CEACAM1 Research Articles

Research Highlights.

Research Highlights.

Abstract CT213: Biomarker analysis from Phase 1/2 study of tusamitamab ravtansine (SAR408701) in patients with advanced non-small cell lung cancer (NSCLC)

Abstract Background Tusamitamab ravtansine is an antibody-drug conjugate of a humanized carcinoembryonic antigen (CEA)-related cell adhesion molecule 5 (CEACAM5)-specific monoclonal antibody linked to DM4. A Phase 1/2 study (NCT02187848) showed tusamitamab ravtansine antitumor activity in pretreated patients (pts) with advanced nonsquamous NSCLC and high CEACAM5 expression. Here, we explore biomarker associations with tumor CEACAM5 expression by immunohistochemistry (IHC), and whether biomarkers predict objective response rate (ORR). Methods We assessed CEACAM5 expression by IHC, RNA sequencing, and whole exome sequencing (WES) on latest archival tumor samples; and circulating CEACAM5 (cCEACAM5) and CEA (cCEA). We enrolled 2 cohorts of pts with IHC CEACAM5 membrane expression at ≥2+ intensity: in ≥50% of tumor cells (high expressors, HEs, n = 64); and in ≥1% to <50% of tumor cells (moderate expressors, MEs, n = 28). Pts received tusamitamab ravtansine 100 mg/m2 IV every 2 weeks. Results cCEA and cCEACAM5 were strongly associated (Spearman rho, 0.9), with weak associations between IHC CEACAM5 and cCEA or cCEACAM5 (Spearman rho, 0.3 and 0.4, respectively). Higher levels of CEACAM5 mRNA were observed in CEACAM5 HEs vs MEs (P=0.0027). EGFR and KRAS genetic alterations by WES were present in 44.8% and 65.5% of CEACAM5 HEs, respectively, and 21.4% and 78.6% of CEACAM5 MEs, respectively. Confirmed partial responses were seen in 13/64 HEs (ORR 20.3%) and 2/28 MEs (ORR 7.1%). In CEACAM5 HEs with available baseline (BL) cCEA data, 25/62 (40.3%) had a cCEA level ≥100 µg/L, with a median value of 71.6 µg/L (range 1-8809); corresponding values in CEACAM5 MEs were 7/28 (25.0%) and 12.4 µg/L (range 0.5-684). In response evaluable CEACAM5 HEs with available BL cCEA data (n = 61), ORR was 10/24 (41.7%) in pts with high cCEA (≥100 µg/L) and 3/37 (8.1%) in pts with low cCEA (<100 µg/L); corresponding ORRs in CEACAM5 MEs were 0/7 and 2/21 (9.5%). Conclusions In CEACAM5 HEs, high cCEA was associated with numerically greater ORR vs low cCEA (41.7% vs 8.1%). Associations were also observed between: cCEA and cCEACAM5; IHC CEACAM5, cCEA, and cCEACAM5; and IHC CEACAM5 and CEACAM5 tumor mRNA levels, but not between IHC CEACAM5 and actionable oncogenic drivers. Clinical Trials Registration: ClinicalTrials.gov NCT02187848 Citation Format: Anas Gazzah, Joon Sang Lee, Emma Wang, Nils Ternès, Hong Wang, Eric Boitier, Aude Lartigau, Mustapha Chadjaa, Colette Dib, Gaëlle Muzard, Sandrine Valence, Anne Remaury, Cintia C. Palu, Anne-Laure Bauchet. Biomarker analysis from Phase 1/2 study of tusamitamab ravtansine (SAR408701) in patients with advanced non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT213.

Investigation of the Role of Carcinoembryonic Antigen-Related Cell Adhesion Molecule-1 in Diabetic Retinopathy

ABSTRACT Purpose Diabetic retinopathy (DR) has become a major cause of blindness with increased prevalence of diabetic mellitus. Carcinoembryonic antigen-related cell adhesion molecule-1 (CEACAM1) plays a part in pathological neovascularization. This study aimed to investigate the role of CEACAM1 in the progression of DR. Methods Aqueous and vitreous samples were collected from proliferative or non-proliferative DR and the control group. Multiplex fluorescent bead-based immunoassays were used to detect the levels of Cytokines. Expression of CEACAM1, VEGF, VEGF receptor 2 (VEGFR2) and hypoxia-induced factor-1α (HIF-1α) were detected in human retinal microvascular endothelial cells (HRECs). Results CEACAM1 and VEGF levels were significantly upregulated in PDR group and positively correlated with PDR progression. Expression CEACAM1 and VEGFR2 were increased in HRECs under hypoxic conditions. The HIF-1α/VEGFA/VEGFR2 pathway was blocked by CEACAM1 siRNA in vitro. Conclusions CEACAM1 might play a role in the pathology of PDR. CEACAM1 might be a therapeutic target for retinal neovasculariztion.

Abstract 2763: Characterization of a novel immunohistochemistry (IHC) assay for CEACAM5 using a commercial antibody

Abstract Introduction: Carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM5) is expressed on the surface of some tumor epithelial cells and is a potential therapeutic target for antibody-drug conjugates such as tusamitamab ravtansine in development. An IHC assay has been fit-for-purpose validated in non-small cell lung cancer (NSCLC) for evaluation of clinical samples using a proprietary CEACAM5-specific murine antibody (Sanofi clone 769) and the Dako/Agilent Autostainer Link 48 platform. Here we compare 5 commercial antibody clones to the validated IHC assay for CEACAM5. Methods: We tested 5 commercial clones (CI-P83-1 [Santa Cruz Biotechnology], 487609 [R&D Systems], OTI1D4 [Thermo Fisher], EPCEAR7 [Abcam], and 327 [Sino Biological]) to evaluate CEACAM5 detection in formalin-fixed, paraffin-embedded (FFPE) human cancer tissues from Discovery Life Sciences using the Leica BOND III platform. The staining pattern and intensity of CEACAM5 reactivity were compared for clone 769 and the commercial clones in the same FFPE samples. Initial IHC testing used gastric cancer tissues with continued assay development in colorectal cancer tissues. Commercially available (HT-29, MKN-45, BxPC-3, and PC-3) cell lines and CHO cells overexpressing other CEACAM targets (Sanofi) were used to test for cross reactivity and specificity. IHC staining in cell line controls and human cancer tissues (small cell lung cancer, NSCLC adenocarcinoma, squamous cell NSCLC, and pancreatic cancer), was performed to further evaluate staining accuracy to CEACAM5 and to better compare clones. Results: Of the 5 antibodies tested, 3 were deemed unsatisfactory due to diffuse cytoplasmic staining or overstaining and were not evaluated further. Clones CI-P83-1 and 487609 best matched the pattern and intensity of clone 769 staining and were evaluated at various dilutions in gastric and colorectal cancer samples to determine optimal antibody concentration and to demonstrate range and linearity of the IHC assays. Clones CI-P83-1 and 487609 accurately and specifically detected CEACAM5 without cross reactivity to similar targets in cell lines. In the human cancer tissues, clone CI-P83-1 tended to show greater parity to clone 769 than clone 487609, which sometimes showed slightly higher CEACAM5 signal than clone 769. Clone CI-P83-1 was appropriately reactive in CEACAM5 expression control cells, highly reactive in CHO cells overexpressing CEACAM5, and lacked cross reactivity in CHO cells overexpressing CEACAM types 1, 6, 7, or 8. A fit-for-purpose validation of clone CI-P83-1 in non-squamous NSCLC with clone 769 showed comparable staining in this indication. Conclusions: Among the 5 tested clones, there were differences in staining pattern, intensity, and cross reactivity. CI-P83-1 was the closest to 769 among the 5 clones. Further testing and validation are required to understand the finer differences between 769 and CI-P83-1. Citation Format: Chin Leng Cheng, Hongli Chen, Karen Kirchner, Gregory Cesarone. Characterization of a novel immunohistochemistry (IHC) assay for CEACAM5 using a commercial antibody [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 2763.

Abstract 4314: CEACAM7 an early detection biomarker for pancreatic ductal adenocarcinoma

Abstract Background: As per key statistics of American Cancer Society 2021, Pancreatic Cancer (PanCa) affects around 60,430 persons (31,950 men and 28,480 women) a year in the U.S. and is tricky to diagnose and treat. PanCa is the 4th leading cause of cancer death with a 5-year survival rate of only 9%, along with a poor prognosis. Generally (around 85-90%) PanCa are adenocarcinomas, such as Pancreatic ductal adenocarcinomas (PDAC). PDAC is now one of the most challenging tumor entities worldwide, characterized as a highly aggressive disease with dismal overall prognosis with a mortality rate near the incidence rate. For PDAC, no early tumor specific biomarker is available, therefore, novel early detection biomarker strategy is immediately required to upgrade the narrow diagnostic portfolio against PanCa. Considering this alarming situation, our team has identified a novel oncogenic protein, Carcinoembryonic antigen-related cell adhesion molecule 7 (CEACAM7). Our studies suggested high CEACAM7 expression in PDAC tumors and its association with patient survival. In this study we have investigated the potential role of CECAM7 for early PDAC diagnosis and its role in PDAC progression. Methodology: This study includes bioinformatics pre-screening using publicly available cancer databases followed by molecular biology techniques. To detect the degree of expression of CEACAM7 in normal pancreatic cell (HPNE) and PDAC progressive cellular models, we have minutely scrutinized the PDAC cell panel (HPAF-2, SU86.86 & Panc-1) & TMAs of PDAC patients, in terms of mRNA & protein expression level of CEACAM7. The confocal microscopy was performed to identify the intensity and localization of CEACAM7 protein. IHC analysis was also performed to identify the protein expressions in the pancreatitis and tumor cores along with their locations, grading and Mean Composite Score. Results: Bioinformatic results confirmed the relevance of CEACAM7 as a potential early diagnostic and prognostic marker of PDAC. HPAF2 (well differentiated) expressed highest mRNA and protein expression analyses, followed by SU86.86 (moderately differentiated) and very low expression in AsPc1 (poorly differentiated), and Panc1 (poorly differentiated). The IHC analyses of TMAs exhibited negligible or faint staining in pancreatitis (chronic and acute) and most of the positive cases were in tumor grading 1 & 2. Conclusion: Our observations clearly cite that CEACAM7 can serve as a potential early diagnostic and/or prognostic marker of PDAC and may also potentiate the sensitivity of the existing biomarker panel of PDAC. However, further studies are warranted to determine its clinical significance. Keywords: Pancreatic cancer, PDAC, CEACAM7, Early detection biomarker, Tumor grading Citation Format: Anupam Dhasmana, Swati Dhasmana, Sheema Khan, Murali M. Yallapu, Meena Jaggi, Subhash C. Chauhan. CEACAM7 an early detection biomarker for pancreatic ductal adenocarcinoma. [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 4314.

Abstract 5100: Combination of κλ bispecific antibodies targeting innate (CEAxCD47, NILK-2401) and adaptive immunity (CEAxCD3, NILK-2301 and CEAxCD28, NILK-3301) for next generation immunotherapy of CEA-expressing cancers

Abstract Background: The CEAxCD3 bispecific antibody (bsAb) NILK-2301 couples CEA (CEACAM5) on cancer cells and CD3 on T-cells inducing T-cell activation (signal 1) and tumor cell killing (TDCC). T-cell activation can be boosted by CEA-targeted CD28-costimulation (NILK-3301; signal 2). NILK-2401, carrying a fully effective IgG1 Fc, induces antibody-dependent phagocytosis (ADCP) and antibody-dependent cytotoxicity (ADCC) of tumor cells by co-targeting CEA and the innate immune checkpoint CD47 (“don’t eat me” signal). We present here next generation immunotherapy to overcome limited single class activity in CEA-expressing solid cancers. Methods: BsAbs were generated using LCB’s fully human κλ body platform. TDCC, ADCP, and ADCC with human PBMC or monocyte-derived macrophages were assessed using CEA+ colorectal (n=3), lung (n=2), and gastric (n=2) cancer lines. Combination activity of NILK-2401 + NILK-2301 (± NILK-3301) was assessed by flow cytometry. In vivo activity was tested in xenograft NOG or NSG/human PMBC-, HIS-, and hSIRPα/hCD47/hCD3/hCD28 transgenic mice. Safety data include binding to other CEACAMs, cytokine release in whole blood, erythrophagocytosis, platelet activation, exclusion of superagonism (NILK-3301), as well as PK- and tolerability in cynomolgus monkeys and Tg32-mice. Results: NILK-2301 induced dose-dependent killing of all tested cell lines, which was also visualized by live cell imaging. Combination of NILK-2301 (1 nM) + NILK-3301 vs. NILK-2301 alone (10 nM) increased TDCC (3-8-fold), T-cell activation (CD25, CD69, HLA-DR), cytokine secretion (interferon-γ, granzyme B, perforin), and CD4+/CD8+ T-cell proliferation. NILK-2401 blocked CD47-SIRPα interaction and induced ADCP/ADCC-mediated elimination of all cell lines. NILK-2301 + NILK-2401 treatment increased maximum activity (Emax) and reduced necessary dose of the T-cell bsAb to reach Emax. E.g., Emax of 30% killing (NILK-2301 alone) was increased in combination with NILK-2401 at 0.1/1/10 µg/mL to 40%, 80%, and 80%. In vivo, NILK-2301 (10 mg/kg IV, BIW) decreased tumor progression. NILK-2301/-3301 combination induced tumor regression in 8/8 mice. NILK-2401 delayed tumor growth vs. mean of control in 100% (15/15) of mice and prevented establishment of detectable tumors (>50mm3) in 53% (8/15). Results of double and quadruple transgenic mice, including triple bsAb combinations, will be presented at the meeting. No relevant safety signals were detected. Conclusions: NILK-2301 and NILK-2401 are active as single agents. Addition of NILK-2401 or NILK-3301 to NILK-2301 significantly increases activity, already at 10 -100x lower CEAxCD3 doses. GMP drug substance has been produced for NILK-2301 and NILK-2401. Generation of the clonal cell line for NILK-3301 clinical material production is ongoing. Citation Format: Anja Seckinger, Lise Nouveau, Sara Majocchi, Valéry Moine, Vanessa Buatois, Bruno Daubeuf, Franck Gueneau, Ulla Ravn, Krzysztof Masternak, Yves Poitevin, Emeline Rousset, Giovanni Magistrelli, Pauline Malinge, Limin Shang, Nicolas Fischer, Klaus Strein, Walter Ferlin, Dirk Hose. Combination of κλ bispecific antibodies targeting innate (CEAxCD47, NILK-2401) and adaptive immunity (CEAxCD3, NILK-2301 and CEAxCD28, NILK-3301) for next generation immunotherapy of CEA-expressing cancers. [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 5100.

Chimeric antigen receptor-T cells are effective against CEACAM5 expressing non-small cell lung cancer cells resistant to antibody-drug conjugates.

Chimeric antigen receptor-T (CAR-T) cells and antibody-drug conjugates (ADCs) are promising therapeutic strategies in oncology. The carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM5) is overexpressed in tumors including non-small cell lung cancer (NSCLC) and pancreatic ductal adenocarcinoma (PDAC), and is an attractive target for therapies based on CAR-T cell or/and ADCs. We previously developed a highly specific antibody-based CAR-T cells targeting CEACAM5 and the tumoricidal effect of CAR-T cells was proved against neuro-endocrine prostate cancer (NEPC) cells expressing CEACAM5. Here, we compare the anti-tumor efficacy of our CAR-T cells with that of an anti-CEACAM5 ADC being clinically evaluated against NSCLC. Our anti-CEACAM5 CAR-T cells showed cytotoxicity in a CEACAM5 surface concentration dependent manner and reduced tumor growth in both ADC-responsive and -non-responsive CEACAM5-expressing NSCLC cells in vitro and in vivo. In contrast, the ADC exhibited cytotoxicity independent on the CEACAM5 cell surface concentration. Even though clinical translation of CEACAM5 targeting CAR-T cell therapies is still in preclinical stage, our CAR-T cell approach could provide a potential therapeutic strategy for CEACAM5-positive cancer patients with resistance to ADCs.

CEACAM7 expression contributes to early events of pancreatic cancer

BackgroundThe trends of pancreatic cancer (PanCa) incidence and mortality are on rising pattern, and it will be a second leading cause of cancer related deaths by 2030. Pancreatic ductal adenocarcinoma (PDAC), major form of PanCa, exhibits a grim prognosis as mortality rate is very close to the incidence rate, due to lack of early detection methods and effective therapeutic regimen. Considering this alarming unmet clinic need, our team has identified a novel oncogenic protein, carcinoembryonic antigen-related cell adhesion molecule 7 (CEACAM7), that can be useful for spotting early events of PDAC. MethodologyThis study includes bioinformatics pre-screening using publicly available cancer databases followed by molecular biology techniques in PDAC progressive cell line panel and human tissues to evaluate CEACAM7 expression in early events of pancreatic cancer. ResultsPanCa gene and protein expression analysis demonstrated the significantly higher expression of CEACAM7 in PDAC, compared to other cancers and normal pancreas. Overall survival analysis demonstrated an association between the higher expression of CEACAM7 and poor patients’ prognosis with high hazard ratio. Additionally, in a performance comparison analysis CEACAM7 outperformed S100A4 in relation to PDAC. We also observed an increase of CEACAM7 in PDAC cell line panel model. However, poorly differentiated, and normal cell lines did not show any expression. Human tissue analysis also strengthened our data by showing strong and positive IHC staining in early-stage tumors. ConclusionOur observations clearly cite that CEACAM7 can serve as a potential early diagnostic and/or prognostic marker of PDAC and may also potentiate the sensitivity of the existing biomarker panel of PDAC. However, further studies are warranted to determine its clinical significance.

Effects of Helicobacter pylori adhesin HopQ binding to CEACAM receptors in the human stomach.

Helicobacter pylori has developed several strategies using its diverse virulence factors to trigger and, at the same time, limit the host's inflammatory responses in order to establish a chronic infection in the human stomach. One of the virulence factors that has recently received more attention is a member of the Helicobacter outer membrane protein family, the adhesin HopQ, which binds to the human Carcinoembryonic Antigen-related Cell Adhesion Molecules (CEACAMs) on the host cell surface. The HopQ-CEACAM interaction facilitates the translocation of the cytotoxin-associated gene A (CagA), an important effector protein of H. pylori, into host cells via the Type IV secretion system (T4SS). Both the T4SS itself and CagA are important virulence factors that are linked to many aberrant host signaling cascades. In the last few years, many studies have emphasized the prerequisite role of the HopQ-CEACAM interaction not only for the adhesion of this pathogen to host cells but also for the regulation of cellular processes. This review summarizes recent findings about the structural characteristics of the HopQ-CEACAM complex and the consequences of this interaction in gastric epithelial cells as well as immune cells. Given that the upregulation of CEACAMs is associated with many H. pylori-induced gastric diseases including gastritis and gastric cancer, these data may enable us to better understand the mechanisms of H. pylori's pathogenicity.

CEACAMS 1, 5, and 6 in disease and cancer: interactions with pathogens.

The CEA family comprises 18 genes and 11 pseudogenes located at chromosome 19q13.2 and is divided into two main groups: cell surface anchored CEA-related cell adhesion molecules (CEACAMs) and the secreted pregnancy-specific glycoproteins (PSGs). CEACAMs are highly glycosylated cell surface anchored, intracellular, and intercellular signaling molecules with diverse functions, from cell differentiation and transformation to modulating immune responses associated with infection, inflammation, and cancer. In this review, we explore current knowledge surrounding CEACAM1, CEACAM5, and CEACAM6, highlight their pathological significance in the areas of cancer biology, immunology, and inflammatory disease, and describe the utility of murine models in exploring questions related to these proteins.

Avoid being trapped by your liver: ischemia-reperfusion injury in liver transplant triggers S1P-mediated NETosis.

Liver transplantation can be a life-saving treatment for end-stage hepatic disease. Unfortunately, some recipients develop ischemia-reperfusion injury (IRI) that leads to poor short- and long-term outcomes. Recent work has shown neutrophils contribute to IRI by undergoing NETosis, a form of death characterized by DNA ejection resulting in inflammatory extracellular traps. In this issue of the JCI, Hirao and Kojima et al. report that sphingosine-1-phosphate (S1P) expression induced by liver transplant-mediated IRI triggers NETosis. They also provide evidence that neutrophil expression of the carcinoembryonic antigen-related cell adhesion molecule-1 (CC1) long isoform inhibited NETosis by controlling S1P receptor-mediated autophagic flux. These findings suggest stimulating regulatory mechanisms that suppress NETosis could be used to prevent IRI.

Increased Carcinoembryonic Antigen-related Cell Adhesion Molecule 5 (CEACAM 5) Production and Its Role in Human Nasal Polyps

A previous study demonstrates that CEACAM5 gene expression is highly upregulated in tissues from chronic rhinosinusitis with nasal polyps (CRSwNP) in microarray analysis. CEACAM5 protein production, localization, and association with other inflammatory markers were investigated in CRS.

Neutrophil CEACAM1 determines susceptibility to NETosis by regulating the S1PR2/S1PR3 axis in liver transplantation.

Neutrophils, the largest innate immune cell population in humans, are the primary proinflammatory sentinel in the ischemia-reperfusion injury (IRI) mechanism in orthotopic liver transplantation (OLT). Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1, CC1, or CD66a) is essential in neutrophil activation and serves as a checkpoint regulator of innate immune-driven IRI cascade in OLT. Although CC1 alternative splicing generates two functionally distinct short and long cytoplasmic isoforms, their role in neutrophil activation remains unknown. Here, we undertook molecular and functional studies to interrogate the significance of neutrophil CC1 signaling in mouse and human OLT recipients. In the experimental arm, we employed a mouse OLT model to document that ablation of recipient-derived neutrophil CC1-long (CC1-L) isotype aggravated hepatic IRI by promoting neutrophil extracellular traps (NETs). Notably, by regulating the S1P-S1PR2/S1PR3 axis, neutrophil CC1-L determined susceptibility to NET formation via autophagy signaling. In the clinical arm, liver grafts from 55 transplant patients selectively enriched for neutrophil CC1-L showed relative resistance to ischemia-reperfusion (IR) stress/tissue damage, improved hepatocellular function, and clinical outcomes. In conclusion, despite neutrophils being considered a principal villain in peritransplant tissue injury, their CC1-L isoform may serve as a regulator of IR stress resistance/NETosis in human and mouse OLT recipients.

CEACAM1 Marks Highly Suppressive Intratumoral Regulatory T Cells for Targeted Depletion Therapy.

Regulatory T cells (Tregs) exert immunosuppressive functions and hamper antitumor immune responses in the tumor microenvironment. Understanding the heterogeneity of intratumoral Tregs, and how it changes with tumor progression, will provide clues regarding novel target molecules of Treg-directed therapies. From 42 patients with renal cell carcinoma and 5 patients with ovarian cancer, immune cells from tumor and peripheral blood were isolated. We performed multicolor flow cytometry and RNA-sequencing to characterize the phenotypes and heterogeneity of intratumoral Tregs. In vitro functional assays were performed to evaluate suppressive capacity of Tregs and effect of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1)-mediated depletion. The CT26 tumor model was used to evaluate the association between intratumoral Tregs and tumor growth, and examine the in vivo role of CEACAM1+ intratumoral Tregs on antitumor immunity. We found that CEACAM1 was selectively expressed on intratumoral Tregs, whereas its expression on peripheral Tregs or other immune cells was low. The CEACAM1+ intratumoral Tregs accumulated with tumor progression, whereas the CEACAM1- subset did not. Notably, we found that CEACAM1 marked intratumoral Tregs that exhibited highly suppressive and activated phenotypes with substantial clonal expansion. Depletion of CEACAM1-expressing cells from tumor-infiltrating leukocytes led to increased effector functions of tumor-infiltrating T cells. Moreover, CEACAM1+ cell depletion further enhanced anti-PD-1-mediated reinvigoration of exhausted CD8+ T cells. CEACAM1 marks highly suppressive subset of intratumoral Tregs, and can be a target for selective depletion of intratumoral Tregs. These results may inform future studies on CEACAM1-mediated depletion in patients with cancer.

Carcinoembryonic Antigen–Related Cell Adhesion Molecule Type 5 Receptor–Targeted Fluorescent Intraoperative Molecular Imaging Tracer for Lung Cancer

Localization of subcentimeter ground glass opacities during minimally invasive thoracoscopic lung cancer resections is a significant challenge in thoracic oncology. Intraoperative molecular imaging has emerged as a potential solution, but the availability of suitable fluorescence agents is a limiting factor. To evaluate the suitability of SGM-101, a carcinoembryonic antigen-related cell adhesion molecule type 5 (CEACAM5) receptor-targeted near-infrared fluorochrome, for molecular imaging-guided lung cancer resections, because glycoprotein is expressed in more than 80% of adenocarcinomas. For this nonrandomized, proof-of-principal, phase 1 controlled trial, patients were divided into 2 groups between August 1, 2020, and January 31, 2022. Patients with known CEACAM5-positive gastrointestinal tumors suggestive of lung metastasis were selected as proof-of-principle positive controls. The investigative group included patients with lung nodules suggestive of primary lung malignant neoplasms. Patients 18 years or older without significant comorbidities that precluded surgical exploration with suspicious pulmonary nodules requiring surgical biopsy were included in the study. SGM-101 (10 mg) was infused up to 5 days before index operation, and pulmonary nodules were imaged using a near-infrared camera system with a dedicated thoracoscope. SGM-101 localization to pulmonary nodules and its correlation with CEACAM5 glycoprotein expression by the tumor as quantified by tumor and normal pulmonary parenchymal fluorescence. Ten patients (5 per group; 5 male and 5 female; median [IQR] age, 66 [58-69] years) with 14 total lesions (median [range] lesion size, 0.91 [0.90-2.00] cm) were enrolled in the study. In the control group of 4 patients (1 patient did not undergo surgical resection because of abnormal preoperative cardiac clearance findings that were not deemed related to SGM-101 infusion), the mean (SD) lesion size was 1.33 (0.48) cm, 2 patients had elevated serum CEA markers, and 2 patients had normal serum CEA levels. Of the 4 patients who underwent surgical intervention, those with 2+ and 3+ tissue CEACAM5 expression had excellent tumor fluorescence, with a mean (SD) tumor to background ratio of 3.11 (0.45). In the patient cohort, the mean (SD) lesion size was 0.68 (0.22) cm, and no elevations in serum CEA levels were found. Lack of SGM-101 fluorescence was associated with benign lesions and with lack of CEACAM5 staining. This in-human proof-of-principle nonrandomized controlled trial demonstrated SGM-101 localization to CEACAM5-positive tumors with the detection of real-time near-infrared fluorescence in situ, ex vivo, and by immunofluorescence microscopy. These findings suggest that SGM-101 is a safe, receptor-specific, and feasible intraoperative molecular imaging fluorochrome that should be further evaluated in randomized clinical trials. ClinicalTrials.gov identifier: NCT04315467.

Abstract B029: CM24, a novel mAb against carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), suppresses Neutrophil Extracellular Trap (NET)-induced migration and metastasis of cancer cells

Abstract Background: NETs are web-like structures involved in cancer immunoediting, progression, metastatic spread and play a key role in the tumor microenvironment. NETs intensify tumor aggressiveness by enhancing cancer migration and invasion capacity and can entrap circulating cancer cells and facilitate metastasis. A better understanding of the crosstalk between cancer and NETs is crucial for the development of novel therapeutic interventions blocking cancer evasion mechanisms and preventing metastatic spread. CEACAM1, plays an important role in tumor immune evasion, metastasis, angiogenesis, and was recently suggested to be a part of the NET structure. In this study, we present for the first time the NET-related activity of CM24. We demonstrate that CM24 binds to the NET structure, significantly suppresses the NET-induced cancer cell migration and inhibits metastases in mice xenografts. Methods: CEACAM1 expression and CM24 binding in whole blood were tested by FACS and analyzed for different immune subpopulations. Expression of CEACAM1 on tumor cells and the tumor microenvironment was tested using IHC and multi-plex IF of tissue microarray (TMA). Direct binding of CM24 to NETs was tested. Primary human neutrophils were stimulated with PMA to produce NETs, fixed and stained for myeloid peroxidase (MPO), DAPI & CM24. Images were captured using confocal microscopy. CM24 effect on NET-induced migration of cancer cells was tested using a Boyden chamber chemoattractant assay in various cancer cells±NETs. In a Xenograft lung lesion model, NOD-SCID mice were IV engrafted with melanoma cells and treated with either CM24 or isotype control ±Tumor Infiltrating Immune cells (TILs). Mice were sacrificed, and lungs were removed to assess tumor burden. Results: Whole blood analysis showed that neutrophils are one of the main cell type that expresses CEACAM1 and binds CM24. CEACAM1 staining of TMAs demonstrated an enhanced expression of CEACAM1 on both the tumor cells and the TILs in the TME as compared to normal tissues. Multiplex IF staining demonstrated CEACAM1 expression on infiltrating T cells and most prominently on the myeloid cells. Direct Binding of CM24 to the NET structure was demonstrated by multiplex staining for MPO (NET marker), DAPI (DNA), and CEACAM1 and captured by confocal microscopy. We next tested the effect of CM24 on cancer cell migration with or without NETs. CM24 significantly suppressed the NET-induced migration of multiple CEACAM1 expressing cancer cells, while the isotype control had no effect. In a xenograft lung lesion model, CM24 significantly inhibited metastasis of human melanoma to the lung. A greater effect of CM24 on lung lesions was shown in the presence of TILs. Conclusions: These findings show that CEACAM1 blockade by CM24 can inhibit NET-mediated cancer cell migration in-vitro and suppress metastasis in-vivo. This novel MOA provides a rationale for the anti-metastatic activity of CM24 and may contribute to improving patient outcome. Clinical studies to evaluate CM24 efficacy are being conducted in PDAC patients. Citation Format: Hadas Reuveni, Hava Ben David, Julie Rumble, Tomer Meirson, Michael Schickler. CM24, a novel mAb against carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), suppresses Neutrophil Extracellular Trap (NET)-induced migration and metastasis of cancer cells [abstract]. In: Proceedings of the AACR Special Conference: Cancer Metastasis; 2022 Nov 14-17; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;83(2 Suppl_2):Abstract nr B029.

Increased CEACAM1 expression on peripheral blood neutrophils in patients with rheumatoid arthritis.

Altered expression of adhesion molecules in immune cells has been demonstrated in rheumatoid arthritis (RA). Carcinoembryonic-antigen-related cell-adhesion molecule 1 (CEACAM1) is an adhesion molecule that acts as a coinhibitory receptor in the immune system. We investigated the role of CEACAM1 in immune cell subsets of patients with RA. Peripheral blood was obtained from 37 patients with RA and 20 healthy controls (HC). The expression of CEACAM1 and T-cell immunoglobulin mucin domain molecule (TIM) -3 on peripheral blood mononuclear cells and neutrophils was analyzed by flow cytometry. Intracellular TIM-3 expression was analyzed using cellular lysates by Western blot analysis. Serum levels of soluble CEACAM1 (sCEACAM1) were estimated by an enzyme-linked immunosorbent assay. CEACAM1 expression was not detected in peripheral blood mononuclear cells, including in CD14(+) monocytes and CD3(+) lymphocytes isolated from patients with RA or HC. However, substantial cell-surface expression of CEACAM1 was detected in peripheral blood neutrophils, and it was significantly elevated in samples from patients with RA without remission compared to those in remission. There was no significant difference in serum levels of sCEACAM1 between patients with RA and HC. Cell-surface expression of TIM-3 was not detected in peripheral blood neutrophils from patients with RA or HC but was seen in CD14(+) monocytes. However, there was no significant difference in TIM-3 expression on monocytes between patients with RA and HC. Our data indicate that cell-surface expression of CEACAM1 on peripheral blood neutrophils are higher in patients with RA and that it is associated with rheumatoid inflammation. Further studies are needed to explore the potential role of CEACAM1 in rheumatoid inflammatory pathways.

The Correlation of Ten Immune Checkpoint Gene Expressions and Their Association with Gastric Cancer Development.

In the immunotherapy based on immune checkpoint inhibition (IC), additional ICs are being studied to increase its effectiveness. An almost unstudied feature is the possible co-expression of ICs, which can determine the therapeutic efficacy of their inhibition. For the selection of promising ICs, information on the association of their expression with cancer development may be essential. We have obtained data on the expression correlation of ADAM17, PVR, TDO2, CD274, CD276, CEACAM1, IDO1, LGALS3, LGALS9, and HHLA2 genes in gastric cancer (GC). All but one, TDO2, have other IC genes with co-expression at some stage. At the metastatic stage, the expression of the IDO1 does not correlate with any other gene. The correlations are positive, but the expressions of the CD276 and CEACAM1 genes are negatively correlated. The expression of TDO2 and LGALS3 is associated with GC metastasis. The expression of TDO2 four-fold higher in metastatic tumors than in non-metastatic tumors, but LGALS3 was two-fold lower. The differentiation is associated with IDO1. The revealed features of TDO2, with a significant increase in expression at the metastatic stage and the absence of other IC genes with correlated expression indicates that the prospect of inhibiting TDO2 in metastatic GC. IDO1 may be considered for inhibition in low-differentiated tumors.

Bispecific antibodies targeting CD40 and tumor-associated antigens promote cross-priming of T cells resulting in an antitumor response superior to monospecific antibodies

BackgroundIndications with poor T-cell infiltration or deficiencies in T-cell priming and associated unresponsiveness to established immunotherapies represent an unmet medical need in oncology. CD40-targeting therapies designed to enhance antigen presentation,...

Antibody Drug Conjugates in Lung Cancer

An antibody-drug conjugate (ADC) comprises a monoclonal antibody that is specific to a tumor cell protein, bound to a cytotoxic agent, known as the payload. The use of ADCs is already common practice in several cancers, thanks to their efficacy and potentially more manageable toxicity profile, resulting from the release of the cytostatic payload directly in the tumors. Currently, early-phase trials of ADCs in non-small cell lung cancer are rapidly gaining ground, with promising results targeting HER2 (human epidermal growth factor 2), HER3, TROP2 (trophoblast cell surface antigen 2), MET, CEACAM5 (carcinoembryonic antigen-related cell adhesion molecule 5), and PTK7 (tyrosine protein kinase-like 7). Unfortunately, in small cell lung cancer, trials targeting the ubiquitous DLL3 (delta-like ligand 3) protein have failed to show clinically relevant results, despite significant toxicity.