Inhibitor SN-38 Research Articles

PLX038A, a long-acting SN-38, penetrates the blood-tumor-brain-barrier, accumulates and releases SN-38 in brain tumors to increase survival of tumor bearing mice

Central nervous system tumors have resisted effective chemotherapy because most therapeutics do not penetrate the blood-tumor-brain-barrier. Nanomedicines between ~ 10 and 100 nm accumulate in many solid tumors by the enhanced permeability and retention effect, but it is controversial whether the effect can be exploited for treatment of brain tumors. PLX038A is a long-acting prodrug of the topoisomerase 1 inhibitor SN-38. It is composed of a 15 nm 4-arm 40 kDa PEG tethered to four SN-38 moieties by linkers that slowly cleave to release the SN-38. The prodrug was remarkably effective at suppressing growth of intracranial breast cancer and glioblastoma (GBM), significantly increasing the life span of mice harboring them. We addressed the important issue of whether the prodrug releases SN-38 systemically and then penetrates the brain to exert anti-tumor effects, or whether it directly penetrates the blood-tumor-brain-barrier and releases the SN-38 cargo within the tumor. We argue that the amount of SN-38 formed systemically is insufficient to inhibit the tumors, and show by PET imaging that a close surrogate of the 40 kDa PEG carrier in PLX038A accumulates and is retained in the GBM. We conclude that the prodrug penetrates the blood-tumor-brain-barrier, accumulates in the tumor microenvironment and releases its SN-38 cargo from within. Based on our results, we pose the provocative question as to whether the 40 kDa nanomolecule PEG carrier might serve as a “Trojan horse” to carry other drugs past the blood-tumor-brain-barrier and release them into brain tumors.

Targeting fatty acid synthase in preclinical models of TNBC brain metastases synergizes with SN-38 and impairs invasion

Fatty acid synthesis (FAS) has been shown to play a key role in the survival of brain-metastatic (BM) breast cancer. We demonstrate that the fatty acid synthase inhibitor TVB-2640 synergizes with the topoisomerase inhibitor SN-38 in triple-negative breast cancer (TNBC) BM cell lines, upregulates FAS and downregulates cell cycle progression gene expression, and slows the motility of TNBC BM cell lines. The combination of SN-38 and TVB-2640 warrants further consideration as a potential therapeutic option in TNBC BMs.

Dual-targeted nanoparticulate drug delivery systems for enhancing triple-negative breast cancer treatment

The efficacy of DNA-damaging agents, such as the topoisomerase I inhibitor SN38, is often compromised by the robust DNA repair mechanisms in tumor cells, notably homologous recombination (HR) repair. Addressing this challenge, we introduce a novel nano-strategy utilizing binary tumor-killing mechanisms to enhance the therapeutic impact of DNA damage and mitochondrial dysfunction in cancer treatment. Our approach employs a synergistic drug pair comprising SN38 and the BET inhibitor JQ-1. We synthesized two prodrugs by conjugating linoleic acid (LA) to SN38 and JQ-1 via a cinnamaldehyde thioacetal (CT) bond, facilitating co-delivery. These prodrugs co-assemble into a nanostructure, referred to as SJNP, in an optimal synergistic ratio. SJNP was validated for its efficacy at both the cellular and tissue levels, where it primarily disrupts the transcription factor protein BRD4. This disruption leads to downregulation of BRCA1 and RAD51, impairing the HR process and exacerbating DNA damage. Additionally, SJNP releases cinnamaldehyde (CA) upon CT linkage cleavage, elevating intracellular ROS levels in a self-amplifying manner and inducing ROS-mediated mitochondrial dysfunction. Our results indicate that SJNP effectively targets murine triple-negative breast cancer (TNBC) with minimal adverse toxicity, showcasing its potential as a formidable opponent in the fight against cancer.

ESG401, a trophoblast cell-surface antigen 2 (TROP2) antibody drug conjugate (ADC), for the treatment of first-line metastatic triple negative breast cancer (mTNBC).

e13132 Background: ESG401, a novel ADC comprising a humanized anti-TROP2 IgG1 monoclonal antibody conjugated to the Topoisomerase I Inhibitor SN-38, with the Drug-Antibody Ratio at 8, by a stable cleavable linker, exhibited preliminary efficacy and tolerability in the Phase Ia trial (Ma, ASCO 2023). This report summarizes current results from the first-line mTNBC cohort of the Phase Ib trial. Methods: Patients (pts) aged ≥18 years with confirmed local advanced/unresectable or metastatic TNBC, without prior metastatic treatment, received ESG401 (16 mg/kg IV on Day 1,8, and 15, with a 28-day cycle) until unacceptable toxicity, progressive disease, or consent withdrawal. Results: As of the cutoff date (Jan 30th, 2024), 23 pts in the cohort received ≥1 dose of ESG401. Median age was 52 years (range: 33-73), 34.8% were ECOG 0, 26.1% were de novo stage IV, 73.9% had visceral disease (8.7% brain, 34.8% liver, 52.2% lung), 43.5% received radiotherapy, and 73.9% received chemotherapy (73.9% Taxanes, 69.6% Anthracycline, 4.3% Platinum)as prior treatments for early-stage disease. Among 14 Efficacy Evaluable (EE) pts, the ORR was 78.6%, including one (7.1%) patient who achieved a complete response. DCR was 100%, and CBR was 85.7%. Eight pts (57.1%) achieved confirmed Partial Response (PR). As of the cutoff date, 14 pts (63.6%) remained on treatment, with the longest on-treatment duration being 8.0 mons. Two (2) pts in this cohort had brain metastases. One pt achieved a complete intracranial response, and the overall response evaluation was PR. The evaluation time for the other pt has not yet been reached. Across the entire Phase Ia/1b trials, including 15 EE pts with brain metastases, the best overall intracranial response rate was 33%, with an intracranial disease control rate of 73%. The response of the intracranial tumor lesion is correlated with the response of the non-CNS lesion. The safety profile of ESG401 remained consistent with previous reports, showing no new or unexpected safety signals. The most common TRAEs were leukopenia (65.2%), neutropenia (69.6%), anemia (43.5%), fatigue (21.7%), nausea (43.5%), and vomiting (34.8%). The most common TRAEs≥ Grade 3 were neutropenia (43.5%). There was no ≥ Grade 3 diarrhea. No interstitial lung disease (ILD) has been reported so far for the entire PhIa/b trials containing 139 pts. Conclusions: ESG401 demonstrates promising antitumor activity in the first-line mTNBC pts, with efficacy comparable to the combination therapy reported by the BEGONIA Study. Additionally, the efficacy of ESG401 in brain metas in the first-line mTNBC pts is in alignment with our previous observations made in the late-line mTNBC and HR+/HER2- BC pts. These results warrant further clinical investigation of ESG401. Clinical trial information: NCT04892342 .

Phase II clinical trial of PLX038 in patients with platinum resistant ovarian, primary peritoneal and fallopian tube cancer.

TPS5630 Background: Platinum-resistant ovarian cancer (PROC) portends a poor prognosis and, although treatments are available, response rates are low. Previous studies have demonstrated that DNA topoisomerase I (TOP1) inhibitors exhibit clinical activity in PROC. Irinotecan also has a 15-25% response rate in some studies of platinum-resistant epithelial ovarian cancer but does not have FDA approval for this indication. Additional studies in preclinical models suggest that TOP1 poisons are more active if administered at low doses on prolonged schedules. PLX038 is a long-acting prodrug of the potent TOP1 inhibitor SN-38. In PLX038, SN-38 is covalently conjugated to circulating polyethylene glycol and is slowly released at a constant rate to provide the active agent with a long half-life, a low Cmax and very high exposure – important facets for optimal safety and efficacy of this drug. Further, in preclinical studies the nanoparticle prodrug accumulates and is retained in tumors, where it slowly releases the SN-38. Methods: This single arm, phase 2 clinical trial is open at Mayo Clinic in Rochester, MN. The primary endpoint is overall response rate and secondary endpoints include progression-free survival and assessment of tolerability of PLX038. Correlative goals include i) measurement of TOP1-DNA covalent complexes in tumor biopsies and circulating tumor cells to assess successful target engagement; ii) assessment of homologous recombination status and SLFN11 expression for correlation with tumor response rate; and iii) pharmacokinetics of SN-38 and SN-38G as well as their association with GI toxicity. Assessment of gut microbiota and association with GI toxicity profile are also being investigated. Eligible patients include women with platinum resistant high grade serous ovarian, primary peritoneal, and fallopian tube cancer who have received fewer than two prior chemotherapy regimens in the platinum-resistant setting. All patients must have an ECOG performance status of 0, 1, or 2 and adequate bone marrow, renal, and hepatic function. As of January 31, 2024, 12 of planned 37 patients have enrolled. Clinical trial information: NCT05465941 .

Antibody-Drug Conjugate Sacituzumab Govitecan Enables a Sequential TOP1/PARP Inhibitor Therapy Strategy in Patients with Breast Cancer.

The antibody-drug conjugate (ADC) sacituzumab govitecan (SG) comprises the topoisomerase 1 (TOP1) inhibitor (TOP1i) SN-38, coupled to a monoclonal antibody targeting trophoblast cell surface antigen 2 (TROP-2). Poly(ADP-ribose) polymerase (PARP) inhibition may synergize with TOP1i and SG, but previous studies combining systemic PARP and TOP1 inhibitors failed due to dose-limiting myelosuppression. Here, we assess the proof-of-mechanism and clinical feasibility for SG and talazoparib (TZP) employing an innovative sequential dosing schedule. In vitro models tested pharmacodynamic endpoints, and in a phase 1b clinical trial (NCT04039230), 30 patients with metastatic triple-negative breast cancer (mTNBC) received SG and TZP in a concurrent (N = 7) or sequential (N = 23) schedule. Outcome measures included safety, tolerability, preliminary efficacy, and establishment of a recommended phase 2 dose. We hypothesized that tumor-selective delivery of TOP1i via SG would reduce nontumor toxicity and create a temporal window, enabling sequential dosing of SG and PARP inhibition. In vitro, sequential SG followed by TZP delayed TOP1 cleavage complex clearance, increased DNA damage, and promoted apoptosis. In the clinical trial, sequential SG/TZP successfully met primary objectives and demonstrated median progression-free survival (PFS) of 7.6 months without dose-limiting toxicities (DLT), while concurrent dosing yielded 2.3 months PFS and multiple DLTs including severe myelosuppression. While SG dosed concurrently with TZP is not tolerated clinically due to an insufficient therapeutic window, sequential dosing of SG followed by TZP proved a viable strategy. These findings support further clinical development of the combination and suggest that ADC-based therapy may facilitate novel, mechanism-based dosing strategies.

Abstract PO2-19-03: Phase 2 study of response-guided neoadjuvant sacituzumab govitecan in patients with localized breast cancer (NeoSTAR)

Abstract Background: Optimizing treatments for localized breast cancer is key to preventing metastatic recurrences and reducing mortality. Sacituzumab Govitecan (SG) is a novel antibody-drug conjugate in which the topoisomerase 1 inhibitor SN-38 (the active metabolite of irinotecan) is coupled to a humanized monoclonal antibody targeting the antigen Trop-2, which is highly expressed in triple negative breast cancer (TNBC). SG is FDA approved for patients with metastatic hormone receptor-positive (HR+) and TNBC. However, efficacy in localized BC is not known. The NeoSTAR clinical trial is evaluating SG in the neoadjuvant (NA) setting for participants with localized breast cancer. Cohort A1 evaluating NA SG monotherapy in localized TNBC is completed, and now cohorts A2 (SG + pembrolizumab (P) for TNBC), B1 (SG monotherapy for HR+ BC), and C (SG + P for HER2- inflammatory breast cancer (IBC)) are enrolling. Trial design: This is a single-arm phase II study of NA SG or SG + P with multiple cohorts across different breast cancer subtypes. In all cohorts, SG is administered via an IV infusion on days 1 and 8 of each 21-day cycle at a starting dose of 10 mg/kg for 4 cycles. For cohorts A2 and C, pembrolizumab is given on day 1 of each cycle. For A2 and B1, after 4 cycles, participants who have biopsy-proven residual disease will have the option to receive additional standard NA therapy at the discretion of the treating physician and subsequently proceed to surgery. Those with a complete response on imaging may proceed directly to surgery. If pathological complete response (pCR) is achieved after 4 cycles of SG + P, participants on A2 will remain on study and receive adjuvant weekly paclitaxel and carboplatin for 12 weeks along with continuation of P to complete a total of one year of the PD-1 inhibitor; participants on B1 will receive adjuvant therapy at the discretion of the treating physician, with the option to continue P to complete a total of one year. On cohort C, after 4 cycles of SG + P, dose-dense doxorubicin + cyclophosphamide + P for 4 cycles will be given before considering surgery. A baseline research biopsy is required for all cohorts, as well as tissue collection following treatment with SG (either at surgery or via biopsy prior to additional NA therapy). Eligibility criteria: Participants ≥ 18 years of age with previously untreated localized breast cancer as determined by the local institution will be enrolled. For A2, participants must either have a primary tumor >2 cm or be node positive. For B1, participants with anatomic clinical stage II-III breast cancer with primary tumor > 1.5 cm and high genomic risk are eligible. For the IBC cohort (C), participants with T4d and any N are eligible. An ECOG performance score of 0 or 1, and adequate bone marrow, hepatic, and renal function is required. Study Objectives: The primary objective is to assess the pCR rate in breast and lymph nodes (ypT0/isN0) with SG + P (A2), SG (B1), or SG + P followed by AC + P (C). Secondary objectives include assessment of overall response rate, evaluation of the safety and tolerability of SG or SG + P (CTCAE v5.0), event-free survival (EFS), and quality of life (EORTC QLQ-C30). Exploratory objectives include assessment of potential predictive biomarkers, including Trop-2 expression, DNA damage response markers and immunological markers, as well as changes in cell free DNA with SG. Statistical methods: The primary analysis is based on the estimated pCR rate with SG or SG + P and will be provided as a proportion (with two-sided 95% confidence interval). Each cohort will follow a Simon two-stage design. EFS will be analyzed using Kaplan-Meier methods and descriptive statistics. Target Accrual: 50 participants (A2), 56 participants (B1), 39 participants (C) Contact: Dr. Laura Spring (LSpring@mgh.harvard.edu) Clinicaltrials.gov #: NCT04230109 Citation Format: Laura Spring, Ana Garrido-Castro, Filipa Lynce, Nadine Tung, Rachel Abelman, Lianne Ryan, Megan Moran, Elizabeth Mittendorf, Leif Ellisen, Sara Tolaney, Aditya Bardia. Phase 2 study of response-guided neoadjuvant sacituzumab govitecan in patients with localized breast cancer (NeoSTAR) [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-19-03.

Abstract CT166: First-in-human phase 1 study of a phosphatidylserine-targeting small molecule drug conjugate (T-1201) in patients with advanced solid tumors

Abstract Background Phosphatidylserine (PS), a phospholipid, is an essential component of bilayer cell membranes and is normally present in the inner cell membrane layer of the cells. In rapid proliferating cancer cells, PS externalizes to the outer cell membrane and has the potential to act as a tumor biomarker and therapeutic target. T-1201 is a small molecule drug conjugate comprised of a phosphatidylserine-targeting head, Zinc (II) bis-dipicolylamine (Zn-DPA), linked to bioactive topoisomerase I inhibitor SN-38. Once T-1201 via Zn-DPA targets tumor cells, then enzymatic cleavage releases the cytotoxic payload of SN38, the active metabolite of irinotecan. This leads to high accumulation of antineoplastic drug directly in the tumor microenvironment. Here, we describe a trial in progress of a first-in-human trial of T-1201 in patients with advanced solid tumors. Methods This first-in-human, open-label, multicenter, phase I trial (NCT04866641) is investigating the safety, tolerability, pharmacokinetics and antitumor activity of T-1201 administrated as monotherapy to subjects with advanced solid tumors. Eligible patients have ≥ 1 measurable lesion per RECIST v1.1. and no effective standard therapy exists. T-1201 is administered intravenously once every 4 weeks (Q4W) in 28-day cycles with dose escalation guided by an accelerated titration design. The study started with 1 subject enrolled per cohort. With 100% dose increment for the next cohort. If 1 subject experiences any ≥ Grade 2 AE or DLT at any dose level during first cycle, the dose escalation will transit to 3+3 design where dose increment will follow a modified Fibonacci sequence. Starting with Cycle 2, the dosing interval can be adjusted to once every two weeks (Q2W) and the dose level after switching to Q2W schedule will be half of that at Q4W. Intra-subject dose escalation is permitted to maximize the likelihood of responses while keeping toxicity acceptable for individual subjects. The dose can be increased after subjects complete at least 2 treatment cycles at a particular dose level without experiencing any > grade 2 drug-related toxicity. The primary endpoints are the maximum tolerated dose (MTD), recommended Phase 2 dose (RP2D) and adverse event rate as measured by CTCAE v5.0. Secondary endpoints include objective response rate (ORR), clinical benefit rate (CBR), duration of response (DOR) and time to tumor progression. This study has begun enrolling patients and is continuing as planned in Taiwan. 77% of planned patients have been enrolled as of November 2023. Citation Format: Hui-Ching Wang, Yu-Min Yeh, Jeng-Shiun Du, Tsung-Jang Yeh, Jui-Hung Tsai, Yu-Chin Gau, Shang Yin Wu, Li-Tzong Chen, Hsiao-Fang Li, Wu-Chou Su. First-in-human phase 1 study of a phosphatidylserine-targeting small molecule drug conjugate (T-1201) in patients with advanced solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT166.

Abstract 3130: In vitro characterization of a novel TROP2-targeting antibody-drug conjugate OBI-992

Abstract BACKGROUND TROP2, a transmembrane glycoprotein widely expressed on epithelial cancers, has emerged as an attractive target for antibody-drug conjugate (ADC) development. Sacituzumab govitecan, a TROP2 ADC conjugated to topoisomerase inhibitor (TOP1) SN-38, has been approved for patients with triple-negative breast cancer. This study aims to investigate the characteristics of a novel TROP2 antibody (R4702) and its ADC OBI-992, which is derived from the conjugation of R4702 with a TOP1 inhibitor exatecan. METHODS Clinical study: Prognostic roles of TROP2 and TOP1 in patients were accessed through the “Kaplan-Meier plotter” database from more than 1055 colon cancer patients. In vitro study: An ELISA study was conducted to investigate the binding epitope of R4702. Cell binding assay and Surface Plasmon Resonance were carried out to measure non-specific binding activity and binding affinity, respectively. DNA damage activity, cytotoxicity, and immunogenic cell death (ICD) effects of three TOP1 inhibitors exatecan, deruxtecan (Dxd), and SN-38, were evaluated. Cytotoxicity of OBI-992 along with other ADC was measured in both 2D and 3D spheroids cancer cells (gastric cancer NCI-N87, pancreatic cancer Capan1, and prostate cancer DU145). TROP2 and BCRP expression as well as potential mutations in TOP1 were investigated in TROP2 ADCs-induced resistant colon cancer cells. The synergistic effects of the combination of OBI-992 with PARP inhibitors were evaluated. RESULTS Clinical study: Positive correlations between high mRNA expressions of both TROP2 (p=0.0075) and TOP1 (p=0.043) with poor overall survival for colon cancer patients were found. In vitro: The R4702 antibody displayed different binding epitopes from sacituzumab and datopotamab. In addition, R4702 showed lower non-specific binding and better binding affinity to TROP2 antigen than datopotamab. The payload exatecan demonstrated excellent ICD effects. Relatively higher cytotoxicity was observed by exatecan when compared to that of Dxd and SN-38. OBI-992 exhibited excellent activities on cell growth inhibition in 2D cell cultures and 3D cell spheroids. TROP2 down-regulation and BCRP upregulation were not found in OBI-992-induced resistant cells. A mutation R364H was identified in TOP1 in the resistance study with OBI-992, which is critical for the binding to TOP1 inhibitors camptothecin analogs. Finally, a synergistic effect of OBI-992 and PARP inhibitors (talazoparib, rucaparib, niraparib, and olaparib) on the enhancement of cancer cell killing and DNA damage was observed. CONCLUSION High expression of both TROP2 and TOP1 is associated with poor survival in patients with colon cancer. R4702 is a novel TROP2 antibody with a unique binding epitope and low non-specific binding. Exatecan and exatecan-conjugated ADC OBI-992 showed high cytotoxic activities. Combination of OBI-992 with PARP inhibitors have potential in TROP2-expressing malignancies. Citation Format: Tzer-Min Kuo, Ting-Yu Chang, Jye-Yu Huang, Wei-Chien Tang, Chun-Jung Lin, Yi-Chen Wu, Chi-Huan Lu, Hao-Cheng Weng, Yu-Jung Chen, Yu-Hsuan Tsao, Cheng-Yen Wei, Lifen Shen, Wan-Fen Li, Ming-Tain Lai. In vitro characterization of a novel TROP2-targeting antibody-drug conjugate OBI-992 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3130.

SN-38, an active metabolite of irinotecan, inhibits transcription of nuclear factor erythroid 2-related factor 2 and enhances drug sensitivity of colorectal cancer cells.

Nuclear factor erythroid 2-related factor 2 (Nrf2) significantly contributes to drug resistance of cancer cells, and Nrf2 inhibitors have been vigorously pursued. Repurposing of existing drugs, especially anticancer drugs, is a straightforward and promising strategy to find clinically available Nrf2 inhibitors and effective drug combinations. Topoisomerase inhibitors SN-38 (an active metabolite of irinotecan), topotecan, mitoxantrone, and epirubicin were found to significantly suppress Nrf2 transcriptional activity in cancer cells. SN-38, the most potent one among them, significantly inhibited the transcription of Nrf2, as indicated by decreased mRNA level and binding of RNA polymerase II to NFE2L2 gene, while no impact on Nrf2 protein or mRNA degradation was observed. SN-38 synergized with Nrf2-sensitive anticancer drugs such as mitomycin C in killing colorectal cancer cells, and irinotecan and mitomycin C synergistically inhibited the growth of SW480 xenografts in nude mice. Our study identified SN-38 and three other topoisomerase inhibitors as Nrf2 inhibitors, revealed the Nrf2-inhibitory mechanism of SN-38, and indicate that clinically feasible drug combinations could be designed based on their interactions with Nrf2 signaling.

Preliminary results from a first-in-human study of ESG401, a trophoblast cell-surface antigen 2 (TROP2) antibody drug conjugate (ADC), in patients with locally advanced/metastatic solid tumors.

1100 Background: ESG401 is an innovative ADC composed of a humanized anti-TROP2 IgG1 monoclonal antibody conjugated to topoisomerase I inhibitor SN-38 via a proprietary stable-cleavable linker with DAR of 8. Methods: Patients (pts) aged ≥18 years with locally advanced/metastatic solid tumors refractory to/relapsed from standard treatment with measurable disease (RECIST v1.1) were eligible. ESG401 is administered intravenously by designated dose and regimen until unacceptable toxicity or progressive disease. The Bayesian Optimal Interval (BOIN) design was used to establish the MTD. Results: As of Feb 3, 2023, 35 pts with a median age of 53 were treated with ≥1 dose of ESG401 during escalation at doses of 2-20 mg/kg once Q3W (Regimen A), or 12-16 mg/kg D1,8,15 in a 4-week cycle (Regimen B). 80% of pts were ECOG = 1, 63% of pts were 3L+; median 4 (2-10) prior lines, 94% of pts had visceral metas (11% brain, 63% liver, 60% lung) at baseline. Only one patient treated with 20 mg/kg experienced DLT. The MTD has not reached either regimen. The most common TRAEs were leukopenia (80%), neutropenia (69%), anemia (66%), fatigue (54%), nausea (51%), and vomiting (46%). The most common ≥ Grade 3 TRAEs were leukopenia (29%) and neutropenia (31%). There was no ≥ Grade 3 thrombocytopenia, diarrhea, skin rash, or oral mucositis. No Interstitial Lung Disease (ILD) was observed. Of 33 efficacy evaluable (EE) pts, 12 partial responses (PR) were observed and 4 pts had achieved SD ≥24 wks. The first dose level found PR (16 mg/kg Q3W) and afterward were taken as therapeutically relevant doses (TRD). The ORR and DCR were 36% (4/11) and 64% (7/11) in 11 TRD pts with TNBC, while 62% (8/13) and 77% (10/13) in 13 TRD pts with HR+/HER2-BC. Two pts with brain metas both got an obvious intracranial response (one pt whose intracranial lesion was significantly reduced to too small to measure). Until the cut-off date, 11 (31%) pts remained on treatment. Three pts were treated over 12 mons and the longest on-treatment duration was 12.9 mons. One pt had consistent PR up to 10.8 mons. Conclusions: The preliminary data demonstrated that ESG401 is safe and well-tolerated with promising signals of efficacy in heavily pretreated pts. These encouraging findings warrant further clinical evaluation. Clinical trial information: NCT04892342 . [Table: see text]

Targeting Replication Stress and Chemotherapy Resistance with a Combination of Sacituzumab Govitecan and Berzosertib: A Phase I Clinical Trial.

Despite promising preclinical studies, toxicities have precluded combinations of chemotherapy and DNA damage response (DDR) inhibitors. We hypothesized that tumor-targeted chemotherapy delivery might enable clinical translation of such combinations. In a phase I trial, we combined sacituzumab govitecan, antibody-drug conjugate (ADC) that delivers topoisomerase-1 inhibitor SN-38 to tumors expressing Trop-2, with ataxia telangiectasia and Rad3-related (ATR) inhibitor berzosertib. Twelve patients were enrolled across three dose levels. Treatment was well tolerated, with improved safety over conventional chemotherapy-based combinations, allowing escalation to the highest dose. No dose-limiting toxicities or clinically relevant ≥grade 4 adverse events occurred. Tumor regressions were observed in 2 patients with neuroendocrine prostate cancer, and a patient with small cell lung cancer transformed from EGFR-mutant non-small cell lung cancer. ADC-based delivery of cytotoxic payloads represents a new paradigm to increase efficacy of DDR inhibitors. See related commentary by Berg and Choudhury, p. 3557.

Abstract CT268: Targeting replication stress and chemotherapy resistance with a combination of sacituzumab govitecan and berzosertib: A phase I clinical trial

Abstract Despite provocative preclinical results, dose-limiting toxicities have precluded rational combinations of cytotoxic chemotherapies that increase DNA damage with DNA damage response (DDR) inhibitors. We hypothesized that tumor-targeted delivery of cytotoxic chemotherapy might enable tolerable and active combinations with DDR inhibitors. We conducted a phase I clinical trial combining ataxia telangiectasia and Rad3-related (ATR) inhibitor berzosertib with sacituzumab govitecan, a trophoblast cell surface antigen 2 (Trop-2) directed antibody drug conjugate (ADC) that delivers high tumoral concentrations of topoisomerase 1 (TOP1) inhibitor SN-38. Depletion of ATR, the main transducer of replication stress is synthetically lethal with double-strand breaks (DSB) generated by TOP1 inhibitors. Patients with DDR gene-mutated or high replication stress solid tumors were enrolled since such tumors are particularly susceptible to ATR inhibition. Primary end point was identification of the maximum tolerated dose of the combination. Efficacy and pharmacodynamics were secondary end points. Using 3 + 3 dose escalation, sacituzumab govitecan (8-10 mg/m2, days 1, 8) and berzosertib (140-210 mg/m2, days 2, 9) were administered to 12 patients across three dose levels in 21-day cycles. The combination was well tolerated, with improved safety profile over conventional chemotherapy-based combinations, which allowed dose escalation to the highest planned dose level. There were no dose limiting toxicities. Common treatment-related adverse events (TRAE) were neutropenia (41.7%), diarrhea (50%), and fatigue (50%). Grade 3 TRAEs occurred in 58.3% of patients and included neutropenia (25%) and diarrhea (8.3%). There were no instances of febrile neutropenia or clinically significant grade 4 TRAEs. Pharmacodynamic studies showed evidence of ATR inhibition and enhanced DNA DSB in response to the combination. While no tumor responses were seen in three patients with DDR defects including BRCA1 and ATM mutations, two patients with neuroendocrine prostate cancer, a highly aggressive subtype of prostate cancer, showed partial or metabolic responses. A patient with EGFR-transformed small cell lung cancer (SCLC) also experienced partial response, together yielding objective responses in 3 of 12 evaluable patients (25%). Ongoing phase II expansion cohorts are evaluating efficacy of sacituzumab govitecan 10mg/m2 and berzosertib 210mg/m2 in patients with SCLC, extra-pulmonary small cell cancers, and DDR-mutated solid tumors. ADC-based delivery of cytotoxic payload represents a new therapeutic paradigm to extend the benefit of DDR inhibitors to target replication stress and chemotherapy resistance, with minimal added toxicities. Clinical trial information: NCT04826341 Citation Format: Melissa L. Abel, Nobukyuki Takahashi, Parth Desai, Cody Peer, Christophe Redon, Samantha Nichols, Rasa Vilimas, Min-Jung Lee, Sunmin Lee, Linda Sciuto, Danielle Pinkiert, Meenakshi Shelat, Chante Graham, Seth Steinberg, William D. Figg, Mirit Aladjem, Jane Trepel, Yves Pommier, Anish Thomas. Targeting replication stress and chemotherapy resistance with a combination of sacituzumab govitecan and berzosertib: A phase I clinical trial [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 CT268.

Abstract LB301: Molecular pharmacology and broad synergy of the novel ATR inhibitor M1774 with DNA damaging anticancer agents

Abstract Ataxia telangiectasia and Rad3-related (ATR) checkpoint kinase orchestrates DNA damage response and repair pathways stimulated by replicative stresses. Recent studies have established that pharmacological inhibition of ATR is clinically promising. As M1774 is an oral ATR inhibitor in clinical development, we explored the molecular basis by which M1774 induces cancer cell death. As a single agent, we found that M1774 suppresses cancer cell viability at nanomolar concentrations with a potency higher than ceralasertib and berzosertib, but lower than gartisertib (M4344) and elimusertib in the small cell lung cancer (SCLC) cell lines H146, H82, and DMS114. We found that M1774 efficiently suppresses the ATR/CHK1 checkpoints. While M1774 alone induced apoptosis and G2/M cell cycle arrest at micromolar concentrations, at a non-toxic low dose, M1774 enhanced TOP1 inhibitor-mediated cancer cell death by preventing replication arrest and inducing DNA damage detected by EdU and γH2AX staining. Tandem mass tagging (TMT) coupled with mass spectrometry revealed that M1774 combined with SN-38 increases the expression of replication-related proteins (TIPIN, CDC45, TIMELESS, and RPA1) and G2/M-related proteins (PLK1 and CCNB1). To establish the synergistic combinations of M1774 with clinical anticancer DNA damaging agents in preclinical models, we performed experiments in cancer cell lines, patient-derived organoids, and xenograft models. Low doses of M1774 significantly synergized with the clinical TOP1 inhibitor SN-38, the TOP2 inhibitor etoposide, cisplatin, and the PARP inhibitor talazoparib in SCLC cell lines. We also found that M1774 significantly reversed chemoresistance to DNA-damaging agents in cancer cells lacking SLFN11 expression, suggesting that SLFN11 expression can be utilized for combination therapy with M1774 as a biomarker. The synergistic efficacy between M1774 and DNA-damaging agents was confirmed in SCLC patient-derived organoids, colon cancer patient-derived organoids, and H82 SCLC xenografts. Together, these results provide insights into the molecular mechanism and potential combination strategies for M1774 in cancer therapy. Citation Format: Ukhyun Jo, Yasuhiro Arakawa, Astrid Zimmermann, Daiki Taniyama, Makito Mizunuma, Lisa M Jenkins, Suresh Kumar, Frank T Zenke, Yves Pommier. Molecular pharmacology and broad synergy of the novel ATR inhibitor M1774 with DNA damaging anticancer agents [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 LB301.

TROP2 expression and SN38 antitumor activity in malignant pleural mesothelioma cells provide a rationale for antibody-drug conjugate therapy

ObjectivesMalignant pleural mesothelioma (MPM) is an aggressive cancer which at large is not amenable to curative surgery. Despite the recent approval of immune checkpoint inhibitor therapy, the response rates and survival following systemic therapy is still limited. Sacituzumab govitecan is an antibody-drug conjugate targeting the topoisomerase I inhibitor SN38 to trophoblast cell-surface antigen 2 (TROP-2)-positive cells. Here we have explored the therapeutic potential of sacituzumab govitecan in MPM models. Materials and methodsTROP2 expression was analyzed in a panel of two well established and 15 pleural effusion derived novel lines by RT-QPCR and immunoblotting, TROP2 membrane-localization was studied by flow cytometry and immunohistochemistry. Cultured mesothelial cells and pneumothorax pleura served as controls. The sensitivity of MPM cell lines to irinotecan and SN38 was studied using cell viability, cell cycle, apoptosis and DNA damage assays. Drug sensitivity of cell lines was correlated with RNA expression of DNA repair genes. Drug sensitivity was defined as an IC50 below 5 nM in the cell viability assay. ResultsTROP2 expression was detected at RNA and protein level in 6 of the 17 MPM cell lines, but not in in cultured mesothelial control cells or in the mesothelial layer of the pleura. TROP2 was detectable on the cell membrane in 5 MPM lines and was present in the nucleus in 6 cell models. Ten of 17 MPM cell lines showed sensitivity to SN38 treatment, among those 4 expressed TROP2. High AURKA RNA expression and high proliferation rate correlated with sensitivity to SN38-induced cell death, DNA damage response, cell cycle arrest and cell death. Sacituzumab govitecan treatment effectively induced cell cycle arrest and cell death in TROP2-positive MPM cells. ConclusionTROP2 expression and sensitivity to SN38 in MPM cell lines support biomarker-selected clinical exploration of sacituzumab govitecan in patients with MPM.

SPDR-4 SLFN11 RENDERS MEDULLOBLASTOMA SENSITIVE TO DNA DAMAGE CHEMOTHERAPIES

Abstract Background Intensive chemotherapeutic regimens with craniospinal irradiation have greatly improved survival in medulloblastoma patients. However, survival markedly differs among molecular subgroups and their biomarkers are unknown. Through unbiased screening, we found Schlafen family member 11 (SLFN11), which is known to improve response to DNA damaging agents in various cancers, to be one of the top prognostic markers in medulloblastomas. Hence, we explored the expression and functions of SLFN11 in medulloblastoma. Methods SLFN11 expression for each subgroup was assessed by immunohistochemistry in 98 medulloblastoma patient samples and by analyzing transcriptomic databases. We genetically or epigenetically modulated SLFN11 expression in medulloblastoma cell lines and determined cytotoxic response to the DNA damaging agents cisplatin and topoisomerase I inhibitor SN-38 in vitro and in vivo. Results High SLFN11 expressing cases exhibited significantly longer survival than low expressing cases. SLFN11 was highly expressed in the WNT-activated subgroup and in a proportion of the SHH-activated subgroup. While WNT activation was not a direct cause of the high expression of SLFN11, a specific hypomethylation locus on the SLFN11 promoter was significantly correlated with high SLFN11 expression. Overexpression or deletion of SLFN11 made medulloblastoma cells sensitive and resistant to cisplatin and SN-38, respectively. Pharmacological upregulation of SLFN11 by the brain-penetrant histone deacetylase-inhibitor RG2833 markedly increased sensitivity to cisplatin and SN-38 in SLFN11-negative medulloblastoma cells. Intracranial xenograft studies also showed marked sensitivity to cisplatin by SLFN11-overexpression in medulloblastoma cells. Conclusions High SLFN11 expression renders favorable outcomes in WNT-activated and a subset of SHH-activated medulloblastoma through enhancing response to cisplatin.

CTNI-58. IMPROVING TRIAL ACCESS FOR PEOPLE WITH RARE CNS TUMORS: AN NCI-CONNECT PHASE I/II TRIAL OF PLX038 IN TUMORS WITH MYC/MYCN AMPLIFICATIONS INCORPORATING CORRELATIVE STUDIES AND PATIENT REPORTED OUTCOMES

Abstract MYC genes encode transcription factors that regulate expression of genes involved in cell division. Supraphysiologic levels of MYC drive oncogenesis in many human cancers and induce the formation of topoisome complexes, essential to managing the torsional strain that would otherwise oppose transcription and replication. MYC/MYCN amplifications are seen in multiple primary CNS tumors including 30% of relapsed medulloblastomas (MB) and a subtype of ependymoma (EPN) where MYCN amplification defines the tumor. PLX038 is a PEGylated macromolecule containing the topoisomerase I inhibitor SN-38. PEGylations lead to longer half-life, tumor accumulation and decreased toxicity. We hypothesize that PLX038 will be efficacious in primary CNS tumors driven by MYC/MYCN amplifications by targeting the MYC-induced topoisome (Das, S.K. 2022 DOI10.1016/j.molcel.2021.11.016). METHODS Trial in development. Design/Objectives: Phase I (N = 12) – determine the maximum tolerated dose and recommended phase 2 dose of PLX038 in recurrent primary CNS tumors using TITE-BOIN design. Phase II - 3 independent cohorts, N = 30 for each group with interim analysis after 10 each: 2A - MYC-N amplified EPN post resection and radiotherapy (PFS); 2B - Recurrent MYCN amplified EPN or MYC/MYCN amplified MB (disease control rate, DCR); 2C - Other recurrent primary CNS tumors with MYC/MYCN amplifications (DCR). Additional measures: PROs from the Office of Patient-Centered Outcomes Research (OpCORe) web-based early phase trial endpoints; biomarkers of PLX038 distribution and response/resistance in peripheral blood, tumor tissue, hair follicles, and abdominal wall fat pad tissue. CONCLUSIONS NCI-CONNECT’s commitment to innovative designs can mitigate some of the barriers to access experimental therapy for people with rare CNS tumors. This basket trial will enroll participants with unifying molecular features and potential susceptibility to PLX038 despite different histological diagnoses. Furthermore, incorporation of comprehensive PROs will help identify clinical benefit and tolerability while correlative studies in tumor and other tissues will help identify biomarkers of response or resistance.

PLX038: A Long-Acting Topoisomerase I Inhibitor With Robust Antitumor Activity in ATM-Deficient Tumors and Potent Synergy With PARP Inhibitors.

Alterations in the ATM gene are among the most common somatic and hereditary cancer mutations, and ATM-deficient tumors are hypersensitive to DNA-damaging agents. A synthetic lethal combination of DNA-damaging agents and DNA repair inhibitors could have widespread utility in ATM-deficient cancers. However, overlapping normal tissue toxicities from these drug classes have precluded their clinical translation. We investigated PLX038, a releasable polyethylene glycol-conjugate of the topoisomerase I inhibitor SN-38, in ATM wild-type and null isogenic xenografts and in a BRCA1-deficient xenograft. PLX038 monotherapy and combination with PARP inhibition potently inhibited the growth of both BRCA1- and ATM-deficient tumors. A patient with an ATM-mutated breast cancer treated with PLX038 and the PARP inhibitor rucaparib achieved rapid, symptomatic, and radiographic complete response lasting 12 months. Single-agent PLX038 or PLX038 in combination with DNA damage response inhibitors are novel therapeutic paradigms for patients with ATM-loss cancers.

Targeted Ultrafine Iron Oxide Nanoparticles for Delivery of the Topoisomerase Inhibitor SN38 and Ovarian Cancer Treatment

Ovarian cancer remains a challenge to decrease mortality and improve diagnostic efficiency in gynecological cancers. To develop a delivery system capable of efficient cancer cell targeting and delivering novel efficacious therapeutics, we assembled folic acid (FA) conjugated ultrafine iron oxide nanoparticles (uIONP) with encapsulation of DNA topoisomerase inhibitor SN38, which target ovarian cancer cells with over-expression of folate receptor alpha (FRα) and deliver SN38 to induce apoptosis. The assembled FA-uIONP-SN38 exhibited higher drug loading efficiency than the larger counterparts with core diameters more than 10 nm. The targeting specificity of FA-uIONP-SN38 for SKOV-3 cancer cells was validated, with HEK293 kidney cells and Raw264.7 macrophages as non-targeted cell line control. It was found that more SKOV-3 cancer cells were killed due to apoptosis by FA-uIONP-SN38 at the same SN38 dosages compared with uIONP-SN38 and free SN38, respectively. The delivery of this inhibitor to SKOV-3 cancer cells by FRα-targeted FA-uIONP carrier was enhanced by about 10-folds with less cytotoxicity comparing to the free drug SN38. The developed FA-uIONP-SN38 holds a great potential as a theranostic approach in treating ovarian cancer.

Targeted Delivery of DNA Topoisomerase Inhibitor SN38 to Intracranial Tumors of Glioblastoma Using Sub‐5 Ultrafine Iron Oxide Nanoparticles (Adv. Healthcare Mater. 14/2022)

Targeted Delivery of DNA Topoisomerase Inhibitor SN38 to Intracranial Tumors of Glioblastoma Using Sub‐5 Ultrafine Iron Oxide Nanoparticles (Adv. Healthcare Mater. 14/2022)