Sacituzumab govitecan plus platinum-based chemotherapy mediates significant antitumor effects in triple-negative breast, urinary bladder, and small-cell lung carcinomas

Abstract

Sacituzumab govitecan (SG) is an antibody-drug conjugate composed of an anti-Trop-2-directed antibody conjugated with the topoisomerase I inhibitory drug, SN-38, via a proprietary hydrolysable linker. SG has received United States Food and Drug Administration (FDA) approval to treat metastatic triple-negative breast cancer (TNBC), unresectable locally advanced or metastatic hormone receptor (HR)-positive, human epidermal growth factor receptor 2 (HER2)-negative breast cancer, and accelerated approval for metastatic urothelial cancer. We investigated the utility of combining SG with platinum-based chemotherapeutics in TNBC, urinary bladder carcinoma (UBC), and small-cell lung carcinoma (SCLC). SG plus carboplatin or cisplatin produced additive growth-inhibitory effects in vitro that trended towards synergy. Immunoblot analysis of cell lysates suggests perturbation of the cell-cycle and a shift towards pro-apoptotic signaling evidenced by an increased Bax to Bcl-2 ratio and down-regulation of two anti-apoptotic proteins, Mcl-1 and survivin. Significant antitumor effects were observed with SG plus carboplatin in mice bearing TNBC or SCLC tumors compared to all controls (P < 0.0062 and P < 0.0017, respectively) and with SG plus cisplatin in UBC and SCLC tumor-bearing animals (P < 0.0362 and P < 0.0001, respectively). These combinations were well tolerated by the animals. Combining SG with platinum-based chemotherapeutics demonstrates the benefit in these indications and warrants further clinical investigation.

Keywords: SN-38; Trop-2; carboplatin; cisplatin; sacituzumab govitecan.

Figure 1. Isobolograms demonstrating changes in the in vitro growth-inhibitory effects of SG and platinum-based chemotherapeutics when combined in TNBC, UBC, and SCLC tumor lines.

Cells were co-cultured with SG plus carboplatin or cisplatin as described in Materials and Methods. After 96 h, changes in growth inhibition were determined for SG or a given chemotherapeutic when each was incubated with a constant amount of drug (i.e., SG dose-response in constant amounts of a chemotherapeutic and vice versa). Isobolograms were graphed on data normalized to IC₅₀-values for each individual drug (i.e., SG or chemotherapeutic). SG was combined with (A) carboplatin or (B) cisplatin. Each assay was performed at least three times for each condition (i.e., 3 assays of SG plus constant chemotherapeutic and 3 assays of chemotherapeutic in constant SG). (□) Effect on IC₅₀ of the chemotherapeutic when incubated with a constant amount of SG. (●) Effect on IC₅₀ of SG when incubated with constant amounts of a chemotherapeutic. Dotted line indicates additive effect of the combination. Area below and above the dotted line represent synergistic and antagonistic interactions, respectively.

Figure 2. Immunoblot assessment of effects on cell-cycle, pro- and anti-apoptosis signaling events mediated by SG plus platinum-based chemotherapeutics in human TNBC, SCLC, and UBC tumor-lines.

Cells were plated overnight in 6-well plates before the addition of SG and chemotherapeutics, either alone or in combination. SG concentrations are shown as SN-38 equivalents based on the protein concentration and DAR of the ADC. After a 24-h incubation, cells were harvested, and cell lysates resolved and transferred for immunoblot analysis as described in Materials and Methods. Bcl-2 and Bax levels were determined on the same blot. (A) Human HCC1806 TNBC cells and (B) DMS 53 SCLC cells exposed to carboplatin (50 and 100 μM) and SG (10 and 100 nM SN-38 equivalents). For HCC1806, β-actin loading control is the same for both Cyclin D1 and survivin blots due to stripping and re-probing the same blot. For clarity, it is reproduced under both blots. Likewise, for DMS 53, the loading control is same for p21 and Cyclin D1. Bcl-2/Bax and survivin also share the same loading control and for clarity, the same β-actin control is reproduced under Bcl-2/Bax and survivin blots. (C) 5637 and (D) RT4 human UBC human tumor lines incubated with cisplatin (0.2 and 2 μM) and SG (10 and 100 nM SN-38 equivalents). 5637 immunoblot was stripped and re-probed and therefore the β-actin loading control is the same for p21, Bcl-2/Bax, Mcl-1 and survivin. For clarity, the β-actin blot is reproduced under the p21 and survivin blots. Likewise, for RT4 p21 and Cyclin D1 share the same β-actin loading control. Bcl-2/Bax, Mcl-1, and survivin also share β-actin loading control. Each assay was performed under the same conditions at least twice.

Figure 3. In vivo efficacy of SG combined with carboplatin or cisplatin in mice bearing human TNBC, SCLC, or UBC xenografts.

Animals were set up with the various tumor xenografts as described in Materials and Methods. Mice were treated with SG i.v., carboplatin i.p., cisplatin i.p., control ADC i.v. (h679-CL2A-SN-38), or combinations at the indicated doses. Brackets in all the figures represent AUC P-values between the two indicated treatment comparisons for a given set of groups. (A, B) Tumor growth curves of mice bearing HCC1806 human TNBC xenografts treated with the combination of SG plus carboplatin. Both SG and carboplatin were administered once weekly for four weeks (red arrows). (A) Mice administered 500 μg SG alone or in combination with carboplatin. (^* P = 0.0006, ^** P < 0.0001, and ^*** P = 0.0062). (B) Mice administered 250 μg SG alone or in combination with carboplatin. These mice were in the same study as in (A) and thus share the same control groups. (^* P = 0.0047 and ^** P = 0.0363). (C) DMS 53 tumor growth curves for mice treated with SG plus carboplatin. SG alone groups received twice weekly injections (blue arrows) while in the combination group, SG was administered on the same schedule as the carboplatin (i.e., weekly x 4 wks; orange arrows). (^* P < 0.0001 and ^** P = 0.0017). (D) Likewise, DMS 53 tumor-bearing mice treated with SG plus cisplatin weekly for four weeks (purple arrows). These mice were in the same study as in (C) and therefore share the same saline and SG monotherapy control groups. (^* P < 0.0001 and ^** P < 0.0001). (E) Mice bearing 5637 human UBC tumors and treated with the combination of SG plus cisplatin weekly for four weeks (red arrows). (^* P = 0.0362, ^** P = 0.0007, and ^*** P = 0.0030). (F) Survival curves for those mice bearing the 5637 tumors and treated with SG plus cisplatin. Dotted grey line denotes the 50% survival threshold. Log-rank analysis was utilized to calculate the P-values. Abbreviations: N.A.: Not Applicable. SD: Standard Deviation.