Trophoblast Cell Surface Antigen Research Articles

Abstract OT1-07-05: TROPiCS-02: phase 3 study of Sacituzumab Govitecan (IMMU-132) in relapsed/refractory hormonal receptor-positive (HR+) human epidermal growth factor receptor 2 negative (HER2-) metastatic breast cancer (MBC)

Abstract Background: Patients (pts) with HR+/HER2- MBC for whom first- and second-line treatments have failed need additional treatment approaches. Trophoblast cell surface antigen 2 (Trop-2) is highly expressed in breast cancer and linked to poor prognosis. Sacituzumab govitecan (SG) is a novel antibody-drug conjugate comprising an anti-Trop-2 monoclonal antibody conjugated to SN-38 (active metabolite of irinotecan) in a high drug-to-antibody ratio of 7.6. It has a unique hydrolysable linker that allows release of SN-38 intracellularly and in the tumor microenvironment. In the HR+/HER2- MBC cohort in the IMMU 132-01 phase 1/2 basket study, SG showed a 31% objective response rate (ORR), median progression-free survival (PFS) of 6.8 mo, and a predictable and manageable safety profile with low discontinuation rates due to adverse events (Kalinsky 2018 SABCS). Trial Design: Trop-2 Investigation in Cancer with Sacituzumab (TROPiCS-02; NCT03901339) is a randomized, open-label, phase 3 study in pts with HR+/HER2- MBC and ≥1 measurable target lesion according to Response Evaluation Criteria in Solid Tumors version 1.1 after failure of ≥2, but ≤4 prior chemotherapy regimens. Pts are randomized 1:1 to receive SG (10 mg/kg intravenously, days 1 and 8 every 21 days) or treatment of physician’s choice (TPC, determined pre-randomization: eribulin, capecitabine, gemcitabine, vinorelbine). Pts continue treatment until progression requiring discontinuation or unacceptable toxicity. Eligible pts are females or males ≥18 y old with documented evidence of HR+/HER2- MBC, Eastern Cooperative Oncology Group score of 0 or 1, and adequate safety laboratories. Pts must have received prior taxanes in any setting, ≥1 prior anticancer hormonal treatment, and ≥1 cyclin dependent kinase 4/6 inhibitor in the metastatic setting, with documented progression after most recent therapy. Pts also must be eligible to receive one of the TPC agents. The primary endpoints are PFS (local assessment) and ORR; additional endpoints include overall survival (OS), duration of response (DOR), and safety. The hazard ratio of the primary endpoint of PFS and its associated 95% confidence interval (CI) will be estimated using a Cox proportional-hazards model; the 2-sided 95% CIs of ORR will be calculated by the Clopper-Pearson exact method. Exploratory endpoints include Trop-2 expression and efficacy in relation to Trop-2 expression and blood and tumor biomarkers. Biomarker samples are taken at baseline, pre-dose at cycle 2, and at disease progression/end of treatment. Approximately 400 pts will be randomized across multiple countries. Citation Format: Hope S Rugo, Aditya Bardia, Sara M. Tolaney, Carlos Arteaga, Javier Cortes, Joohyuk Sohn, Frederik Marmé, Quan Hong, Scott Hofsess, Martin Olivo, André Fabrice, Peter Schmid. TROPiCS-02: phase 3 study of Sacituzumab Govitecan (IMMU-132) in relapsed/refractory hormonal receptor-positive (HR+) human epidermal growth factor receptor 2 negative (HER2-) metastatic breast cancer (MBC) [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr OT1-07-05.

Preclinical Activity of Sacituzumab Govitecan, an Antibody-Drug Conjugate Targeting Trophoblast Cell-Surface Antigen 2 (Trop-2) Linked to the Active Metabolite of Irinotecan (SN-38), in Ovarian Cancer.

Background: Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy. Sacituzumab govitecan (SG) is a novel antibody-drug-conjugate (ADC) targeting trophoblast-antigen-2 (Trop-2), a cell surface glycoprotein highly expressed in many epithelial tumors, to deliver SN-38, the active metabolite of irinotecan. This study aimed to evaluate Trop-2 expression in EOC tissues and the preclinical activity of SG against primary EOC cell lines and xenografts.Methods: Trop-2 expression was assessed in 90 formalin-fixed-paraffin-embedded tumors and nine primary tumor cell lines by immunohistochemistry (IHC) and flow cytometry, respectively. Trop-2 expression and cell viability after exposure to SG in primary tumor cell lines, non-targeting control ADC, and SG-parental antibody hRS7 were evaluated using flow-cytometry-based-assays. Antibody-dependent-cell-cytotoxicity (ADCC) against Trop-2+ and Trop-2– EOC cell lines was tested in vitro using 4 h Chromium-release-assays. In vivo activity of SG was evaluated against Trop-2+ EOC xenografts.Results: Moderate-to-strong staining was seen in 47% (42/90) of ovarian tumors by IHC while 89% (8/9) of the primary EOC cell lines overexpressed Trop-2 by flow cytometry. EOC Trop-2+ were significantly more sensitive to SG compared to control ADC (p < 0.05). Both SG and hRS7 mediated high ADCC activity against Trop-2+ cell lines. SG also induced significant bystander killing of Trop-2– tumor cells admixed with Trop-2+ EOC cells. In in vivo experiments SG treatment demonstrated impressive anti-tumor activity against chemotherapy-resistant EOC xenografts.Conclusion: SG demonstrates remarkable preclinical activity against biologically aggressive and chemotherapy-resistant EOC cell lines and a significant bystander effect against EOC cell lines with heterogenous Trop-2 expression. Clinical trials are warranted.

Knockdown of Trop2 inhibits proliferation and migration and induces apoptosis of endometrial cancer cells via AKT/β-catenin pathway.

Endometrial cancer (EC) is the most common gynaecologic malignancy in western countries and has been reported to account for about 7% of female malignant tumours and 20% to 30% of female genital system malignant tumours. Accumulating evidence showed the expression of human trophoblast cell surface antigen 2 (Trop2) was abnormal in many cancers; however, the expression and role of Trop2 in EC are not clear. The Trop-2 protein expression was detected by western blot in EC cells. Cell proliferation, apoptosis, and migration were measured by CCK-8, flow cytometry, and Transwell assay, respectively. The epithelial mesenchymal transition (EMT) and AKT/β-catenin signalling pathway-related proteins in EC cell lines were detected by western blot assay following Trop2 gene silencing. The present study revealed that the Trop2 protein was highly expressed in EC cell lines compared with human endometrial epithelial cells. The Trop2 mRNA and protein were obviously decreased following transfection with Trop2-siRNA sequence in KLE and Ishikawa cells. Meanwhile, Trop2 gene silencing in KLE and Ishikawa cells strongly inhibited cell proliferation and migration and increased cell apoptosis. Investigation into the molecular mechanism indicated that the Trop2 gene silencing suppressed EMT and AKT/β-catenin signalling pathway activation. SIGNIFICANCE OF THE STUDY: These findings suggested that Trop2 silencing inhibited EC cell proliferation and migration and promoted cell apoptosis. The mechanism might be related to the inhibition of the AKT/β-catenin signalling pathway in EC cells. Therefore, Trop2 may be a potential therapeutic target for the treatment of EC.

Sacituzumab govitecan, an antibody-drug conjugate targeting trophoblast cell-surface antigen 2, shows cytotoxic activity against poorly differentiated endometrial adenocarcinomas invitro and invivo.

Endometrial cancer is the most common gynecologic malignancy in developed countries. The antibody–drug conjugate (ADC) sacituzumab govitecan (SG) targets trophoblast cell‐surface antigen‐2 (Trop‐2) – a cell‐surface glycoprotein highly expressed in many epithelial tumors – and delivers the active metabolite of irinotecan SN‐38 to Trop‐2‐positive tumor cells. We evaluated Trop‐2 expression in endometrial endometrioid carcinoma (EC) tissues and the activity of SG against primary poorly differentiated EC cell lines and xenografts. Trop‐2 expression was assessed in 143 formalin‐fixed–paraffin‐embedded tumors and seven primary tumor cell lines by immunohistochemistry and flow cytometry, respectively. Cell viability of primary tumor cell lines was assessed following exposure to SG, or control antibodies. Antibody‐dependent cell cytotoxicity (ADCC) against Trop‐2‐positive and Trop‐2‐negative EC cell lines was measured in vitro using 4‐h chromium release assays. A Trop‐2‐positive EC xenograft model was used to determine the in vivo activity of SG. Moderate‐to‐strong staining was detected in 84% (120/143) of EC samples, whereas 43% (3/7) of the primary EC cell lines tested overexpressed Trop‐2. EC cell lines overexpressing Trop‐2 were significantly more sensitive to SG compared to control ADC (P = 0.014 and P = 0.005). Both SG and the unconjugated parental antibody hRS7 mediated high ADCC against Trop‐2‐positive cell lines. Moreover, SG induced significant bystander killing of Trop‐2‐negative tumors cocultured with Trop‐2‐positive tumors. In the xenograft model, intravenous administration of SG twice weekly for three weeks was well tolerated and demonstrated impressive tumor growth inhibition against poorly differentiated, chemotherapy‐resistant EC xenografts (P = 0.011). In summary, SG is a novel ADC with remarkable preclinical activity against poorly differentiated EC cell lines overexpressing Trop‐2. These findings warrant future clinical trials.

MiR-488-3p sponged by circ-0000495 and mediated upregulation of TROP2 in head and neck squamous cell carcinoma development.

TROP2 (trophoblast cell surface antigen 2) overexpression has been reported in many human cancers. The correlation between TROP2 and tumor aggressiveness has implied it could be a prognostic indicator. However, the roles of TROP2 and their underlying mechanisms remain of great interest in head and neck squamous cell carcinoma (HNSCC) biology. In the current study, the prognostic significance of TROP2 in HNSCC archival samples was determined using immunohistochemistry. Quantitative reverse transcriptase PCR (qRT-PCR) was used to measure the phenotypic effects of TROP2 knockdown, miR-488-3p re-expression, and circRNAs expression. Cell viability, migration/invasion as well as in vivo tumor formation assays were accessed. The interactions of miRNAs-TROP2 or circRNAs-miRNAs were determined by qRT-PCR, western blot analysis and luciferase assays. TROP2 was demonstrated overexpression in HNSCC patients and cancer cell lines. High expression of TROP2 was significantly associated with patient relapse. TROP2 promoted tumor cell proliferation, migration, invasion, and tumor growth, through AKT and MAPK pathways. Further investigation revealed that TROP2 is a direct target of miR-488-3p, while circ-0000495 bounds to miR-488-3p. Our study unraveled a novel mechanism by which down-regulation of miR-488-3p sponged by circ-0000495 releases its epigenetic silencing to TROP2. The increased TROP2 promotes tumor proliferation, therefore, providing evidence in support of targeting the circ-0000495/miR-488-3p/TROP2 axis in contributing to HNSCC therapy and preventing tumor metastasis.

Can TROP2 be used as a prognostic marker in endometrioid endometrial carcinoma?

Endometrioid-type endometrial carcinoma is the most common malignancy of the female genital tract in developed countries. The prognosis greatly depends on the grade and stage of the disease. In some patients, the disease recurs in a short time after the surgical/medical therapy. Hence, it is important to predict the patients who will have worse prognosis at the beginning, to choose the appropriate treatment; resuming the search of new prognostic markers. Therefore, our study aimed to detect trophoblast cell surface antigen 2 (TROP2) as a new prognostic marker. The patients who underwent a hysterectomy and diagnosed with endometrioid-type endometrial carcinoma were evaluated retrospectively and TROP2 immunostain was performed to their tumoral slides. We evaluated TROP2 expressions in 102 patients immunohistochemically who underwent hysterectomy with the diagnosis of endometrioid-type endometrial carcinoma histopathologically and correlated them with the other generally accepted prognostic parameters. The Kolmogorov-Smirnov test and Q-Q plot test were used to verify the normality of the distribution of continuous variables. The Chi-square/Fisher's exact tests were used for categorical variables. Analyses were performed with SPSS Statistics for Windows, Version 20. High overexpression of TROP2 was seen in larger, higher-grade, deeper-invasive tumors, tumors with vascular invasion, and pelvic-lymph-node metastasis. These results were statistically significant (P ≤ 0.05). Overexpression of TROP2 in endometrioid-type endometrial carcinoma seems to be a poor prognostic factor; it may be useful in determining the biologically more aggressive tumors before the treatment. This early determination is very important to choose the appropriate surgery, adjuvant-treatments, and follow-up.

In vitro and in vivo activity of sacituzumab govitecan, an antibody-drug conjugate targeting trophoblast cell-surface antigen 2 (Trop-2) in uterine serous carcinoma

ObjectiveUterine serous carcinoma (USC) is an aggressive variant of endometrial cancer with poor prognosis. Sacituzumab govitecan (SG) is a novel antibody-drug-conjugate (ADC) targeting trophoblast cell-surface antigen 2 (Trop-2), a transmembrane-calcium-signal-transducer, to deliver SN-38, the active metabolite of irinotecan. The objective of this study was to evaluate the expression of Trop-2 in USC and the preclinical activity of SG against primary USC cell-lines and xenografts. MethodsWe used immunohistochemistry (IHC) and flow-cytometry-based assays to evaluate Trop-2 expression and cell-viability in USC tissue and primary tumor-cell-lines after exposure to SG, non-targeting control ADC, and naked antibody hRS7-IgG. Antibody-dependent-cell-cytotoxicity (ADCC) against Trop-2+ and Trop-2- USC cell-lines was evaluated in vitro using 4-hr-Chromium-release-assays. In vivo activity of SG was tested against Trop-2+ USC xenografts by intravenous administration of SG, control ADC, and hRS7. ResultsTrop-2 expression by IHC was detected in 95.1% of USC samples (99/104). Primary tumor cell-lines overexpressing Trop-2 were significantly more sensitive to SG when compared to control ADC (p <0.05). Both SG and hRS7 mediated ADCC in Trop2+ USC cell-lines while no cytotoxicity was detected against Trop-2- cells. SG induced significant bystander killing of Trop-2- tumors when admixed with Trop-2+ tumors. SG caused growth-inhibition and increased survival in SG treated mice harboring Trop-2+ xenografts when compared to controls (p <0.05). ConclusionsSG is remarkably active against USC overexpressing Trop-2 in vitro and in vivo. Our results combined with SG clinical responses recently reported against multiple chemotherapy resistant human tumors further support clinical development of SG in USC patients with advanced/recurrent disease.

Abstract C026: DS-1062a, a novel TROP2-targeting antibody-drug conjugate with a novel DNA topoisomerase I inhibitor DXd, demonstrates potent antitumor activity in preclinical models

Abstract Background: A trophoblast cell surface antigen 2 (TROP2) is a 36-kDa single-pass transmembrane protein overexpressed in various epithelial tumors including non-small cell lung cancer (NSCLC) with relatively low and restricted expression in normal tissues, and is associated with aggressive tumor behavior. Therefore, TROP2 could be an attractive target for cancer therapy. We created DS-1062a, TROP2-targeting antibody-drug conjugate (ADC) with Daiichi Sankyo DXd technology using a novel DNA topoisomerase I inhibitor DXd. In this study, the pharmacological activity and the mechanism of action of DS-1062a were evaluated in preclinical in vitro and in vivo models. Methods: In vitro cell growth inhibitory and in vivo antitumor activities of DS-1062a were evaluated using TROP2-high and -low tumor cell lines and xenograft mouse models. Internalization and intracellular trafficking of DS-1062a in tumor cells were analyzed by immunofluorescence microscopy. Induction of DNA damage and apoptosis to tumor cells by DXd payload released from DS-1062a were assessed by western blot and immunohistochemistry. Pharmacokinetic profiles of DS-1062a were also analyzed in xenograft mouse model. Results: DS-1062a showed in vitro cell growth inhibitory activity to TROP2-high tumor cells, but not to TROP2-low tumor cells. It was observed with confocal microscopy that DS-1062a was co-localized with a lysosomal marker LAMP-2 after internalizing into TROP2-high tumor cells. The amount of DXd payload released from TROP2-high tumor cells after the in vitro treatment with DS-1062a was higher than that of TROP2-low tumor cells. DNA damage and apoptosis were induced in TROP2-high tumor cells after the in vitro treatment with DXd and DS-1062a, but not with isotype control IgG ADC. Similarly, DS-1062a exhibited strong antitumor activity with tumor regression in several TROP2-high tumors including NSCLC and DXd accumulation and DNA damage in TROP2-high tumors were observed in DS-1062a-treated tumors, but not in isotype control IgG ADC-treated tumors. Pharmacokinetic profiles of DS-1062a in xenograft mouse model were preferable. Conclusion: Based on these preclinical results, DS-1062a could provide a valuable therapy with a potential benefit in TROP2-expressing cancers in the clinical setting. A first-in-human phase 1 study in patients with advanced solid tumors is in progress (NCT03401385). Citation Format: Daisuke Okajima, Junko Yamaguchi, Michiko Kitamura, Reiko Kamei, Takanori Maejima, Tomoko Shibutani, Satoru Yasuda, Tadashi Toki, Tsuyoshi Karibe, Tomomichi Fujitani, Takashi Nakada, Riki Goto, Yutaka Noguchi, Yuki Abe, Toshinori Agatsuma. DS-1062a, a novel TROP2-targeting antibody-drug conjugate with a novel DNA topoisomerase I inhibitor DXd, demonstrates potent antitumor activity in preclinical models [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr C026. doi:10.1158/1535-7163.TARG-19-C026

TROP2 increases growth and metastasis of human oral squamous cell carcinoma through activation of the PI3K/Akt signaling pathway.

Most malignant neoplasms of the oral cavity are oral squamous cell carcinoma (OSCC), which is a type of highly malignant tumor with a propensity for forming distant metastases. Trophoblast cell surface antigen 2 (TROP2) is a transmembrane protein that is overexpressed in several types of tumor cells, although its role and regulatory mechanism in OSCC have not been determined. The aim of the present study was to examine the effects of TROP2 in human OSCC cell lines. The present study demonstrated that TROP2 protein expression was upregulated in OSCC cell lines. Transfection of short hairpin RNA (shRNA) targeting TROP2 (sh-TROP2) reduced cell proliferation, migration and invasion of OSCC cell lines, whereas overexpression of TROP2 increased proliferation, migration and invasion. sh-TROP2 transfection in OSCC cell lines inhibited tumor growth in OSCC mouse models. Furthermore, TROP2 expression activated the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway in human OSCC cells. These results suggest that TROP2 induces cell growth, migration and invasion through activation of the PI3K/Akt signaling pathway in OSCC cells.

High Expression of Trophoblast Cell Surface Antigen 2 in Triple-negative Breast Cancer Identifies a Novel Therapeutic Target

Abstract Objectives Patients with triple-negative breast cancer (TNBC) respond poorly to current therapeutic modalities. The newly FDA-approved drug conjugate, Sacituzumab Govitecan, targeting antitrophoblastic cell surface antigen 2 (Trop-2), offers a new modality to treat these subtypes of breast carcinoma (BC). Our study examines expression of Trop-2 by immunohistochemistry (IHC) in TNBC. Methods Forty cases of TNBC were selected from our files (34 ductal, 4 pleomorphic lobular, 1 metaplastic, and 1 mixed ductal and lobular carcinoma). A control group included 11 luminal A-like, 11 luminal B-like, and 8 HER2-like BC. Immunostaining for Trop-2 was performed on 4-micron-thick tissue sections using goat polyclonal antibody (R&amp;D Systems). Three pathologists individually scored the % and intensity of membranous staining. The % of staining was defined as &lt;10%, 10%-50%, and &gt;50%. The intensity of staining was scored as 3+ (crisp circumferential membranous staining), 2+ (medium membranous staining), 1+ (patchy weak membranous staining), and 0 (no staining). A Kruskal-Wallis H-test and pairwise comparison with Bonferroni correction was performed to compare % and intensity of staining in TNBC to control group. Results TNBC exhibited stronger Trop-2 staining in both intensity and extent when compared to the control. The result was significantly different, χ2 (4) = 17.427, P = .001. A pairwise comparison with Bonferroni correction found significant difference between TNBC (42.55) and luminal A (16.65) (P &lt; .001). There was significant agreement (P &lt; .001) among 3 independent pathologists for assessing % and intensity of staining. Conclusion The extent and intensity of staining for Trop-2 is higher in TNBC than in other molecular subtypes of BC. These findings suggest that patients with TNBC may potentially benefit from this targeted therapy. Furthermore, identification of Trop-2 in other molecular subtypes may expand the therapeutic potential of this new drug. Further studies are needed to determine the efficacy of this marker.

Trophoblast cell surface antigen 2 (Trop-2) phosphorylation by protein kinase C α/δ (PKCα/δ) enhances cell motility

Dysfunction of tight junctions is a critical step during the initial stage of tumor progression. Trophoblast cell surface antigen 2 (Trop-2) belongs to the family of tumor-associated calcium signal transducer (TACSTD) and is required for the stability of claudin-7 and claudin-1, which are often dysregulated or lost in carcinogenesis. Here, we investigated the effects of Trop-2 phosphorylation on cell motility. Analyses using HCT116 cells expressing WT Trop-2 (HCT116/WT) or Trop-2 alanine-substituted at Ser-303 (HCT116/S303A) or Ser-322 (HCT116/S322A) revealed that Trop-2 is phosphorylated at Ser-322. Furthermore, coimmunoprecipitation and Transwell assays indicated that Trop-2 S322A interacted with claudin-7 the strongest, and a phosphomimetic variant, Trop-2 S322E, the weakest and that HCT116/S322E cells have the highest motility and HCT116/S322A cells the lowest. All cell lines had similar levels of claudin-7 mRNA, but levels of claudin-7 protein were markedly decreased in the HCT116/S322E cells, suggesting posttranscriptional control of claudin-7. Moreover, claudin-7 was clearly localized to cell-cell borders in HCT116/S322A cells but was diffusely distributed on the membrane and partially localized in the cytoplasm of HCT116/S322E and HCT116/WT cells. These observations suggested that Trop-2 phosphorylation plays a role in the decrease or mislocalization of claudin-7. Using protein kinase C (PKC) inhibitors and PKC-specific siRNAs, we found that PKCα and PKCδ are responsible for Trop-2 phosphorylation. Of note, chemical PKC inhibition and PKCα- and PKCδ-specific siRNAs reduced motility. In summary, our findings provide evidence that Trop-2 is phosphorylated at Ser-322 by PKCα/δ and that this phosphorylation enhances cell motility and decreases claudin-7 localization to cellular borders.

Induction of Trop-2 expression through the binding of galectin-3 to MUC1

Both mucin 1 (MUC1) and trophoblast cell surface antigen 2 (Trop-2) are overexpressed in various epithelial tumor cells, and their high expression is correlated with a poor prognosis. Both proteins were expressed in a human breast cancer cell line, MCF-7 cells, but neither was in a human colon cancer cell line, HCT116 cells. When MUC1 cDNA was introduced into HCT116 cells (HCT116/MUC1), expression of Trop-2 was induced. Reciprocally, treatment of MCF-7 cells with MUC1 siRNA reduced the level of Trop-2. Mithramycin A, an inhibitor of specificity protein 1 (Sp1) transcription factor, effectively inhibited the expression of Trop-2. Consistently, treatment with Sp1 siRNA reduced the expression of Trop-2. To reveal the relationship between MUC1 and Sp1, coimmunoprecipitation assays were performed. Sp1 was coimmunoprecipitated with MUC1 and the level of coimmunoprecipitated Sp1 increased in relation to the level of induced Trop-2. It is known that galectin-3 is an endogenous ligand of MUC1. Binding of galectin-3 to MUC1 elevated the recruitment of Sp1 to MUC1, and knockdown of galectin-3 reduced the level of Trop-2. These results suggest that the binding of galectin-3 to MUC1 enhances the recruitment of Sp1, leading to promotion of the transcription of Trop-2.

First-in-human phase 1 study of DS-1062a in patients with advanced solid tumors.

9051 Background: DS-1062a is a trophoblast cell-surface antigen 2 (TROP2)-targeting antibody drug conjugate. TROP2 is highly expressed in epithelial cancers, including non-small cell lung cancer (NSCLC). Overexpression of TROP2 may be associated with poor survival in some solid tumors. Preclinical studies showed promising antitumor activity of DS-1062a in mouse models with TROP2-positive tumors. Preliminary results are reported in the dose escalation part of this phase 1 study. Methods: This is an ongoing phase 1 study of DS-1062a in patients (pts) with advanced solid tumors in the US and Japan (NCT03401385). Adult (age ≥20 years in Japan or ≥18 years in US) pts with measurable disease per RECIST v1.1 and sufficient tumor tissue sample for TROP2 measurement were eligible. Pts were excluded if they had multiple primary malignancies or untreated brain metastases. Endpoints included safety, pharmacokinetics, and efficacy. Results: As of November 16, 2018, 22 pts with advanced NSCLC were treated with DS-1062a at doses of 0.27-mg/kg (n = 4), 0.5-mg/kg (n = 5), 1.0-mg/kg (n = 7), and 2.0-mg/kg (n = 6). Overall, mean (standard deviation) treatment duration and cumulative dose were 7.8 (4.1) weeks and 181.8 (141.4) mg, respectively, with a median of 2 (range 1–6) cycles initiated. The majority (n = 18; 81.8%) of pts had ≥1 treatment-emergent adverse event (TEAE). The most common TEAE, fatigue (n = 9), was the only grade ≥3 treatment-related TEAE (n = 1; 2.0-mg/kg). Serious TEAEs, reported in 5 pts in the 0.27-mg/kg (n = 2), 0.5-mg/kg (n = 2), and 2.0-mg/kg (n = 1) cohorts, were not treatment-related. TEAEs leading to discontinuation occurred in 1 pt in the 0.27-mg/kg cohort (pleural effusion; not treatment-related). One pt died due to progressive disease. Peak concentration (Cmax) and area under the curve from time 0 to last measurable concentration (AUClast) increases were generally dose proportional. Of 18 tumor evaluable pts, 1 had a partial response (PR; 1.0-mg/kg), and 8 had stable disease (SD). Conclusions: DS-1062a was well tolerated at doses up to 2.0-mg/kg. An observable PR and multiple pts with SD warrant further evaluation of DS-1062a. The maximum tolerated dose has not been reached, and this study is ongoing. Clinical trial information: NCT03401385.

Sacituzumab Govitecan-hziy in Refractory Metastatic Triple-Negative Breast Cancer

BackgroundStandard chemotherapy is associated with low response rates and short progression-free survival among patients with pretreated metastatic triple-negative breast cancer. Sacituzumab govitecan-hziy is an antibody–drug conjugate that combines a humanized monoclonal antibody, which targets the human trophoblast cell-surface antigen 2 (Trop-2), with SN-38, which is conjugated to the antibody by a cleavable linker. Sacituzumab govitecan-hziy enables delivery of high concentrations of SN-38 to tumors.MethodsWe conducted a phase 1/2 single-group, multicenter trial involving patients with advanced epithelial cancers who received sacituzumab govitecan-hziy intravenously on days 1 and 8 of each 21-day cycle until disease progression or unacceptable toxic effects. A total of 108 patients received sacituzumab govitecan-hziy at a dose of 10 mg per kilogram of body weight after receiving at least two previous anticancer therapies for metastatic triple-negative breast cancer. The end points included safety; the objective response rate (according to Response Evaluation Criteria in Solid Tumors, version 1.1), which was assessed locally; the duration of response; the clinical benefit rate (defined as a complete or partial response or stable disease for at least 6 months); progression-free survival; and overall survival. Post hoc analyses determined the response rate and duration, which were assessed by blinded independent central review.ResultsThe 108 patients with triple-negative breast cancer had received a median of 3 previous therapies (range, 2 to 10). Four deaths occurred during treatment; 3 patients (2.8%) discontinued treatment because of adverse events. Grade 3 or 4 adverse events (in ≥10% of the patients) included anemia and neutropenia; 10 patients (9.3%) had febrile neutropenia. The response rate (3 complete and 33 partial responses) was 33.3% (95% confidence interval [CI], 24.6 to 43.1), and the median duration of response was 7.7 months (95% CI, 4.9 to 10.8); as assessed by independent central review, these values were 34.3% and 9.1 months, respectively. The clinical benefit rate was 45.4%. Median progression-free survival was 5.5 months (95% CI, 4.1 to 6.3), and overall survival was 13.0 months (95% CI, 11.2 to 13.7).ConclusionsSacituzumab govitecan-hziy was associated with durable objective responses in patients with heavily pretreated metastatic triple-negative breast cancer. Myelotoxic effects were the main adverse reactions. (Funded by Immunomedics; IMMU-132-01 ClinicalTrials.gov number, NCT01631552.)

Silencing of the TROP2 gene suppresses proliferation and invasion of hepatocellular carcinoma HepG2 cells.

ObjectivesOverexpression of human trophoblast cell surface antigen 2 (Trop2) has been observed in many cancers; however, its roles in proliferation, apoptosis, migration, and invasion of hepatocellular carcinoma (HCC) remain unclear. Thus, this study aimed to characterize the function of Trop2 in HCC.MethodsTrop2 protein expression was detected by immunohistochemistry in HCC tissues. Cell proliferation, apoptosis, and invasion were respectively measured by CCK-8, flow cytometry, Transwell, and wound healing assays. Expression levels of epithelial–mesenchymal transition-related proteins and Trop2 protein in HCC cell lines were detected by western blotting after silencing of the TROP2 gene.ResultsTrop2 protein was highly expressed in HCC tissues and HCC cell lines. Trop2 mRNA and protein expression levels decreased in HepG2 and HCCLM3 cells after transfection with Trop2 siRNA. Silencing of the TROP2 gene in HepG2 and HCCLM3 cells strongly inhibited cell proliferation and migration, while enhancing cell apoptosis. Investigation of the molecular mechanism revealed that silencing of the TROP2 gene suppressed epithelial–mesenchymal transition of HepG2 and HCCLM3 cells.ConclusionsThe results of the present study may improve understanding of the role of Trop2 in regulation of cell proliferation and invasion, and may aid in development of novel therapy for HCC.

The diagnostic significance of trophoblast cell-surface antigen-2 expression in benign and malignant thyroid lesions.

Thyroid cancers are the most common malignancy of the endocrine system. Over-expression of trophoblast cell-surface antigen 2 (TROP-2) in various tumors has been found to correlate with poor prognosis and aggressive tumor behavior. The aim of this study was to evaluateTROP-2 expression in thyroid neoplasms. This study contained 152 cases, including 48 follicular nodular disease (FND), 29 follicular adenoma (FA), 57 papillary thyroid carcinoma (PTC), 12 follicular thyroid carcinoma (FTC), 3 medullary thyroid carcinoma (MTC), 2 poorly differentiated thyroid carcinoma (PDTC) and 1 undifferentiated thyroid carcinoma (UDTC). TROP-2 expression was investigated via immunohistochemistry in sections prepared from paraffin blocks of the cases. The cases comprised 32 (21%) males and 120 (79%) females with a mean age of 46.8 years (range, 15-85 years). TROP-2 expression was observed in 74.6% of the malignant lesions of the thyroid except for medullary carcinoma, poorly differentiated and undifferentiated thyroid carcinoma. Immunoreactivity was 3.4% in FA, 41.7% of cases with FTC and 81.8% in PTC follicular variant (PTC fv). The difference between FA/FTC and FA/PTC follicular variant were both significant (P < 0.005, P < 0.001, respectively). There was no difference between FTC/PTC fv (P = 0.089). TROP-2 can be considered a useful marker for distinguishing PTC fv cases from follicular nodular disease and follicular adenoma cases because of its high sensitivity in the identification of papillary carcinomas of the thyroid.

Sacituzumab govitecan: antibody-drug conjugate in triple-negative breast cancer and other solid tumors.

Patients with metastatic triple-negative breast cancer (mTNBC) that has progressed on first-line therapy have a poor prognosis with limited therapeutic options. Sacituzumab govitecan (SG) is a novel antibody-drug conjugate (ADC) that has shown promising efficacy in mTNBC. SG is comprised of SN-38, the active metabolite of irinotecan, conjugated via a hydrolyzable linker to the humanized RS7 antibody targeting trophoblast cell surface antigen 2 (Trop-2), a glycoprotein that is expressed at high levels in many epithelial solid tumors. It has received breakthrough therapy status by the U.S. Food and Drug Administration (FDA) for the treatment of patients with pretreated mTNBC. In this review, we summarize available data regarding the pharmacology, pharmacokinetics, safety and efficacy of SG and describe ongoing and future clinical studies investigating this agent.

TROP2 promotes the proliferation and metastasis of glioblastoma cells by activating the JAK2/STAT3 signaling pathway.

Trophoblast cell surface antigen 2 (TROP2), a single transmembrane domain protein, is often found to be highly expressed in various types of human cancers. However, the biological function and molecular mechanism of TROP2 in glioblastoma have not been fully elucidated, particularly in regards to cell proliferation and metastasis of glioblastoma cells. In the present study, it was demonstrated that TROP2 expression was increased in glioblastoma tissues and glioblastoma cell lines by immunohistochemical analysis and western blot analysis. High TROP2 expression was significantly correlated with the poor survival of glioblastoma patients. MTT assay, BrdU incorporation assay, flow cytometry and Transwell assay were performed to demonstrate that knockdown of TROP2 in glioblastoma cells inhibited cell proliferation and metastasis. We found that the effects of TROP2-knockdown on glioblastoma cells were associated with the inhibition of JAK2 and STAT3 phosphorylation and decreased transcription of STAT3 target genes. In addition, blocking the activation of JAK2/STAT3 signaling by WP1066 negated the effects of TROP2 overexpression. Furthermore, exogenous IL-6, which functions as a potent activator of JAK2/STAT3 signaling, was able to rescue the phosphorylation of JAK2 and STAT3 in TROP2-silenced glioblastoma cells and regulate phenotypic changes in these cells. Therefore, we revealed a novel mechanism by which TROP2 activates the JAK2/STAT3 pathway to promote the growth and metastasis of glioblastoma cells. These data offer insight into the function of TROP2 in glioblastoma and indicate that TROP2 is a promising biomarker and therapeutic target for glioblastoma patients.

Down-regulation of TROP-2 Predicts Poor Prognosis of Hepatocellular Carcinoma Patients.

Hepatocellular carcinoma (HCC) is one of the most common and lethal cancer types worldwide, especially in Asian countries. Genetic alterations, including hyperactivation of oncogenes and loss of expression of tumor suppressor genes, greatly contribute to the initiation and progression of HCC. Here we report that down‐regulation of trophoblast cell surface antigen 2 (TROP‐2) was frequently detected in HCC. Transcriptome sequencing of non‐tumor and HCC patient samples revealed down‐regulation of TROP‐2 in tumor tissues. Immunohistochemical staining showed nearly undetectable levels of TROP‐2 in HCC tissues but distinct and strong staining of TROP‐2 in adjacent non‐tumor tissues. The frequent down‐regulation of TROP‐2 expression was further confirmed in an in‐house cohort of 205 pairs of HCC patient samples and in the Cancer Genome Atlas (TCGA) databases. Furthermore, the down‐regulation of TROP‐2 was associated with poor overall survival of HCC patients, severe adjacent organ invasion, and poor differentiation of HCC. Using bisulfite genomic sequencing and methylation‐specific polymerase chain reaction analyses, we show that higher levels of promoter methylation were detected in the DNA samples of HCC tissues (low TROP‐2 expression) than that of the non‐tumor tissues (high TROP‐2 expression). Conclusion: Taken together, our data suggest that promoter hypermethylation contributes to the frequent down‐regulation of TROP‐2 in HCC, and that TROP‐2 down‐regulation predicts poor prognosis of HCC patients.

Effect of human trophoblast cell-surface antigen 2 gene expression by RNA interference on proliferation and apoptosis of tongue squamous cell carcinoma and its mechanism

Objective: To investigate the effect and its mechanism of human trophoblast cell-surface antigen 2 (Trop2) gene expression was inhibited in squamous cell carcinoma of tongue on the proliferation and apoptosis of cancer cells. Methods: A total of 46 patients from February 2014 to May 2016 received radical treatment of tongue cancer from oral and maxillofacial surgery of the First Affiliated Hospital of Zhengzhou University were enrolled in this study. Real time PCR and Western blotting were used to detect mRNA and protein expression of Trop2 in tongue squamous cell carcinoma and corresponding adjacent tissues; NC-siRNA and Trop2-siRNA were transfected into human tongue squamous cell carcinoma CAL-27 cells, a blank control group (control) was set, the expression of Trop2, Ki-67, cyclin D1, cleaved caspase3, Notch1, Hes1 protein after transfected for 48 h were detected by Western bloting; cell proliferation was detected by cell counting kit-8; cell cycle and apoptosis rate were detected by flow cytometry. Results: The mRNA (5.72±1.13) and protein expression (0.77±0.06) of Trop2 gene in tongue squamous cell carcinoma were significantly higher than those in adjacent tissues (0.92±0.15, 0.11±0.01, P<0.05); Trop2 protein expression after transfeced Trop2-siRNA (0.08±0.01) in CAL-27 cells decreased significantly (0.46±0.05); the survival rate (64.28±4.12)%, S cells (14.54±1.02)% and Ki-67 (0.12±0.01), cyclin D1 (0.04±0.01) protein expression in Trop2-siRNA group were significantly lower than those in control group [(100.00±1.02)%, (27.33±1.11)%, (0.24±0.02), (0.20±0.02)] and NC-siRNA group [(96.55±2.43)%, (26.67±1.23)%, (0.26±0.03), (0.21±0.024)], the apoptosis rate (23.55±1.45)%, G0/G1 cells (72.32±0.94)% and the expression of cleaved caspase 3 (0.16±0.02), Notch1 (0.62±0.06) and Hes1 (0.50±0.05) protein were significantly higher than control group and NC-siRNA group (P<0.05). Conclusions: The expression of Trop2 gene was inhibited in tongue squamous carcinoma cells can reduce the proliferation of cancer cells, block the cells in phase G1, and promote the apoptosis of cells. The mechanism is related to the up regulation of Notch1 signaling pathway.