Bifunctional Fusion Protein Research Articles

Abstract 443: Bintrafusp alfa enhances the antitumor efficacy of anti-VEGF and VEGF receptor kinase inhibitors in preclinical solid tumor models

Abstract The targeting of pathways that regulate tumor angiogenesis is an effective strategy for the treatment of solid tumors. Transforming growth factor (TGF)-β and vascular endothelial growth factor (VEGF) are master regulators of tumor angiogenesis that limit the efficacy of cancer therapies and suppress the anti-tumor immune response. Combination therapies consisting of immune checkpoint inhibitors and tumor angiogenesis inhibitors are emerging as effective treatments for solid cancers. Bintrafusp alfa (M7824) is a bifunctional fusion protein composed of the extracellular domain of the TGF-βRII receptor (a TGF-β “trap”) fused to a human IgG1 antibody blocking PD-L1. Here, we tested the antitumor efficacy of a combination therapy consisting of bintrafusp alfa plus a VEGF receptor kinase inhibitor (pazopanib, lenvatinib, regorafenib, axitinib, or nintedanib), or a VEGF neutralizing antibody (B20) in multiple preclinical tumor models. The combination of bintrafusp alfa with lenvatinib, regorafenib, axitinib, or nintedanib showed significantly greater antitumor efficacy than the respective monotherapies in the CT26 mouse syngeneic colon carcinoma model. In the 4T1 mouse breast carcinoma model, the combination of bintrafusp alfa with anti-VEGF, lenvatinib, or regorafenib was significantly more efficacious than the respective monotherapies. Additionally, the combination of bintrafusp alfa with pazopanib also showed superior antitumor efficacy in an orthotopic Renca mouse tumor model. These data indicate that bintrafusp alfa may be a viable combination partner with multiple angiogenesis inhibitors and provide a rational for the continued development of these combination therapies to treat solid cancers. Citation Format: Tsz-Lun Yeung, Adam Lazorchak, Huakui Yu, Jin Qi, Guozhong Qin, Bo Marelli, Kai Fu, Yan Lan. Bintrafusp alfa enhances the antitumor efficacy of anti-VEGF and VEGF receptor kinase inhibitors in preclinical solid tumor models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 443.

Abstract 693: ASKG843, a bifunctional fusion protein of PD1/TGF-βRII, demonstrates potent in vitro/in vivo activities, and excellent pharmacokinetic properties in non-human primates (NHPs) with good safety profiles

Abstract Introduction: Antibodies targeting the immune checkpoint PD1/PD-L1 are emerging as effective cancer immunotherapies, but most cancers fail to respond to these as single agents. Tumor cells and the tumor microenvironment (TME) frequently express transforming growth factor-β (TGF-β) to either drive immune cell dysfunction in the microenvironment or to exclude immune cell infiltration into tumors. Concurrently targeting the immunosuppressive/immune-excluding TGF-β pathway represents a rational and promising strategy to enhance PD1/PD-L1-based immunotherapies. ASKG843 is a bifunctional molecule designed to simultaneously block PD1/PD-L1 pathway and disable all three TGF-β isoforms in the TME. Here we report the functional evaluation of ASKG843 in vitro, anti-tumor activity in vivo, pharmacokinetic (PK) and pharmacodynamic (PD) properties in cynomolgus monkeys as well as its safety profiles following 5-repeat weekly dosing in NHPs. Methods: ASKG843 was constructed comprising αhPD1 Ab with the C-terminus of the heavy chain fused to the ECD of human TGF-βRII via flexible linkers. Bispecific binding properties were evaluated by Fortebio and bispecific ELISA. Respective signaling blockade for PD1 and TGF-β were assessed using either PD1/NFAT-luciferase reporter system or TGF-β/SMAD-luciferase reporter cell line. The anti-tumor activity of ASKG843 was determined in human PBMC-engrafted NSG mice-bearing A375 xenografts. The PK/PD and safety profiles of ASKG843 were assessed in NHPs. Results: Fortebio and bispecific ELISA analyses demonstrated the ability of ASKG843 to simultaneously bind both human PD1 and TGF-β1-3. ASKG843 not only dose-dependently blocked PD1/PD-L1 inhibitory axis with similar potency to parental αPD1 Ab in PD1 bioassay, but also thwarted TGF-β canonical signaling with IC50 =14.7 ng/mL in TGF-β bioassay. In human PBMC-reconstituted A375 xenograft model ASKG843 promoted strong anti-tumor response with efficacy comparable to the reference molecule (PD-L1/TGF-βRII) at the same dose/dosing frequency. In cynomolgus monkeys ASKG843 demonstrated superior PK to PD-L1/TGF-βRII reference molecule resulting in sustained depletion of TGF-β in peripheral blood for at least 10 days following single injection at 5 mg/kg dose, and no toxicity-related sign observed which contrasted with the reference molecule. Most encouragingly ASKG843 maintained very good safety profiles in cynomolgus monkeys following multiple injections at dose up to 100 mg/kg. Conclusions: ASKG843 demonstrated potent bi-functional activities in vitro and in vivo endowed by its design. Superior PK to the reference molecule yet maintaining better safety profiles in NHPs support further development of ASKG843 as an effective immunotherapeutic candidate. Citation Format: Liqin Liu, Shiwen Zhang, Kurt Shanebeck, Mouzhong Xu, Yunxia Yuan, Ming Li, Lu Li, Ray Chuang, Yong Wen, Yuefeng Lu, Jeff Lu. ASKG843, a bifunctional fusion protein of PD1/TGF-βRII, demonstrates potent in vitro/in vivo activities, and excellent pharmacokinetic properties in non-human primates (NHPs) with good safety profiles [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 693.

Functionalization of Cellulose-Based Hydrogels with Bi-Functional Fusion Proteins Containing Carbohydrate-Binding Modules.

Materials with novel and enhanced functionalities can be obtained by modifying cellulose with a range of biomolecules. This functionalization can deliver tailored cellulose-based materials with enhanced physical and chemical properties and control of biological interactions that match specific applications. One of the foundations for the success of such biomaterials is to efficiently control the capacity to combine relevant biomolecules into cellulose materials in such a way that the desired functionality is attained. In this context, our main goal was to develop bi-functional biomolecular constructs for the precise modification of cellulose hydrogels with bioactive molecules of interest. The main idea was to use biomolecular engineering techniques to generate and purify different recombinant fusions of carbohydrate binding modules (CBMs) with significant biological entities. Specifically, CBM-based fusions were designed to enable the bridging of proteins or oligonucleotides with cellulose hydrogels. The work focused on constructs that combine a family 3 CBM derived from the cellulosomal-scaffolding protein A from Clostridium thermocellum (CBM3) with the following: (i) an N-terminal green fluorescent protein (GFP) domain (GFP-CBM3); (ii) a double Z domain that recognizes IgG antibodies; and (iii) a C-terminal cysteine (CBM3C). The ability of the CBM fusions to bind and/or anchor their counterparts onto the surface of cellulose hydrogels was evaluated with pull-down assays. Capture of GFP-CBM3 by cellulose was first demonstrated qualitatively by fluorescence microscopy. The binding of the fusion proteins, the capture of antibodies (by ZZ-CBM3), and the grafting of an oligonucleotide (to CBM3C) were successfully demonstrated. The bioactive cellulose platform described here enables the precise anchoring of different biomolecules onto cellulose hydrogels and could contribute significatively to the development of advanced medical diagnostic sensors or specialized biomaterials, among others.

Targeting Tissue Factor to Tumor Vasculature to Induce Tumor Infarction.

Simple SummaryAmong multiple other functional roles of tissue factor (TF) and other coagulation proteins in the development and targeting of malignant disease, some scientific groups are attempting to modify TF and target the molecule or truncated forms of the molecule to tumor vasculature to selectively induce local blood vessel thromboembolic occlusion resulting in tumor infarction. This review briefly describes the characteristics and development of some of these proteins and structures, including tTF-NGR, which as the first drug candidate from this class has entered clinical trials in cancer patients.Besides its central functional role in coagulation, TF has been described as being operational in the development of malignancies and is currently being studied as a possible therapeutic tool against cancer. One of the avenues being explored is retargeting TF or its truncated extracellular part (tTF) to the tumor vasculature to induce tumor vessel occlusion and tumor infarction. To this end, multiple structures on tumor vascular wall cells have been studied at which tTF has been aimed via antibodies, derivatives, or as bifunctional fusion protein through targeting peptides. Among these targets were vascular adhesion molecules, oncofetal variants of fibronectin, prostate-specific membrane antigens, vascular endothelial growth factor receptors and co-receptors, integrins, fibroblast activation proteins, NG2 proteoglycan, microthrombus-associated fibrin-fibronectin, and aminopeptidase N. Targeting was also attempted toward cellular membranes within an acidic milieu or toward necrotic tumor areas. tTF-NGR, targeting tTF primarily at aminopeptidase N on angiogenic endothelial cells, was the first drug candidate from this emerging class of coaguligands translated to clinical studies in cancer patients. Upon completion of a phase I study, tTF-NGR entered randomized studies in oncology to test the therapeutic impact of this novel therapeutic modality.

MICA-G129R: A bifunctional fusion protein increases PRLR-positive breast cancer cell death in co-culture with natural killer cells.

Breast cancer cells were reported to up-regulate human prolactin receptor (PRLR) to assist their growth through the utilization of prolactin (PRL) as the growth factor, which makes PRLR a potential therapeutic target for breast cancer. On the other hand, advanced cancer cells tend to down-regulate or shed off stress signal proteins to evade immune surveillance and elimination. In this report, we created a fusion protein consisting of the extracellular domain of MHC class I chain-related protein (MICA), a stress signal protein and ligand of the activating receptor NKG2D of natural killer (NK) cells, and G129R, an antagonistic variant of PRL. We hypothesize that the MICA portion of the fusion protein binds to NKG2D to activate NK cells and the G129R portion binds to PRLR on breast cancer cells, so that the activated NK cells will kill the PRLR-positive breast cancer cells. We demonstrated that the MICA-G129R fusion protein not only binds to human natural killer NK-92 cells and PRLR-positive human breast cancer T-47D cells, but also promotes NK cells to release granzyme B and IFN-γ and enhances the cytotoxicity of NK cells specifically on PRLR-positive cells. The fusion protein, therefore, represents a new approach for the development of breast cancer specific immunotherapy.

SHR-1701, a bifunctional fusion protein targeting PD-L1 and TGF-β, for advanced NSCLC with EGFR mutations: Data from a multicenter phase 1 study.

9055 Background: Despite the development of targeted therapies for advanced NSCLC harboring EGFR mutations ( EGFR+), acquired resistance remains inevitable. Immune checkpoint inhibitor as monotherapy has limited efficacy. Blockade of the TGF-β pathway which plays a key role in immune suppression may enhance the tumor response to anti-PD-1/PD-L1 antibodies. Here, we assessed SHR-1701, a novel bifunctional fusion protein composed of a mAb against PD-L1 fused with the extracellular domain of TGF-β receptor II, in advanced NSCLC pts including one separate EGFR+ cohort. Methods: This phase 1 study includes a 3+3 dose-escalation and dose-expansion period of pretreated advanced NSCLC and multiple clinical expansion cohorts of different tumor types, genetic aberrations, or prior therapies. During the dose-escalation and dose-expansion period, pathologically confirmed pts received SHR-1701 at 3, 10, or 20 mg/kg Q3W or 20 mg/kg Q2W by intravenous infusion. The primary objectives were to determine the safety profile, maximum tolerated dose (MTD), and recommended phase 2 dose (RP2D) of SHR-1701. In the EGFR+ NSCLC clinical expansion cohort, histologically or cytologically confirmed advanced pts after at least 1L standard EGFR TKI received SHR-1701 at RP2D, and the primary endpoint was objective response rate (ORR). Treatment beyond progression was allowed. Results: During the dose-escalation and dose-expansion period, 30 pts were recruited: all stage IV; 83.3% had ≥2 metastasis sites; 76.7% had received ≥2L prior systemic therapy. One dose-limiting toxicity (immune-mediated pneumonitis) in the 20 mg/kg Q2W group was observed, and the MTD was not reached. Population pharmacokinetics and exposure‐response analysis of SHR-1701 based on this study and another phase 1 study for advanced solid tumors (NCT03710265) demonstrated 30 mg/kg Q3W as the RP2D. In the EGFR+ NSCLC cohort, 27 pts were enrolled: all stage IV; 77.8% had ≥2 metastasis sites; 70.4% had received ≥2L prior systemic therapy; 29.6% had 19-Del, 14.8% 19-Del and T790M, 7.4% 20-ins, 29.6% L858R, 18.5% L858R and T790M. With a median SHR-1701 exposure of 8.7 weeks (range, 3.0-24.0), 4 of the 24 pts who had at least one post-baseline radiographic assessment achieved objective responses, including 3 ongoing confirmed and 1 unconfirmed partial response. ORR was 16.7% (95% CI, 4.7%-37.4%), and disease control rate was 50.0% (95% CI, 29.1%-70.9%). Grade 3 treatment-related adverse events (TRAEs) occurred in 2 (7.4%) pts, including anemia, hypokalemia, and asthenia (1 [3.7%] for each). There were no grade 4 or 5 TRAEs. No pts discontinued treatment due to TRAEs. Conclusions: SHR-1701 monotherapy shows a manageable safety profile and encouraging antitumor activity in advanced EGFR+ NSCLC pts after failure of at least 1L standard EGFR TKI. Further investigation of SHR-1701 combination therapy for EGFR+ NSCLC is warranted. Clinical trial information: NCT03774979.

Evaluation of bintrafusp alfa, a bifunctional fusion protein targeting TGF-β and PD-L1, in cervical cancer: Data from phase 1 and phase 2 studies.

5509 Background: The accelerated FDA approval of pembrolizumab validated the efficacy of anti–PD-(L)1 therapy for pts with recurrent/metastatic cervical cancer; however, the objective response rate (ORR) with pembrolizumab was 14.3% in pts with PD-L1 expressing tumors. HPV infection is implicated in > 95% of cervical cancers and is linked to upregulation of TGF-β signaling. Bintrafusp alfa is a first-in-class bifunctional fusion protein composed of the extracellular domain of the TGF-βRII receptor (a TGF-β “trap”) fused to a human IgG1 mAb blocking PD-L1. We report pooled safety and efficacy in pts with immune checkpoint inhibitor–naive, recurrent/metastatic cervical cancer treated with bintrafusp alfa in phase 1 (INTR@PID 001; NCT02517398) and phase 2 (study 012; NCT03427411) studies. Methods: Pts with pretreated, immune checkpoint inhibitor–naive, recurrent/metastatic cervical cancer received bintrafusp alfa 0.3-30 mg/kg (phase 1 dose escalation) or 1200 mg Q2W (phase 1 expansion/phase 2) until progressive disease, unacceptable toxicity, or withdrawal. Treatment past progression was allowed. Primary endpoints were safety for the dose-escalation part of the phase 1 study and best overall response per RECIST 1.1 for the expansion part of phase 1 and phase 2 studies. Secondary endpoints for the expansion part of the phase 1 and 2 studies included safety. Results: As of May 15, 2020 (phase 1) and December 22, 2020 (phase 2), 39 pts had received bintrafusp alfa for a median duration of 2.8 months (range, 0.5-19.3). The median follow-up to data cutoff was 35.0 months and 24.1 months for the phase 1 and phase 2 studies, respectively. All pts had received prior anticancer therapy; 16 pts (41.0%) had received ≥3 prior anticancer regimens. There were 2 complete responses and 9 partial responses (PRs; ORR per RECIST 1.1, 28.2%). Median duration of response was 11.7 months (range, 1.4-41.2), and 5 pts (45.5%) had ongoing responses (duration 1.5-41.2 months). An additional delayed PR was observed (duration 23.7 months). Reponses occurred irrespective of tumor histology or prior bevacizumab or radiation treatment. Median overall survival (mOS) was 13.4 months (95% CI, 5.5 to not reached); 24-month OS rate was 33.2%. Any-grade treatment-related adverse events (TRAEs) occurred in 33 pts (84.6%). Grade 3 TRAEs occurred in 8 pts (20.5%; anemia, colitis, gastroparesis, upper gastrointestinal hemorrhage, keratoacanthoma, cystitis noninfective, hematuria, pneumonitis, rash macular [n = 1 each]); 1 patient (2.6%) had a grade 4 TRAE (asymptomatic hypokalemia related to the above grade 3 gastroparesis). No treatment-related deaths occurred. Conclusions: Bintrafusp alfa had a manageable safety profile and demonstrated clinical activity in pts with heavily pretreated, immune checkpoint inhibitor–naive recurrent/metastatic cervical cancer. Clinical trial information: NCT02517398 , NCT03427411.

Long-term follow-up of bintrafusp alfa, a bifunctional fusion protein targeting TGF-β and PD-L1, in advanced squamous cell carcinoma of the head and neck (SCCHN).

6020 Background: Bintrafusp alfa is a first-in-class bifunctional fusion protein composed of the extracellular domain of the TGF-βRII receptor (a TGF-β “trap”) fused to a human IgG1 mAb blocking PD-L1. A previous report of an expansion cohort from a phase 1 study (NCT02517398) suggested that bintrafusp alfa had a manageable safety profile and early signs of clinical activity in patients with heavily pretreated, advanced SCCHN after a median follow-up of 86.4 weeks. Here we report long-term efficacy and safety for this cohort. Methods: Patients with advanced SCCHN that progressed/recurred after platinum therapy in the recurrent/metastatic setting, or < 6 months after platinum therapy in the locally advanced setting, received bintrafusp alfa 1200 mg every 2 weeks until confirmed progressive disease, unacceptable toxicity, or trial withdrawal. The primary endpoint was confirmed best overall response assessed per RECIST 1.1 assessed by independent review committee (IRC); safety was a secondary endpoint. Results: As of May 15, 2020, 32 patients had received bintrafusp alfa for a median of 2.8 months (range, 0.5-29.9 months), no patient remained on treatment, and median follow-up to data cutoff was 41.7 months (range, 39.8-43.5 months). The objective response rate (ORR; 13%) was unchanged since the previous report; median duration of response (DOR) was increased at 21.4 months (95% CI, 5.5 months to not reached [NR]). While the clinical activity of bintrafusp alfa may be improved in patients with HPV-positive tumors (Table), outcomes were generally similar between PD-L1 subgroups (≥1% vs < 1% tumor cells). The overall safety profile was consistent with the previous report for this cohort, without grade 4 nor 5 treatment-related adverse events (TRAEs); no new TRAEs of grade 3 or that led to discontinuation of bintrafusp alfa were reported. Conclusions: With a median follow-up of over 3 years in patients with heavily pretreated advanced SCCHN, bintrafusp alfa showed sustained clinical activity and 3-year OS of 24.0%, which compares favorably to historical data. Clinical activity appeared to be greater in patients with HPV-positive tumors than those with HPV-negative tumors. The safety profile was manageable and consistent with earlier analysis. Further investigation of bintrafusp alfa in SCCHN and other HPV-associated cancers is ongoing. Clinical trial information: NCT02517398. [Table: see text]

A phase I/II study of bintrafusp alfa and NHS-IL12 in combination with docetaxel in adults with metastatic castration sensitive (mCSPC) and castration-resistant prostate cancer (mCRPC).

TPS5096 Background: Immune checkpoint inhibition is successful in a small subpopulation of men with prostate cancer. This could be related to barriers to immune response in the tumor microenvironment. Immunocytokines present an opportunity to specifically target the pleiotropic tumor microenvironment impacting immune cells beyond T-cells. NHS-IL12 is an immunocytokine that carries IL-12 (shown to impact natural killer and myeloid cells) and binds to necrotic tissue with exposed histones. Phase 1 studies have indicated immune and even PSA responses in prostate cancer to NHS-IL12 (Strauss J, CCR 2019). Preclinical data has demonstrated synergy with docetaxel, which is standard therapy in both mCSPC and mCRPC. Further synergy has been shown with a novel first-in-class bifunctional fusion protein (bintrafusp alfa) composed of the extracellular domain of human TGF-β receptor II (TGFβRII), which effectively functions to sequester or “trap” all three TGF-β isoforms (Lind H, JITC 2020), fused to a monoclonal antibody against PD-L1. This study will examine the potential of a novel combination of chemotherapy, checkpoint inhibition, and immunocytokines in metastatic prostate cancer. Methods: The study will evaluate safety of NHS-IL12 with docetaxel at escalating doses followed by the addition of bintrafusp alfa in all metastatic patients. Once safety of the combination is established, patients will enroll in 2 cohorts with either mCSPC or mCRPC. Eligible patients include mCSPC (≤134 days of starting ADT) and mCRPC (must have been previously treated with abiraterone or enzalutamide), with good performance status (ECOG of ≤ 2). Patients with brain metastases or who are immunocompromised are excluded. For mCSPC, the primary endpoint will evaluate the increase in the proportion of patients who have a PSA less than 0.2 ng/mL 7 months after the start of ADT, which is based on the prognostic value of PSA less than 0.2 ng/mL at 7 months in mCSPC (Harshman L, JCO 2017). The secondary endpoint is biochemical and radiographic time to progression. The primary endpoint for the mCRPC patients will evaluate progression free survival with secondary endpoints examining the percentage of patients with a 50% PSA decline from baseline and radiographic response rates per RECIST. Exploratory analysis will analyze changes in immune cell subsets after treatment as well as immune status of the tumor microenvironment. Clinical trial information: NCT04633252.

First-in-human phase I study of the bifunctional EGFR/TGFβ fusion protein BCA101 in patients with EGFR-driven advanced solid cancers.

3074 Background: Therapeutic targeting of EGFR has demonstrated efficacy in common advanced malignancies such as colorectal cancer (CRC), squamous cell of the head and neck (SCCHN), and non-small cell lung cancer (NSCLC). TGFβ has pro-tumorigenic effects on migration, invasion and tumor-specific immunosuppression, which may enhance the oncogenic effects of activated EGFR. Inhibition of EGFR signaling can lead to TGFβ upregulation as a resistance mechanism. BCA101 is a bifunctional recombinant fusion protein consisting of a chimeric anti-EGFR antibody and an extracellular domain (ECD) of human TGFβRII which demonstrated anti-tumor activity in several preclinical models. Methods: Patients with EGFR-driven advanced solid cancers refractory to standard therapies received BCA101 at escalating doses from 64 mg to 1000 mg intravenously (IV) weekly across 6 dose levels using a 3+3 design to determine dose limiting toxicities (DLT, established within 21 days of initial dosing), maximum tolerated dose (MTD) and/or recommended dose (RD). Secondary endpoints include detailed pharmacokinetic (PK), pharmacodynamic (PD) studies in serial tumor and/or blood samples and assessment of anti-tumor activity. Results: As of 2/11/2021, 21 patients received single agent BCA101 at 64 (n = 3), 240 (n = 7), 500 (n = 2), 750 (n = 3), 800 (n = 3) or 1000 (n = 3) mg IV weekly, including patients with CRC (n = 6), SCCHN (n = 5) uveal melanoma (n = 2), ovarian cancer (n = 2), glioblastoma multiforme (n = 2), conjunctival melanoma, chordoma, pancreatic cancer and anal squamous cell carcinoma (all n = 1). These patients had 1-7 prior lines of antineoplastic therapy (median 4), including 3 patients with prior EGFR inhibitor exposure. Adverse events (AE) related to BCA101 observed in > 1 patient included grade (G) 1-2 rash (n = 9), G 1-2 lipase elevation (n = 2). G3 vitreous hemorrhage at the 240 mg dose level has been the only DLT. The MTD has not been reached and the RD will be based on safety, exposure and pending PD data. Saturation of the clearance was observed at doses above 500mg. Dose proportional increase in Cmax and AUC were observed with doses of 750-1000mg. Best RECISTv1.1 response was stable disease (SD) in 3/10 evaluable patients, with 1 patient on drug ≥4 months. Conclusions: BCA101 is well tolerated at biologically active doses. BCA101 is now being tested in combination with the PD-1 antibody pembrolizumab in patients with SCCHN and anal carcinoma. Clinical trial information: NCT04429542.

A phase I study of bintrafusp alfa (M7824) and NHS-IL12 (M9241) alone and in combination with stereotactic body radiation therapy (SBRT) in adults with metastatic non-prostate genitourinary malignancies.

TPS4599 Background: The majority of non- prostate genitourinary (GU) cancers are lethal when metastatic and rare GU cancers have limited treatment options. Bintrafusp alfa is a bifunctional fusion protein composed of human TGF-β receptor II, which sequesters or “traps” all three TGF-β isoforms and a monoclonal PD-L1 antibody. NHS-IL12 is an immunocytokine composed of two IL-12 heterodimers, each fused to the H-chain of the NHS76 antibody. The NHS76 IgG1 antibody has affinity for both single- and double-stranded DNA (dsDNA) allowing for targeted delivery of pro-inflammatory cytokine, IL-12, to necrotic portions of tumor with DNA exposure to promote local immunomodulation. Preclinical data suggest synergy between these two agents. There is also evidence suggesting that stereotactic body radiation therapy (SBRT) can promote anti-tumor immune responses both locally and systemically while also synergizing with immune checkpoint inhibitors. Therefore, the combination of Bintrafusp alfa, NHS-IL12 and radiation is a potential strategy for metastatic non-prostate GU tumors. Methods: This is an open label, non-randomized, three-stage phase I trial of bintrafusp alfa and NHS-IL12 or bintrafusp alfa and NHS-IL12 in combination with either sequential or concurrent SBRT. Bintrafusp alfa (IV 1200 mg q2w) and SBRT (8 Gy x 3 fractions) are planned with a deescalating NHS-IL12 (subQ q4w) dose schedule. The accrual ceiling has been set at 66 patients. The trial will enroll patients with a pathologically confirmed diagnosis of metastatic non-prostate genitourinary cancer with an ECOG ≤ 2 (KPS ≥60%). Participants may have had prior cancer immunotherapy but excluding prior treatment with bintrafusp alfa and/or NHS-IL12. 9 patients will receive treatment in cycles consisting of 4 weeks. The primary objective is to determine the safety and highest tolerated doses with acceptable toxicity (recommended phase II dose) of bintrafusp alfa and NHS-IL12 alone or in combination with SBRT administered sequentially or concurrently in patients with metastatic non-prostate genitourinary cancers. Secondary objectives are objective response rate (ORR), progression free survival (PFS) and overall survival (OS). Exploratory objectives are to determine peripheral immune modulation and the status of the immune microenvironment using cytokine analysis, circulating tumor cells, multiplex immunohistochemistry, T-cell receptor sequencing, and RNA-sequencing. The study is open and enrolling. Clinical trial information: NCT04235777.

TGF-β and PD-L1 inhibition combined with definitive chemoradiotherapy in esophageal squamous cell carcinoma: A phase II clinical trial (NCT04595149).

TPS4154 Background: Esophageal cancer is the 6th leading cause of mortality worldwide, with an overall 5-year survival rate of 10%. This is in part due to more than 50% of patients presenting with irresectable or metastatic disease. With the introduction of definitive chemoradiation, 3-year survival rates of patients with irresectable tumors have risen to up to 40% (Hulshof et al., ASCO GI Oral Presentation, 2020). However, treatment still fails in the majority of patients due to locoregional recurrences or development of metastatic disease. Recently, it has been shown that addition of TGF-β inhibition to chemoradiation may improve treatment efficacy (Steins et al., Int J Cancer, 2019). Additionally, PD-L1 inhibition has emerged as a relevant therapeutic strategy in specific patient subgroups, such as squamous cell carcinoma (Kato et al., The Lancet Oncology, 2019). In this phase II study, we will investigate the feasibility of addition of bintrafusp alfa, a bifunctional fusion protein blocking TGF-β and PD-L1, to definitive chemoradiation in patients with esophageal squamous cell carcinoma. Methods: To assess feasibility, 52 patients will receive definitive chemoradiation combined with three doses of bintrafusp alfa at week 1, 4 and 7 (Table). Feasibility is defined as ≥80% of patients completing 2 cycles of bintrafusp alfa and will be tested with a one-sample test for a binomial proportion, comparing the observed percentage to the fixed reference value of unfeasibility (62%). Secondary endpoints are toxicity, progression-free survival, overall survival and quality of life. Additionally, exploratory endpoints include development of biomarkers to predict treatment response. Eligible for inclusion are patients with surgically irresectable (T1-4a N+ M0) squamous cell carcinoma of the esophagus or gastro-esophageal junction, or patients with resectable tumors refraining from radical surgery. Patients with M1 disease solely on the basis of supraclavicular metastasis are eligible. Patients with locoregional recurrences are eligible, provided that full dose of radiation can be safely delivered. Currently, 2 of planned 52 patients have been enrolled. Clinical trial information: NCT04595149. [Table: see text]

First-in-human phase I/II trial of PRGN-2009 vaccine as monotherapy or with bintrafusp alfa in patients with recurrent/metastatic (R/M) human papillomavirus (HPV)-associated cancers (HPVC) and as neoadjuvant/induction therapy in locoregionally advanced (LA) HPV oropharyngeal (OP) and sinonasal (SN) squamous cell cancer (SCC).

TPS6092 Background: R/M HPVC (cervical, anal, oropharyngeal, etc.) are incurable by current therapies. For newly diagnosed LA HPV-OPSCC standard-of-care (SOC) is radiotherapy ± chemotherapy (C/RT) or surgery ± adjuvant C/RT, with considerable risk of relapse. Newly diagnosed LA SNSCC treatment follows the OPSCC paradigm, and detection of HPV appears to confer improved prognosis. Neoadjuvant PD-1 immune checkpoint blockade (ICB) before surgery may improve RFS and is being evaluated in a multicenter phase III clinical trial (Keynote-689). PRGN-2009 (P) is a novel gorilla adenovirus vaccine containing 35 non-HLA-restricted epitopes of HPV 16 and 18 shown to induce HPV specific responses (preclinical models). Bintrafusp alfa (BA) is a bifunctional fusion protein targeting TGF-β and PD-L1 with promising activity in HPVC. This trial will evaluate the safety and activity of P/ P + BA in patients with previously treated R/M HPVC and as neoadjuvant/induction therapy before SOC surgery or C/RT in newly diagnosed LA HPV-OPSCC and HPV-SNSCC. Methods: This is a first-in-human, investigator-initiated, single-center phase I/II trial. Pts with previously treated (incl. ICB) R/M HPVC are eligible for Phase I: P dose escalation arm (3+3 design, 6-12 patients) testing 2 dose levels (1x1011, 5x1011 viral particle units, SC Q2W three times, then Q4W), and combination arm (10 patients) testing P (recommended phase 2 dose (RP2D), same schedule) + BA (1200 mg IV Q2W). Treatment (both arms) will continue until disease progression, unacceptable toxicity, decision to withdraw. Primary endpoint is safety. Secondary endpoints include ORR (RECIST 1.1), PFS, and OS. For Phase II, patients with newly diagnosed stage II/III (AJCC Cancer Staging Manual, 8th ed.) HPV-OPSCC and stage II/III/IVA/IVB HPV-SNSCC planned for SOC C/RT or surgery will be eligible for two treatment arms of 20+2 patients each (sequential): P arm and P + BA, to evaluate the treatment activity. All patients will have pre-treatment biopsy, receive two cycles of the study treatment at the NCI Clinical Center two weeks apart, followed by post-treatment biopsy and SOC treatment (at the referring institution) 4 weeks after the first study treatment. Primary endpoint is post-treatment ≥2-fold increase in tumor-infiltrating CD3+ cells. Secondary endpoints include RFS, OS. Exploratory endpoints for both arms include analyses of immune subsets, soluble factors, and HPV-specific immune responses in peripheral blood and tissue where available, and in Phase II sequencing (exome, scRNA), immune spatial profiling with multiplex immunofluorescence, and salivary HPV DNA. Clinical trial registry: NCT04432597. Clinical trial information: NCT04432597.

Phase II evaluation of the triple combination of PDS0101, M9241, and bintrafusp alfa in patients with HPV 16 positive malignancies.

2501 Background: There are more than 630,000 cases of HPV associated malignancies including cervical, oropharyngeal and anal cancer worldwide annually. HPV 16 is responsible for the majority of these cases. About 15-20% of HPV associated malignancies respond to PD-(L)1 inhibitors, but for the overwhelming majority of patients who progress on these immunotherapies there is no effective standard of care therapy. Preclinical studies have shown that the triple combination of PDS0101 (Versamune-HPV), a liposomal multipeptide therapeutic vaccine targeting HPV 16 E6/E7, M9241, a tumor-targeting immunocytokine composed of IL-12 heterodimers fused to a monoclonal antibody targeting free DNA in necrotic tumor regions, and bintrafusp alfa, a bifunctional fusion protein targeting TGF-β and PD-L1, resulted in maximum HPV-specific T cell responses, T cell tumor infiltration and tumor reduction as compared to any one or two of these agents alone. Methods: Fourteen pts with HPV 16+ relapsed or refractory advanced cancer were enrolled to the triple combination of PDS0101, M9241 and bintrafusp alfa (NCT04287868). Pts received bintrafusp alfa at 1200 mg flat dose i.v. every 2 weeks, M9241 at 16.8 mcg/kg s.c. every 4 weeks and PDS0101 given as two separate 0.5 ml s.c. injections every 4 weeks. Dose reductions of M9241 to 8 mcg/kg were allowed as well as skipped doses of any agent for ongoing toxicities. Results: Fourteen pts with advanced HPV 16+ cancers (5 cervical, 2 vaginal/vulvar, 4 anal, 3 oropharyngeal) were treated. 4/14 (28.6%) pts had a grade 3 treatment related toxicity including grade 3 hematuria in 2 pts with cervical ca and prior pelvic radiation and grade 3 AST/ALT elevation in 2 pts, one with anal ca and one with vaginal ca. For one patient with grade 3 AST/ALT elevation dose reduction of M9241 from 16.8 to 8 mcg/kg allowed for continued treatment with AST/ALT remaining at grade 1 or less. One additional patient had transient asymptomatic grade 4 neutropenia. No other treatment related grade 3 or greater toxicities were noted. 10/14 (71%) pts have had objective responses: 1 CR (anal ca) and 9 PRs (3 cervical, 2 vulvar/vaginal, 2 anal, 2 oropharyngeal) with 9/10 of these responses ongoing after a median 5 month of follow up. Of the 14 pts, 6 pts have checkpoint naïve disease and 8 pts have checkpoint refractory disease. 5/6 (83%) pts with checkpoint naïve disease and 5/8 (63%) pts with checkpoint refractory disease have had objective responses. Analyses of immune responses and other immune correlates are ongoing. Conclusions: Triple combination of PDS0101, M9241 and bintrafusp alfa appears to have a manageable safety profile along with early evidence of notable clinical activity for pts with both checkpoint naïve as well as checkpoint refractory HPV 16+ advanced malignancies. Clinical trial information: NCT04287868.

Phase 1 study of SHR-1701, a bifunctional fusion protein targeting PD-L1 and TGF-β, in patients with advanced solid tumors.

2503 Background: Dual inhibition of PD-1/PD-L1 and TGF-β pathways is a promising therapeutic strategy for multiple tumor types. SHR-1701 is a novel bifunctional anti-PD-L1/TGF-βRII agent. This dose escalation and expansion phase 1 study aimed to evaluated the safety and preliminary anti-tumor acitivity of SHR-1701 in refractory solid tumors. Methods: The dose escalation period was initiated by accelerated titration (1 mg/kg Q3W) and then switched to 3+3 scheme (3, 10, 20, and 30 mg/kg Q3W and 30 mg/kg Q2W). The dose expanded at doses of 10, 20, and 30 mg/kg Q3W and 30 mg/kg Q2W. The primary objectives were to determine the safety profile, MTD, and RP2D of SHR-1701. Results: 17 pts (1 mg/kg Q3W [n = 1]; 3, 10, 20 and 30 mg/kg Q3W [n = 3 each]; 30 mg/kg Q2W [n = 4]) were enrolled in dose escalation part. No DLT was observed and MTD was not reached. Another 32 pts (10 mg/kg Q3W [n = 8]; 20 and 30 mg/kg Q3W [n = 9 each]; 30 mg/kg Q2W [n = 6]) were enrolled in dose expansion part. Of 49 enrolled pts, 33 pts (67.3%) had received ≥2 lines of prior systemic therapy. As of data cutoff on Oct 30, 2020, the median duration of SHR-1701 exposure was 6.0 weeks (range, 2.0-78.6). The most common reported TRAEs were increased ALT/AST, anemia, hypothyroidism, and increased bilirubin/conjugated bilirubin, with incidence > 15%. The incidence of irAEs reported by the investigator was 46.9% and 4 pts received systemic corticosteroids. Hypothyroidism and rash were the most common irAEs with incidence > 10%. The incidence of Grade ≥3 TRAEs was 18.4%. The incidence of Grade ≥3 irAEs was 10.2%. 1 pt suffered early death for liver failure more likely caused by tumor progression. PK analysis showed a linear dose-exposure relationship with SHR-1701 dosing from 1 to 30 mg/kg. The peripheral PD-L1 target occupancy rate exceeded 90%, and nearly complete TGF-β1 trapping was detected in all dose groups. Of 49 enrolled pts, 45 pts completed at least once efficacy evaluation. The ORR was 17.8% (95% CI, 8.0%-32.1%), with 8 pts achieving PR (2 lung adenocarcinoma, 1 HCC, 1 ESCC, 1 dMMR-CRC, 1 renal cancer, 1 epiglottis cancer, and 1 pancreatic acinar cell carcinoma). The DCR was 40.0% (18/45; 95% CI, 25.7%-55.7%). Majority of responses (7/8) were still ongoing, and the median DoR had not been reached yet. Based on data of saftety, PK, PD, and efficacy, we recommended 30 mg/kg Q3W as the RP2D. Conclusions: SHR-1701 showed acceptable safety profile and encouraging antitumor activity in refractory solid tumors, establishing the foundation for further exploration. Clinical trial information: NCT03710265.

Novel GLP-1/anti-apolipoprotein B bifunctional fusion protein alleviates diabetes and diabetic complications in combination with low-intensity ultrasound

AimsTo engineer and screen a novel GLP-1/anti-apolipoprotein B (apoB) bifunctional fusion protein with therapeutic potential on alleviating diabetes and diabetic complication in combination with low-intensity ultrasound. Main methodsAnti-apoB antibodies were screened by phage display technology and further fused to mutated GLP-1 (7–37) via light or heavy fusion to generate bifunctional fusion protein (termed aBG). The optimal design of aBG fusion protein was further confirmed by in vitro epitope competition assay and cAMP accumulation assay. Subsequently, chronic study in DIO mice were subjected to assess the long-term efficacy of screened fusion protein. Key findingsThe selected GLP-1/anti-apoB fusion protein, aBG-8, exerted either the highest binding affinities for GLP-1R and apoB, or the greatest LDL-C uptake capacity and GLP-1R activation activity. After 60-day treatment in DIO mice, aBG-8 was proved to exert the promising improvement on hyperglycemia, hyperlipidemia, and obesity in DIO mice. Furthermore, combined therapy of aBG-8 and low-intensity ultrasound could accelerate skin wound closure in diabetic mice. SignificanceA novel long-lasting bifunctional fusion molecule, aBG-8, was designed with the enormous potential on alleviating diabetes and diabetic complications in combination with low-intensity ultrasound.

Abstract PO-088: Bintrafusp alfa enhances anti-tumor efficacy in combination with radiation therapy plus the ataxia telangiectasia mutated (ATM) inhibitor, M4076

Abstract Ionizing radiation therapy (RT) is a standard of care for many cancer indications. RT promotes the activation of transforming growth factor (TGF)-β, a pleotropic cytokine that mediates RT-induced fibrosis and suppresses anti-tumor immunity. Additionally, RT induces the expression of the immunosuppressive checkpoint PD-L1 on tumor cells. Bintrafusp alfa (M7824) is a first-in-class bifunctional fusion protein composed of the extracellular domain of the TGF-βRII receptor, functioning as a TGF-β “trap”, fused to a human IgG1 antibody blocking PD-L1. Bintrafusp alfa enhances anti-tumor immunity elicited by RT and reverses RT-induced fibrosis in preclinical models. Inhibition of ataxia telangiectasia mutated (ATM) increases the cancer cell's sensitivity to radiotherapy by blocking cellular pathways which facilitate the response to, and repair of, RT induced DNA double strand breaks. We hypothesize that a combination therapy consisting of bintrafusp alfa, RT and the ATM kinase inhibitor M4076 will show greater anti-tumor efficacy than the respective mono- or dual-combination therapies in immune competent preclinical solid tumor models. The anti-tumor efficacy of bintrafusp alfa, RT, and M4076 triple combination therapy was explored in the immune therapy resistant 4T1 murine syngeneic breast carcinoma model. This triple combination therapy significantly increased anti-tumor efficacy and prolonged overall survival relative to the bintrafusp alfa + RT or M4076 + RT dual combination therapies. The triple combination was further evaluated in the more immunogenic MC38 colon carcinoma model. MC38 tumor bearing mice treated with the triple combination therapy showed a significantly greater tumor growth inhibition and long term tumor free survival when compared with the bintrafusp alfa + RT or M4076 + RT treatment groups. Finally, the bintrafusp alfa + RT + M4076 combination therapy also showed significantly greater tumor growth inhibition and long term tumor free survival when compared to the combination of anti-PD-L1 + RT + M4076. The bintrafusp alfa triple combination resulted in 8 of 10 mice showing long term tumor free survival, whereas the anti-PD-L1 triple combination failed to elicit tumor free survival (0 of 10 mice tumor free). These studies demonstrate the superior anti-tumor efficacy of the Bintrafusp alfa + RT + M4076 triple combination therapy over that of the dual combination therapies of RT + bintrafusp alfa or RT + M4076. These data in the MC38 model also suggest that bintrafusp alfa may promote a superior anti-tumor immune response relative to that of anti-PD-L1 when in combination with RT and M4076. Together, these data support the rational to develop a bintrafusp alfa + RT + M4076 combination therapy for the treatment of solid tumors. Citation Format: Adam Lazorchak, Huakui Yu, Guozhong Qin, Jin Qi, Bo Marelli, Yan Lan. Bintrafusp alfa enhances anti-tumor efficacy in combination with radiation therapy plus the ataxia telangiectasia mutated (ATM) inhibitor, M4076 [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-088.

Study on the activity of recombinant mutant tissue-type plasminogen activator fused with the C-terminal fragment of hirudin.

In the present study, bifunctional fusion proteins were designed by fusing the kringle 2 and protease domains of tissue-type plasminogen activator (tPA) to the C-terminal fragment of hirudin. The thrombolytic and anticoagulant activities of these recombinant proteins from mammalian cells were investigated using in vitro coagulation models and chromogenic assays. The results showed that all assayed tPA mutants retained catalytic activity. The C-terminal fragment of hirudin may have weak affinity to thrombin and thus was insufficient to suppress thrombin-mediated fibrin agglutination. The strength of the thrombolytic activity only relied on the selected tPA sequences, and the fibrinolytic efficiency of single-chain protein significantly decreased. Our data indicate that truncated tPA combined with a hirudin peptide may provide a framework for the further development of a new antithrombotic agent.

A phase Ib single-arm study of bintrafusp alfa for the treatment of pretreated, locally advanced/unresectable or metastatic urothelial cancer.

TPS501 Background: Urothelial cancer (UC) is the most common histological subtype of bladder cancer. For patients with metastatic UC, platinum-containing chemotherapy (CT) is the first-line standard of care. With 5 PD-(L)1 inhibitors approved for second-line treatment of platinum-refractory advanced UC, objective response rates (ORRs) in this setting (15% to 21%) and in first-line cisplatin-ineligible patients (23% to 31%) suggest that there remains a significant unmet need for improved outcomes. TGF-β signaling has been associated with resistance to PD-(L)1 inhibitors in patients with UC. Bintrafusp alfa (BA) is a first-in-class bifunctional fusion protein composed of the extracellular domain of the TGF-βRII receptor (a TGF-β “trap”) fused to a human IgG1 mAb blocking PD-L1. In murine models, BA resulted in improved antitumor activity vs TGF-β or PD-L1 monotherapies alone or in combination. In 2 phase I trials (NCT02517398 and NCT02699515), BA demonstrated a manageable safety profile and encouraging clinical efficacy in >670 patients with advanced solid tumors. Here we describe a phase Ib study (NCT04349280) that will assess the clinical activity and safety profile of BA in platinum-experienced patients with locally advanced/unresectable or metastatic UC. Colocalized, simultaneous inhibition of the TGF-β and PD-L1 pathways by BA is hypothesized to elicit greater antitumor activity than anti–PD-(L)1 therapies in patients with UC. Methods: This open-label, multicenter, single-arm trial is accruing patients with histologically confirmed locally advanced/unresectable or metastatic UC (ECOG PS ≤1) with disease progression or recurrence after platinum-based CT. Patients must not have received >2 lines of systemic therapy for metastatic disease or prior therapy targeting T-cell costimulation, or checkpoint or TGF-β pathways. Patients with pneumonitis or a history of noninfectious pneumonitis that required systemic immunosuppression are ineligible. Patients will receive BA 1200 mg every 2 weeks until progression, unacceptable toxicity, death, or study withdrawal, or for up to 2 years. The primary endpoint is investigator-assessed confirmed ORR per RECIST 1.1; key secondary endpoints include duration of response and progression-free survival per the investigator and IRC, overall survival, safety, immunogenicity, and pharmacokinetics. Exploratory biomarker analyses will be conducted using patient samples collected during screening and prior to administration of BA. The base ORR used for the null hypothesis is 21%, and the target ORR is 40%. Using a predictive probability design and an estimated enrollment of 40 patients, the type I error rate is 5.70% and the power is 85.39%. As of September 2020, 3 sites in 2 countries have been activated and no patients have been enrolled. Clinical trial information: NCT04349280.

Abstract OT-32-01: A phase 2, open-label study of bintrafusp alfa monotherapy in patients with HMGA2-expressing triple-negative breast cancer

Abstract Background: Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with few targeted treatment options and a poor prognosis. Despite approvals of the anti-PD-L1 monoclonal antibody (mAb) atezolizumab in combination with nab-paclitaxel for unresectable, locally advanced/metastatic TNBC that expresses PD-L1 (tumor-infiltrating immune cells ≥1% of tumor area), many recent studies of other anti-PD-(L)1 therapies in advanced TNBC have shown limited efficacy, likely due to intrinsic therapeutic resistance. Transforming growth factor β (TGF-β), which promotes cancer progression by inducing angiogenesis, fibrosis, and epithelial-mesenchymal transition (EMT), may attenuate the efficacy of or promote resistance to anti-PD-(L)1 therapies. Bintrafusp alfa is a first-in-class bifunctional fusion protein composed of the extracellular domain of the TGF-βRII receptor (a TGF-β “trap”) fused to a human IgG1 mAb blocking PD-L1. In a cohort of 33 patients with heavily pretreated, advanced TNBC that progressed during/after first-line therapy, bintrafusp alfa was safe and resulted in antitumor activity (NCT02517398). Exploratory biomarker analysis showed that high mobility group AT-hook 2 (HMGA2) expression was 32-fold higher in tumor samples from patients who experienced disease control than from patients who had progressive disease in that cohort. Elevated expression of HMGA2, a protein associated with TGF-β signaling and a known regulator of EMT, is associated with metastasis and poor survival in breast cancer. We present the study design of a phase 2 trial to evaluate the efficacy and safety of bintrafusp alfa in patients with pretreated metastatic TNBC that expresses high levels of HMGA2. Trial Design: This phase 2, multicenter, open-label study will evaluate bintrafusp alfa monotherapy in patients with HMGA2-expressing TNBC that progressed on ≥1 line of systemic therapy for their metastatic disease. Patients will receive bintrafusp alfa 1200 mg every 2 weeks until confirmed progression, unacceptable toxicity, or trial withdrawal. Eligibility Criteria: Patients must have histologically confirmed TNBC defined by ASCO-CAP guidelines (estrogen receptor: immunohistochemistry [IHC] <1%; progesterone receptor: IHC <1%; human epidermal growth factor receptor 2: in situ hybridization nonamplified or IHC 0/1), high tumor HMGA2 expression, ECOG performance status ≤1, and measurable disease by RECIST 1.1. Patients must have experienced disease progression while receiving the most recent therapy prior to enrollment. HMGA2 expression will be centrally determined on archival or fresh tumor tissue by RT-PCR. Prescreening for HMGA2 expression while receiving preceding treatment is allowed; a fresh tumor biopsy prior to study entry may be requested for exploratory biomarker analysis. Patients with prior exposure to immunotherapy are not eligible. Specific Aims: The primary endpoint is independent review committee-assessed objective response rate per RECIST 1.1. Key secondary endpoints include safety, duration of response, durable response rate, progression-free survival, and overall survival. Additional exploratory biomarker characteristics will also be investigated. Statistical Methods: Descriptive statistics, including mean, median, standard deviation, and range, will be used to characterize continuous variables. Frequency counts and percentages will be used to characterize categorical variables. Accrual: Planned enrollment is 29 patients. Contact Information: Leisha A. Emens, MD, PhD Email: emensla@upmc.edu Citation Format: Leisha A Emens, Margaret E. Gatti-Mays, Joyce O’Shaughnessy, Luc Dirix, Giovanni Faggioni, Andrea Fontana, Jerome Martin-Babau, Christoph Helwig, Alice Huang, Riham Iadevaia, Laureen S Ojalvo. A phase 2, open-label study of bintrafusp alfa monotherapy in patients with HMGA2-expressing triple-negative breast cancer [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr OT-32-01.