LP-184, a Novel Acylfulvene Molecule, Exhibits Anticancer Activity against Diverse Solid Tumors with Homologous Recombination Deficiency.

Homologous recombination (HR)-related gene alterations are present in a significant subset of prostate, breast, ovarian, pancreatic, lung, and colon cancers rendering these tumors as potential responders to specific DNA damaging agents. A small molecule acylfulvene prodrug, LP-184, metabolizes to an active compound by the oxidoreductase activity of enzyme prostaglandin reductase 1 (PTGR1), which is frequently elevated in multiple solid tumor types. Prior work demonstrated that cancer cell lines deficient in a spectrum of DNA damage repair (DDR) pathway genes show increased susceptibility to LP-184. Here, we investigated the potential of LP-184 in targeting multiple tumors with impaired HR function and its mechanism of action as a DNA damaging agent. LP-184 induced elevated DNA double-strand breaks in HR deficient (HRD) cancer cells. Depletion of key HR components BRCA2 or ataxia telangiectasia mutated (ATM) in cancer cells conferred up to 12-fold increased sensitivity to the LP-184. LP-184 showed nanomolar potency in a diverse range of HRD cancer models, including prostate cancer organoids, leiomyosarcoma cell lines, and patient-derived tumor graft models of lung, pancreatic, and prostate cancers. LP-184 demonstrated complete, durable tumor regression in 10 patient-derived xenograft (PDX) models of HRD triple-negative breast cancer (TNBC) including those resistant to PARP inhibitors (PARPi). LP-184 further displayed strong synergy with PARPi in ovarian and prostate cancer cell lines as well as in TNBC PDX models. These preclinical findings illustrate the potential of LP-184 as a pan-HRD cancer therapeutic. Taken together, our results support continued clinical evaluation of LP-184 in a large subset of HRD solid tumors. SIGNIFICANCE: New agents with activity against DDR-deficient solid tumors refractory to standard-of-care therapies are needed. We report multiple findings supporting the potential for LP-184, a novel alkylating agent with three FDA orphan drug designations, to fill this void clinically: strong nanomolar potency; sustained, durable regression of solid tumor xenografts; synthetic lethality with HR defects. LP-184 adult phase IA trial to assess safety in advanced solid tumors is ongoing.

Conditional Dependency of LP-184 on Prostaglandin Reductase 1 is Synthetic Lethal in Pancreatic Cancers with DNA Damage Repair Deficiencies.

The greater efficacy of DNA-damaging drugs for pancreatic adenocarcinoma (PDAC) relies on targeting cancer-specific vulnerabilities while sparing normal organs and tissues due to their inherent toxicities. We tested LP-184, a novel acylfulvene analog, for its activity in preclinical models of PDAC carrying mutations in the DNA damage repair (DDR) pathways. Cytotoxicity of LP-184 is solely dependent on prostaglandin reductase 1 (PTGR1), so that PTGR1 expression robustly correlates with LP-184 cytotoxicity in vitro and in vivo. Low-passage patient-derived PDAC xenografts with DDR deficiencies treated ex vivo are more sensitive to LP-184 compared with DDR-proficient tumors. Additional in vivo testing of PDAC xenografts for their sensitivity to LP-184 demonstrates marked tumor growth inhibition in models harboring pathogenic mutations in ATR, BRCA1, and BRCA2. Depletion of PTGR1, however, completely abrogates the antitumor effect of LP-184. Testing combinatorial strategies for LP-184 aimed at deregulation of nucleotide excision repair proteins ERCC3 and ERCC4 established synergy. Our results provide valuable biomarkers for clinical testing of LP-184 in a large subset of genetically defined characterized refractory carcinomas. High PTGR1 expression and deleterious DDR mutations are present in approximately one third of PDAC making these patients ideal candidates for clinical trials of LP-184.

Preclinical Efficacy of LP-184, a Tumor Site Activated Synthetic Lethal Therapeutic, in Glioblastoma.

PURPOSE: Glioblastoma (GBM) is the most common brain malignancy with median survival <2 years. Standard-of-care temozolomide has marginal efficacy in approximately 70% of patients due to MGMT expression. LP-184 is an acylfulvene-derived prodrug activated by the oxidoreductase PTGR1 that alkylates at N3-adenine, not reported to be repaired by MGMT. This article examines LP-184 efficacy against preclinical GBM models and identifies molecular predictors of LP-184 efficacy in clinical GBM. EXPERIMENTAL DESIGN: LP-184 effects on GBM cell viability and DNA damage were determined using cell lines, primary PDX-derived cells and patient-derived neurospheres. GBM cell sensitivities to LP-184 relative to temozolomide and MGMT expression were examined. Pharmacokinetics and CNS bioavailability were evaluated in mice with GBM xenografts. LP-184 effects on GBM xenograft growth and animal survival were determined. Machine learning, bioinformatic tools, and clinical databases identified molecular predictors of GBM cells and tumors to LP-184 responsiveness. RESULTS: LP-184 inhibited viability of multiple GBM cell isolates including temozolomide-resistant and MGMT-expressing cells at IC50 = approximately 22-310 nmol/L. Pharmacokinetics showed favorable AUCbrain/plasma and AUCtumor/plasma ratios of 0.11 (brain Cmax = 839 nmol/L) and 0.2 (tumor Cmax = 2,530 nmol/L), respectively. LP-184 induced regression of GBM xenografts and prolonged survival of mice bearing orthotopic xenografts. Bioinformatic analyses identified PTGR1 elevation in clinical GBM subtypes and associated LP-184 sensitivity with EGFR signaling, low nucleotide excision repair (NER), and low ERCC3 expression. Spironolactone, which induces ERCC3 degradation, decreased LP-184 IC50 3 to 6 fold and enhanced GBM xenograft antitumor responses. CONCLUSIONS: These results establish LP-184 as a promising chemotherapeutic for GBM with enhanced efficacy in intrinsic or spironolactone-induced TC-NER-deficient tumors.

LP-284, a small molecule acylfulvene, exerts potent antitumor activity in preclinical non-Hodgkin's lymphoma models and in cells deficient in DNA damage repair.

Despite advances in therapies treating non-Hodgkin's lymphoma (NHL), 20~40% of patients experience relapsed or refractory disease. While solid tumors with homologous recombination deficiencies have been successfully targeted with synthetic lethal agents such as poly-ADP ribose polymerase (PARP) inhibitors, such synthetic lethality strategy has not yet been approved to treat patients with NHL. Here we investigated the mechanism of action (MoA) and therapeutic potential of a new-generation acylfulvene compound, LP-284, in both in vitro and in vivo NHL models. One of LP-284's MoA includes inducing the repair of double-strand DNA break (DSB). We found that LP-284 exerts nanomolar potency in a panel of hematological cancer cell lines including fifteen NHL cell lines. In vivo, LP-284 treatment prolongs the survival of mantle cell lymphoma (MCL) cell line JeKo-1 derived xenograft mice by two-fold and shows increased efficacy over bortezomib and ibrutinib. In addition, LP-284 is capable of inhibiting tumor growth of JeKo-1 xenografts that are refractory to bortezomib or ibrutinib. We further showed that LP-284 is particularly lethal in cells with deficient DNA damage response and repair, a targetable vulnerability in NHL.

The acylfulvene alkylating agent, LP-184, retains nanomolar potency in non-small cell lung cancer carrying otherwise therapy-refractory mutations.

More than 40% of non-small cell lung cancer (NSCLC) patients lack actionable targets and require non-targeted chemotherapeutics. Many become refractory to drugs due to underlying resistance-associated mutations. KEAP1 mutant NSCLCs further activate NRF2 and upregulate its client PTGR1. LP-184, a novel alkylating agent belonging to the acylfulvene class is a prodrug dependent upon PTGR1. We hypothesized that NSCLC with KEAP1 mutations would continue to remain sensitive to LP-184. LP-184 demonstrated highly potent anticancer activity both in primary NSCLC cell lines and in those originating from brain metastases of primary lung cancers. LP-184 activity correlated with PTGR1 transcript levels but was independent of mutations in key oncogenes (KRAS and KEAP1) and tumor suppressors (TP53 and STK11). LP-184 was orders of magnitude more potent in vitro than cisplatin and pemetrexed. Correlative analyses of sensitivity with cell line gene expression patterns indicated that alterations in NRF2, MET, EGFR and BRAF consistently modulated LP-184 sensitivity. These correlations were then extended to TCGA analysis of 517 lung adenocarcinoma patients, out of which 35% showed elevated PTGR1, and 40% of those further displayed statistically significant co-occurrence of KEAP1 mutations. The gene correlates of LP-184 sensitivity allow additional personalization of therapeutic options for future treatment of NSCLC.

A machine learning-based gene signature of response to the novel alkylating agent LP-184 distinguishes its potential tumor indications.

BACKGROUND: Non-targeted cytotoxics with anticancer activity are often developed through preclinical stages using response criteria observed in cell lines and xenografts. A panel of the NCI-60 cell lines is frequently the first line to define tumor types that are optimally responsive. Open data on the gene expression of the NCI-60 cell lines, provides a unique opportunity to add another dimension to the preclinical development of such drugs by interrogating correlations with gene expression patterns. Machine learning can be used to reduce the complexity of whole genome gene expression patterns to derive manageable signatures of response. Application of machine learning in early phases of preclinical development is likely to allow a better positioning and ultimate clinical success of molecules. LP-184 is a highly potent novel alkylating agent where the preclinical development is being guided by a dedicated machine learning-derived response signature. We show the feasibility and the accuracy of such a signature of response by accurately predicting the response to LP-184 validated using wet lab derived IC50s on a panel of cell lines. RESULTS: We applied our proprietary RADR® platform to an NCI-60 discovery dataset encompassing LP-184 IC50s and publicly available gene expression data. We used multiple feature selection layers followed by the XGBoost regression model and reduced the complexity of 20,000 gene expression values to generate a 16-gene signature leading to the identification of a set of predictive candidate biomarkers which form an LP-184 response gene signature. We further validated this signature and predicted response to an additional panel of cell lines. Considering fold change differences and correlation between actual and predicted LP-184 IC50 values as validation performance measures, we obtained 86% accuracy at four-fold cut-off, and a strong (r = 0.70) and significant (p value 1.36e-06) correlation between actual and predicted LP-184 sensitivity. In agreement with the perceived mechanism of action of LP-184, PTGR1 emerged as the top weighted gene. CONCLUSION: Integration of a machine learning-derived signature of response with in vitro assessment of LP-184 efficacy facilitated the derivation of manageable yet robust biomarkers which can be used to predict drug sensitivity with high accuracy and clinical value.