data cut-off date. One treatment emergent serious adverse event is peripheral neuropathy. Of the 23 patients, 14 were evaluable for efficacy. Among 14 evaluated patients, 4 patients achieved partial response (PR) and 9 patients had stable disease (SD), with disease control rate (DCR) of 92.8%. Among 3 HER2 high-expressing patients who were previously treated with trastuzumab deruxtecan, 1 patient achieved PR and 2 patients had SD with DCR of 100.0%. Among 8 HER2 low-expressioning (IHC 1+) patients, 3 patients achieved PR (2 patients received prior trastuzumab deruxtecan and 1 patient received prior SHR-A1811) and 4 patients had SD with DCR of 87.5%. More data will be presented at the ASCO meeting. Conclusions: BB-1701 shows promising antitumor activity and a manageable safety profile in HER2 expressing breast cancer patients who had previously been treated with HER2-ADC (containing TOP-I inhibitor). Clinical trial information: CTR20241422 .

Epidermal Growth Factor Receptor (EGFR) is a type I receptor tyrosine kinase critical for the development and normal function of epithelial cells. However, its overexpression or hyperactivity in certain cancers can lead to tumor growth and metastasis. While existing EGFR-targeting therapies can effectively inhibit tumor progression, they often cause significant toxicity due to EGFR expression in normal tissues. To mitigate these side effects, we have developed BB-1705, an antibody-drug conjugate (ADC) that utilizes eribulin as its cytotoxic agent. Eribulin is an FDA-approved treatment for metastatic breast cancer and advanced liposarcoma. BB-1705 employs an affinity-optimized anti-EGFR antibody to selectively deliver eribulin to tumor cells, sparing normal tissues by not effectively blocking the EGFR signaling pathway in those areas. This strategy aims to enhance the therapeutic index of EGFR-targeted treatments while minimizing adverse effects on normal tissues. In vitro studies demonstrated that BB-1705 exhibits potent cytotoxicity against a range of cancer cell lines with medium to high levels of EGFR expression, while showing reduced efficacy against normal human keratinocytes, likely due to their lower EGFR expression and the antibody's affinity profile. Notably, BB-1705 also induced a significant bystander effect on EGFR-low/null cells when co-cultured or co-inoculated with EGFR-high cells. In vivo experiments using xenograft models revealed that a single dose of BB-1705 resulted in substantial antitumor activity. When administered repeatedly via a clinically relevant route and schedule, BB-1705 demonstrated a favorable pharmacokinetic profile and safety in cynomolgus monkeys with manageable hematological side effects and minimal skin toxicity observed even at the highest tested dose (12 mg/kg, Q3W x 3). In summary, our preclinical findings suggest that BB-1705 has the potential to optimize the efficacy and safety profile of EGFR-targeting therapies for cancers characterized by medium to high EGFR expression levels. A clinical trial evaluating BB-1705 is currently underway, aiming to assess its therapeutic potential in treatment of EGFR-expressing human cancers including TNBC, NSCLC and head and neck cancers. Bing Xia, Lixia Cao, Jianfeng Yang, Cui Feng, Fangdun Jiang, Chen Li, Lixia Gu, Yifan Yang, Jing Tian, Jiasheng Geng, Likun Gong, Tingting Liu, Qiuping Qin, Ziping Wei, Yuhong Zhou. Preclinical studies of BB-1705: An EGFR-targeting eribulin-containing ADC with an affinity optimized antibody to minimize potential toxicities to normal tissues [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4779.

B7-H3, a member of the B7 family of proteins, is overexpressed in various solid tumors, including small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC), while exhibiting minimal expression in healthy tissues. This unique expression profile positions B7-H3 as a promising target for antibody-drug conjugates (ADCs). Multiple B7H3-targeting ADCs are currently being tested in different stages of clinical trials and several of them have reported antitumor activities in heavily treated patients. In this study, we introduce BB-1712, a novel B7-H3-targeting ADC that incorporates eribulin as its cytotoxic payload, with the potential to treat patients not sensitive to ADCs containing DNA topoisomerase I inhibitors as payload toxin such as DS-7300a. BB-1712 was designed to leverage eribulin's distinct mechanism of action, which effectively inhibits tumor growth through multiple mode of actions including promotion of apoptosis, tumor microenvironment modulation, immunogenic cell death and bystander killing. We conducted extensive preclinical evaluations of BB-1712, focusing on its in vitro cytotoxicity and in vivo tumor inhibitory activities against a range of B7-H3-expressing cancer cell lines and xenograft models.Our results demonstrated that BB-1712 exhibited binding activity to B7-H3-positive cell lines similarly to that of Infinatamab, and displayed potent cytotoxic effects across multiple human cancer models, including non-small cell lung cancer, breast cancer, gastric cancer and small cell lung cancer cell lines. In xenograft studies, BB-1712 achieved significant tumor regression at well-tolerated doses without obvious toxicity. Notably, BB-1712 effectively overcame resistance to DS-7300a in both SCLC and NSCLC models previously treated with this agent. Pharmacokinetic and toxicology assessments in non-human primates revealed a favorable safety profile for BB-1712 at clinically relevant dosages and administration schedules.In summary, our findings suggest that BB-1712 has promising preclinical efficacy and safety profiles as well as the potential to circumvent resistance mechanisms associated with current therapies. These results support the advancement of BB-1712 into clinical trials for patients with B7-H3-expressing solid tumors. Yang Wang, Bing Xia, Lixia Cao, Jianfeng Yang, Cui Feng, Fangdun Jiang, Chen Li, Lixia Gu, Yifan Yang, Jing Tian, Jiasheng Geng, Qi Zhang, Ziping Wei, Yuhong Zhou. Preclinical studies of BB-1712: A B7H3-targeting eribulin-containing ADC with the potential to overcome resistance to DNA topoisomerase I inhibitor-containing ADCs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6173.

Background: Several HER2-targeting antibody-drug conjugates (ADC) have gained market approval for the treatment of HER2-expressing metastasis. Promising responses have been reported with the new generation of ADCs in patients who do not respond well to other HER2-targeting therapeutics. However, these ADCs still face challenges of resistance and/or severe adverse effects associated with their particular payload toxins. Eribulin, a therapeutic agent for the treatment of metastatic breast cancer and liposarcoma, is a new choice of ADC payload with a distinct mechanism of action and safety profile. Methods: We've generated a novel HER2-tageting eribulin-containing ADC, BB-1701. The potency of BB-1701 was tested in vitro and in vivo against cancer cells where HER2-expressing levels vary in a large range. Bystander killing effect and toxin-induced immunogenic cell death (ICD) of BB-1701 were also tested. Results: In comparison with HER2-targeting ADCs with DM1 and Dxd payload, eribulin-containing ADC demonstrated higher in vitro cytotoxicity in HER2-low cancer cell lines. BB-1701 also effectively suppressed tumors in models resistant to DM1 or Dxd containing ADCs. Mode of action studies showed that BB-1701 had a significant bystander effect on HER2-null cells adjacent to HER2-high cells. In addition, BB-1701 treatment induced ICD. Repeated doses of BB-1701 in nonhuman primates showed favorable pharmacokinetics and safety profiles at the intended clinical dosage, route of administration, and schedule. Conclusions: The preclinical data support the test of BB-1701 in patients with various HER2-expressing cancers, including those resistant to other HER2-targeting ADCs. A phase I clinical trial of BB-1701 (NCT04257110) in patients is currently underway. Statement of Significance: A HER2-targeting eribulin-containing ADC had exhibited good plasma stability; BB-1701 had potent antitumor activities against multiple HER2-expressing cancers, especially those insensitive to other HER2-targeting ADCs; BB-1701 had bystander killing effect and induced immunogenic cell death; BB-1701 also showed favorable pharmacokinetics and safety profiles in nonhuman primates.

Satellite Communication on the Move (SOTM) is essential for dynamic environments, such as military operations and disaster relief. Conventionalsystems with parabolic reflector antennas suffer from slow mechanical scanningand reduced tracking efficiency, especially for low Earth orbit (LEO) satellites,while phased array antennas offer faster, more precise tracking but remain underutilized. A key challenge in SOTM is achieving precise beam alignment betweenthe satellite and moving terminals, which directly impacts tracking accuracy.Several traditional existing beam alignment techniques are computationallyexpensive and impractical for real-time use. In this work, we propose a novel andefficient beam alignment algorithm that exploits the inherent low-rank structureof the beamspace energy matrix. By modeling the alignment process as amaximization problem with non-convex constraints, we leverage randomizedmatrix approximation to reconstruct a low-rank representation of the beamspace.This allows us to identify a subspace containing the optimal beam direction. Wethen use Particle Swarm Optimization (PSO) to refine the beam directionestimate. As illustrated in the numerical simulations, our approach improvesbeam alignment accuracy while reducing computational complexity comparing totraditional methods. This advancement has broad implications for enhancing theperformance of SOTM systems in dynamic environments.

1110 Background: BB-1701 is an antibody-drug conjugate (ADC) consisting of a humanized anti-HER2 antibody and eribulin. After BB-1701 is internalized in HER2 expressing cancer cells, free eribulin is cleaved from the ADC by cathepsin b and causes cytotoxicity to cancer cells. The neighboring cells are affected with bystander effects, like cytotoxicity on tumor cells and non-cytotoxic effects on microenvironment by free eribulin. Here, we report updated results from the ongoing cohort expansion of the first-in-human phase I study (NCT 04257110). Methods: Patients enrolled were ≥18 years of age; had confirmed locally advanced/metastatic HER2 low-expressing (IHC 2+ and FISH negative, or IHC 1+; tested before enrollment) breast cancer; disease progression after ≥2 lines of prior standard therapies; an ECOG PS <2; and measurable lesion(s) (per RECIST v1.1). Four dose levels of BB-1701 (1.0, 1.2, 1.4 and 1.6 mg/kg) were administered during the cohort expansion. Results: As of 31 January 2024, 40 pts with HER2 low-expressing breast cancer were enrolled, 5 at 1.0 mg/kg, 20 at 1.2 mg/kg, 5 at 1.4 mg/kg and 10 at 1.6 mg/kg. Median age was 55 years, 97.5% ps were female, and 30.0%/70.0% pts had ECOG PS 0/1. The median number of prior lines of systemic therapy was 3. The most common (≥20%) all grade treatment-related adverse events (TRAEs) were peripheral neuropathy, AST increased, ALT increased, anemia, WBC decreased and hypertriglyceridemia. The most common (≥ 5%) grade 3 TRAEs were peripheral neuropathy and peripheral sensory neuropathy. There were no grade 4 or grade 5 events. Four treatment related serious adverse events were peripheral sensory neuropathy, anemia, spinal compression fracture and infusion related reaction. One patient discontinued BB-1701 treatment due to peripheral neuropathy. Of the 40 pts, 38 were evaluable for efficacy. At 1.0 mg/kg dose level, there were 2 partial response (PR), 2 stable disease (SD), with disease control rate (DCR) of 80.0% and 6-month PFS rate of 20%. At 1.2 mg/kg dose level, there were 5 PR, 11 SD with DCR of 88.9% and 6-month PFS rate of 50%. At 1.4 mg/kg dose level, there were 2 PR, 2 SD with DCR of 80.0% and 6-month PFS rate of 40%. At 1.6 mg/kg dose level, there were 1 complete response (CR), 2 PR with objective response rate of 30.0%, DCR of 60.0% and 6-month PFS rate of 50%. Among the 10 pts at 1.6 mg/kg dose level, 1 pt who had previously been treated with disitamab vedotin maintain CR and has been treated by BB-1701 for 9 months, and 1 pt who had previously been treated with trastuzumab deruxtecan maintain PR and has been treated by BB-1701 for 12 months. Conclusions: BB-1701 shows promising antitumor activity and a manageable safety profile in HER2 low-expressing breast cancer, including patients who received prior anti-HER2 ADCs. Clinical trial information: NCT04257110 .

Abstract Background: BB-1701 is an antibody-drug conjugate (ADC) consisting of a humanized IgG1κ monoclonal anti-HER2 antibody with the same sequence as trastuzumab and eribulin linked together by a cathepsin-cleavable valine-citrulline linker. Eribulin is an inhibitor of microtubule approved for the treatment of metastatic breast cancer and advanced liposarcoma, following prior therapies. When BB-1701 targets HER2 expressing cancer cells and it is internalized, free eribulin is cleaved from the ADC by cathepsin b and causes cytotoxicity to cancer cells. The neighboring cells were affected with bystander effects, like cytotoxicity on tumor cells and non-cytotoxic effects on microenvironment, by free eribulin released from dying cancer cells. Here, we report preliminary results from the ongoing cohort expansion of a phase I study of BB-1701 in patients with HER2 low breast cancer (ClinicalTrials.gov Identifier: NCT 04257110). Methods: Patients ≥ 18 years of age with confirmed locally advanced/metastatic HER2 low breast cancer (IHC 2+ and FISH negative or IHC 1+), that had progressed on at least two lines of prior standard therapies, with ECOG PS ≤ 2 and measurable disease (RECIST 1.1) were enrolled. HER2 expressions was determined by IHC and FISH before enrollment. Total of four dose levels of BB-1701 (1.0 mg/kg Q3W, 1.2 mg/kg Q3W, 1.4 mg/kg Q3W and 1.6 mg/kg Q3W) were administered during the cohort expansion. The safety, tolerability and preliminary anti-tumor activity of BB-1701 were assessed during study. Radiographic assessment was conducted every 6 weeks. Results: As of 30 June 2023, 40 patients with HER2 low breast cancer were enrolled in the four dose levels, 5 patients at 1.0 mg/kg Q3W, 20 patients at 1.2 mg/kg Q3W, 5 patients at 1.4 mg/kg Q3W and 10 patients at 1.6 mg/kg Q3W. Median age was 55 years (range 30-74), 97.5% were female, and 30.0%/70.0% patients were ECOG PS 0/1. The median prior lines of systemic therapy were 3 (range 2 – 9). All subjects experienced at least one treatment emergent adverse events and treatment-related adverse events (TRAEs) occurred in 30 subjects. The most common all grade TRAEs (≥20%) were peripheral neuropathy, aspartate aminotransferase increased, alanine aminotransferase increased, anemia, white blood cell count decreased. Grade 3 TRAEs were peripheral neuropathy (3 subjects), peripheral sensory neuropathy (2 subjects), neutrophil count decreased (2 subjects), platelet count decreased (1 subject), white blood cell count decreased (1 subject). There were no grade 4 or grade 5 events. The treatment related serious adverse events (SAEs) were peripheral sensory neuropathy (1 subject) and infusion related reaction (1 subject). There was no interstitial lung disease reported. No patients discontinued the study treatment due to AE. Of the 40 patients enrolled, 38 patients were evaluable for antitumor activity. At 1.0 mg/kg dose level, of 5 patients treated, 2 achieved partial response (PR), 2 had stable disease (SD), best overall response rate (BOR) was 40.0% (2/5) and disease control rate (DCR) was 80.0% (4/5). At 1.2 mg/kg dose level, of 18 patients treated, 5 achieved PR, 11 had SD with BOR of 27.8% (5/18) and DCR of 88.9% (16/18). At 1.4 mg/kg dose level, of 5 patients treated, 2 achieved PR, 2 had SD with BOR of 40.0% (2/5), and DCR was 80.0% (4/5). One patient who achieved PR was previously progressed on sacituzumab govitecan. At 1.6 mg/kg dose level, of 10 patients treated, 1 achieved CR, 3 achieved PR, 2 had SD with BOR of 40.0% (4/10), and DCR of 60.0% (6/10). Among these patients, the patient who achieved CR was previously treated by disitamab vedotin, and one patient who achieved PR was previously treated by trastuzumab deruxtecan. Conclusions: BB-1701 has demonstrated promising preliminary antitumor activity in HER2 low breast cancer including patients who received prior anti-HER2 ADCs, and a manageable safety profile. Citation Format: Fei Ma, Pamela Munster, Xiaoxiang Guan, Jingfen Wang, Herui Yao, Erika Hamilton, Xian Wang, Yehui Shi, Xinjun Liang, Nong Xu, Huoling Tang, Yuhong Zhou, Ziping Wei. Report on Cohort Expansion of Phase I Study to Investigate the Safety, Tolerability, Pharmacokinetics and Antitumor Activity of BB-1701 in Patients with Locally Advanced/Metastatic HER2 Low Breast Cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr RF02-05.

Several HER2-targeting antibody-drug conjugates (ADC) have gained market approval for the treatment of HER2-expressing metastasis. However, these ADC drugs still face challenges of acquired resistance and/or severe adverse effects associate with their payload toxins. Eribulin, a market approval agent for the treatment of metastatic breast cancer and liposarcoma , is a new choice of ADC payload with distinct mechanism of action (MOA). Here we report preclinical studies of BB-1701, a novel HER2-tageting eribulin-containing ADC. BB-1701 exhibited potent and specific cytotoxicity activities to various types of cancer cells where HER2-exprssing levels vary in large ranges. BB-1701 induced tumor-growth suppression, even regression in in vivo models with a single dose of 1-5 mg/kg dosage. In comparison with other HER2-targeting ADCs with different toxin payloads (DM1, MMAE and Dxd), eribulin-containing ADC demonstrated higher potency to cancer cell lines with low HER2 expressing levels (with lower IC50). BB-1701 also showed effective tumor suppression in models insensitive to T-DM1 or T-Dxd. Mode of action studies revealed significant bystander cytotoxicity of BB-1701 to HER2-low cells when co-cultured with HER2-high cells. Additionally, BB-1701 treatment increased ATP releasing and Calreticulin expression, both were deemed immunogenic cell death (ICD) markers. Lastly, repeat dose pharmacokinetic and toxicology studies in nonhuman primates revealed favorable PK and safety profiles of BB-1701 in the clinical intend dosage route and schedule. In conclusion, these comprehensive preclinical data have strongly supported BB-1701 as a potent antitumor agent to cancers including HER2-low expressing and/or cancers insensitive to other HER2-targeting ADCs. A phase I clinical study of BB-1701 in those patients is ongoing. Citation Format: Yang Wang, Bing Xia, Lixia Cao, Jianfeng Yang, Cui Feng, Fangdun Jiang, Chen Li, Lixia Gu, Yifan Yang, Jing Tian, Cindy Cheng, Keiji Furuuchi, James Fulmer, Arielle Verdi, Katherine Rybinski, Allis Soto, Earl Albone, Toshimitsu Uenaka, Likun Gong, Tingting Liu, Qiuping Qin, Ziping Wei, Yuhong Zhou. Preclinical development of BB-1701, a HER2-tageting eribulin-containing ADC with potent bystander effect and ICD activity [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 1887.

3029 Background: BB-1701 is an antibody-drug conjugate consisting of a humanized IgG1κ monoclonal antibody and eribulin. When BB-1701 targets HER2-expressing cancer cells and is internalized, free eribulin is cleaved from the ADC by cathepsin b and causes cytotoxicity to cancer cells. The neighboring cells were affected with bystander effects, like cytotoxicity on tumor cells and non-cytotoxic effects on microenvironment by free eribulin. Here, we report preliminary results from this phase 1 study (NCT04257110). Methods: This dose escalation study followed an accelerated titration and traditional "3 + 3" design with 6 dose levels from 0.4 to 2.6 mg/kg Q3W. Patients with confirmed advanced/metastatic HER2 overexpressing solid tumors, who had progressed on, or were intolerant to prior standard therapies, aged ≥ 18 years, with ECOG PS ≤ 2, and measurable disease, were enrolled. HER2 expressions were determined by IHC or next generation sequencing. Tumor imaging was conducted every 9 weeks. The primary objective is the assessment of safety and tolerability, and the secondary objective is the assessment of PK and preliminary anti-tumor activity. Results: A total of 29 patients were enrolled in this study. Tumor types were breast cancer, gastric/gastroesophageal junction cancer, colorectal cancer, gallbladder cancer, ovarian cancer and urothelial cancer. The medium prior lines of systemic therapy were 3 (range 1 – 6). During this study, no DLT was observed and the MTD was not reached. The most commonly reported (≥20%) treatment-related adverse events (TRAEs) were peripheral neuropathy, aspartate aminotransferase increased, neutrophil count decreased, white blood cell count decreased, alanine aminotransferase increased, hypertriglyceridemia, anemia, platelet count decreased. The most commonly reported ≥ grade 3 TRAEs (≥10 %) was peripheral neuropathy. There were two grade 4 adverse events: neurological symptom and sepsis. There was no interstitial lung disease observed during the study. Nine patients discontinued the study treatment due to peripheral neuropathy. Thirteen patients out of 26 patients evaluable for antitumor activity achieved partial response (PR), and 11 patients had stable diseases (SD). The best overall response rate (BOR) was 50.0% (13/26) and the disease control rate (DCR) was 92.3% (24/26). For 17 breast cancer patients in the study, 12 patients achieved PR, and 5 patients had SD. The BOR was 70.5% (12/17) and the DCR was 100% (17/17). For 5 gastric cancer patients in the study, all patients had SD. The duration of treatment ranged from 2 weeks to 66 weeks. Conclusions: BB-1701 was generally well tolerated up to 2.6 mg/kg dose levels and has shown promising antitumor activity in patients with HER2-overexpression solid tumors. Clinical trial information: NCT04257110 .

Recombinant influenza Virus-Like Particle (VLP) vaccines are promising vaccine candidates to prevent influenza, contain two major viral antigenic glycoproteins, Hemagglutinin (HA) and Neuraminidase (NA), on the surface of recombinant VLPs. Accurate quantitation of the mass of these antigenic proteins is important to ensure the product quality and proper dosing. Currently, Single Radial Immunodiffusion (SRID) is a recognized assay for determination of the HA immuno-reactive concentration (potency) in vaccine products, based on immuno-reactivity of HA with strain-specific antisera. The SRID assay, however, requires availability of strain-specific and properly calibrated reagents, which can be time-consuming to generate and calibrate. In addition, the assay is not suitable for quantitation of low abundant proteins, such as NA. In order to accelerate the overall production cycle, we have developed and optimized a high-resolution (HR) LC-MS method for absolute quantitation of both HA and NA protein concentrations in influenza VLP vaccine candidates. In this work, we present the method development, optimization and verification of its suitability for the intended purpose, as a prerequisite for its potential application in Quality Control, by assessing specificity, precision and accuracy, detection characteristics, and dynamic linear range. The method can be also used for other HA/NA containing preparations including in-process samples, purified proteins, whole virus preparations, nano-particle and egg-based vaccine preparations, or for calibration of SRID reference antigens.

Aspartate (Asp) isomerization is a common post-translational modification of recombinant therapeutic proteins that can occur during manufacturing, storage, or administration. Asp isomerization in the complementarity-determining regions of a monoclonal antibody may affect the target binding and thus a sufficiently robust quality control method for routine monitoring is desirable. In this work, we utilized a liquid chromatography-mass spectrometry (LC/MS)-based approach to identify the Asp isomerization in the complementarity-determining regions of a therapeutic monoclonal antibody. To quantitate the site-specific Asp isomerization of the monoclonal antibody, a UV detection-based quantitation assay utilizing the same LC platform was developed. The assay was qualified and implemented for routine monitoring of this product-specific modification. Compared with existing methods, this analytical paradigm is applicable to identify Asp isomerization (or other modifications) and subsequently develop a rapid, sufficiently robust quality control method for routine site-specific monitoring and quantitation to ensure product quality. This approach first identifies and locates a product-related impurity (a critical quality attribute) caused by isomerization, deamidation, oxidation, or other post-translational modifications, and then utilizes synthetic peptides and MS to assist the development of a LC-UV-based chromatographic method that separates and quantifies the product-related impurities by UV peaks. The established LC-UV method has acceptable peak specificity, precision, linearity, and accuracy; it can be validated and used in a good manufacturing practice environment for lot release and stability testing.LAY ABSTRACT:Aspartate isomerization is a common post-translational modification of recombinant proteins during manufacture process and storage. Isomerization in the complementarity-determining regions (CDRs) of a monoclonal antibody A (mAb-A) has been detected and has been shown to have impact on the binding affinity to the antigen. In this work, we utilized a mass spectrometry-based peptide mapping approach to detect and quantitate the Asp isomerization in the CDRs of mAb-A. To routinely monitor the CDR isomerization of mAb-A, a focused peptide mapping method utilizing reversed phase chromatographic separation and UV detection has been developed and qualified. This approach is generally applicable to monitor isomerization and other post-translational modifications of proteins in a specific and high-throughput mode to ensure product quality.

A new analytical method has been developed for the quantitative determination of ethylene glycol‐containing nonionic surfactants, such as polyethylene glycol 8000, polysorbate 80, and Pluronic F‐68. These surfactants are commonly used in pharmaceutical protein preparations, thus, testing in the presence of protein is required. This method is based on the capillary gas chromatographic analysis of ethylene glycol diacetate formed by hydrolysis and acetylation of surfactants that contain ethylene glycol. Protein samples containing free surfactants were hydrolyzed and acetylated with acetic anhydride in the presence of p ‐toluene sulfonic acid. Acetylated ethylene glycol was extracted with dichloromethane and analyzed by gas chromatography using a flame ionization detector. The amount of nonionic surfactant in the sample was determined by comparing the released ethylene glycol diacetate signal to that measured from calibration standards. The limits of quantitation of the method were 5.0 μg/mL for polyethylene glycol 8000 and Pluronic F‐68, and 50 μg/mL for polysorbate 80. This method can be applied to determine the polyethylene glycol content in PEGylated proteins or the final concentration of polysorbate 80 in a protein drug in a quality control environment.