Pancreatic Xenografts Research Articles

Exploring the Interplay Between Radioimmunoconjugates and Fcγ Receptors in Genetically Engineered Mouse Models of Cancer.

Fcγ receptors (FcγR) are responsible for many of the interactions between immunoglobulins (IgG) and immune cells. In biomedicine, this interplay is critical to the activity of several types of immunotherapeutics; however, relatively little is known about how FcγRs affect the in vivo performance of radiolabeled antibodies. A handful of recent preclinical studies suggest that binding by FcγR-and particularly FcγRI-can affect the pharmacokinetic profiles of 89Zr-labeled radioimmunoconjugates, but there are no extant studies in immunocompetent or genetically engineered mouse models of cancer. In the investigation at hand, we synthesized and characterized 89Zr-labeled probes based on wild-type and aglycosylated variants of the CA19-9-targeting antibody 5B1 and evaluated their in vivo behavior in several murine models of cancer, including immunocompetent and FcγR-humanized mice. The aglycosylated desferrioxamine (DFO)-bearing immunoconjugate DFO-N297A5B1 displayed identical binding to CA19-9-expressing cells compared to its wild-type analogue (DFO-5B1) but exhibited dramatically attenuated affinity for several FcγR. Positron emission tomography imaging and biodistribution studies with [89Zr]Zr-DFO-5B1 and [89Zr]Zr-DFO-N297A5B1 were subsequently performed in several strains of mice bearing CA19-9-expressing BxPC3 human pancreatic ductal adenocarcinoma and B16F10-FUT3 murine melanoma xenografts. Significant differences in the pharmacokinetics of the two radioimmunoconjugates were observed in tumor-bearing immunocompromised NSG mice, but these differences failed to materialize in immunocompetent C57BL/6 and FcγR-humanized C57BL/6 mice with B16F10-FUT3 xenografts. We hypothesize that these observations are related to the presence or absence of endogenous IgG. NSG mice completely lack endogenous IgG, and thus their mFcγR are free to bind radioimmunoconjugates and alter their pharmacokinetic behavior. In contrast, C57BL/6 and FcγR-humanized C57BL/6 mice both have endogenous IgG that occupy their FcγR (murine for the former and human for the latter), precluding interactions with radioimmunoconjugates. Ultimately, these data suggest that understanding the interplay between radiolabeled antibodies and FcγR is critical during the preclinical evaluation of radioimmunoconjugates.

Inhibition of phospholipase D1 reduces pancreatic carcinogenesis in mice partly through a FAK-dependent mechanism.

Phospholipase D (PLD) plays a critical role in cancer progression. However, its role in pancreatic cancer remains unclear. Thus, we evaluated the role of PLD1, one of two classical isoforms of PLD, in pancreatic carcinogenesis in vivo. The role of PLD1 in tumor growth was evaluated by subcutaneously transplanting human MIA PaCa-2 cells expressing endogenous PLD1 levels (Ctr KD cells) or cells in which PLD1 was knocked down (Pld1 KD cells) into immunodeficient mice. Twenty days post-implantation, tumors that arose from Pld1-KD cells were significantly smaller, compared to controls (Ctr KD). Then, we assessed the role of PLD1 in the tumor microenvironment, by subcutaneously implanting mouse LSL-KrasG12D/+;Trp53R172H/+;Pdx-1-Cre (KPC) cells into wild-type (WT) or PLD1 knockout (Pld1-/-) mice. Compared to WT, tumor growth was attenuated in Pld1-/- mice by 39%, whereas treatment of Pld1-/- mice with gemcitabine reduced tumor growth by 79%. When PLD1 was ablated in LSL-KrasG12D;Ptf1Cre/+ (KC) mice, no reduction in acinar cell loss was observed, compared to KC mice. Finally, treatment of KC mice with a small molecule inhibitor of PLD1 and PLD2 (FIPI) significantly reduced acinar cell loss and cell proliferation, compared to vehicle-treated mice. Mechanistically, the effect of PLD on tumor growth is mediated, partly, by the FAK pathway. In conclusion, while PLD1 is a critical regulator of pancreatic xenograft and allograft growth, playing an important role at the tumor and at the microenvironment levels, inhibition of PLD1 and PLD2 are necessary to reduce pancreatic carcinogenesis in KC mice, and might represent a novel therapeutic target.

A potent nano-strategy for dual energy deprivation to inhibit pancreatic cancer progression

Pancreatic cancer is a highly malignant tumor that poses significant threats to public health, and glycolysis plays a crucial role in its energy metabolism. Here, glycolysis was confirmed to be directly associated with poor prognosis through the use of clinical samples from 130 patients with pancreatic ductal adenocarcinoma (PDAC), and the effectiveness of zinc ions (Zn2+) in inhibiting glycolysis-related genes was further validated. Therefore, polyvinyl pyrrolidone (PVP)-modified zinc sulfide nanomedicines (ZnS-PVP) were developed for dual energy suppression by targeting glycolysis and mitochondrial respiration in pancreatic cancer. On the one hand, the released Zn2+ efficiently inhibited glycolysis in pancreatic cancer cells through the PI3K-Akt-mTOR-HIF-1α signaling axis. On the other hand, acid-responsive release of hydrogen sulfide (H2S) gas damaged mitochondria and further reduced energy compensation by inhibiting oxidative phosphorylation. This two-pronged energy deprivation nano-strategy effectively eliminated pancreatic cancer cells and was proven to overcome chemotherapeutic resistance. Moreover, ZnS-PVP administration combined with immune checkpoint blockade (ICB) therapy significantly suppressed tumor progression in mouse orthotopic pancreatic tumor models, as also demonstrated in a pancreatic cancer patient-derived xenograft (PDX) model. Our work highlights the positive role of bioactive metal ions in targeting tumor energy metabolism and the great potential of nano-strategy for energy deprivation in the treatment of pancreatic cancer.

αvβ3 integrin-targeted magnetic resonance imaging in a pancreatic cancer mouse model using RGD-modified liposomes encapsulated with Fe-deferoxamine.

Magnetic resonance (MR) imaging is a powerful imaging modality for obtaining anatomical information with high spatial and temporal resolution. In the drug delivery system (DDS) framework, nanoparticles such as liposomes are potential candidates for MR imaging. We validated that RGD peptides are possible targeting molecules for pancreatic cancer with αvβ3 integrin expression. This study aimed to evaluate RGD-modified liposomes loaded with ferrioxamine B for pancreatic cancer imaging. We synthesized four types of RGD-modified liposomes encapsulated with ferrioxamine B (SH-, H-, M-, and L-RGD-liposomes). The binding affinity of RGD-modified liposomes was evaluated in a competitive inhibition study using 125I-echistatin. To investigate the pharmacokinetics of RGD-modified liposomes, a biodistribution study using RGD-liposomes labeled with 111In was carried out in a human pancreatic cancer PANC-1 xenograft mouse model. Finally, MR was performed using ferrioxamine-B-loaded liposomes. RGD-liposomes inhibited the binding of 125I-echistatin to RGD. The biodistribution study revealed that 111In-RGD-liposomes accumulated significantly in the liver and spleen. Among the 111In-RGD-liposomes, 111In-H-RGD-liposomes showed the highest tumor-to-normal tissue ratio. In the MR study, H-RGD-liposomes loaded with ferrioxamine B showed higher tumor-to-muscle signal ratios than RKG-liposomes loaded with ferrioxamine B (control). We successfully synthesized RGD-liposomes that can target αvβ3 integrin.

Development of novel imipridone derivatives with potent anti-cancer activities as human caseinolytic peptidase P (hClpP) activators

Based on a clinically staged small molecular hClpP activator ONC201, a class of imipridone derivatives was designed and synthesized. These compounds were evaluated in a protease hydrolytic assay, as well as cell growth inhibition assays in three cancer cell lines, MIA PACA-2, HCT116, and MV4-11. A number of compounds that can more potently activate hClpP and more effectively inhibit cell growth in the three cancer cell lines than ONC201 were identified. The most potent compound, ZYZ-17, activated hClpP with an EC50 value of 0.24 µM and inhibited the growth of the three cancer cell lines with IC50 values of less than 10 nM. Mechanism studies for ZYZ-17 revealed that it potently activates cellular hClpP, efficiently induces the degradation of hClpP substrates, and robustly induces apoptosis in the three cancer cell lines. Furthermore, ZYZ-17 demonstrated a promising pharmacokinetic (PK) profile and exhibited highly potent in vivo antitumor activity in a pancreatic cancer MIA PACA-2 xenograft model in BALB/c nude mice.

Cardiac Atrophy, Dysfunction, and Metabolic Impairments: A Cancer-Induced Cardiomyopathy Phenotype

Muscle atrophy and weakness are prevalent features of cancer. Although extensive research has characterized skeletal muscle wasting in cancer cachexia, limited studies have investigated how cardiac structure and function are affected by therapy-naive cancer. Herein, orthotopic, syngeneic models of epithelial ovarian cancer and pancreatic ductal adenocarcinoma, and a patient-derived pancreatic xenograft model, were used to define the impact of malignancy on cardiac structure, function, and metabolism. Tumor-bearing mice developed cardiac atrophy and intrinsic systolic and diastolic dysfunction, with arterial hypotension and exercise intolerance. In hearts of ovarian tumor-bearing mice, fatty acid-supported mitochondrial respiration decreased, and carbohydrate-supported respiration increased-showcasing a substrate shift in cardiac metabolism that is characteristic of heart failure. Epithelial ovarian cancer decreased cytoskeletal and cardioprotective gene expression, which was paralleled by down-regulation of transcription factors that regulate cardiomyocyte size and function. Patient-derived pancreatic xenograft tumor-bearing mice show altered myosin heavy chain isoform expression-also a molecular phenotype of heart failure. Markers of autophagy and ubiquitin-proteasome system were upregulated by cancer, providing evidence of catabolic signaling that promotes cardiac wasting. Together, two cancer types were used to cross-validate evidence of the structural, functional, and metabolic cancer-induced cardiomyopathy, thus providing translational evidence that could impact future medical management strategies for improved cancer recovery in patients.

Onychocolone A produced by the fungus Onychocola sp. targets cancer stem cells and stops pancreatic cancer progression by inhibiting MEK2-dependent cell signaling

Pancreatic cancer (PC) shows a high fatality rate that can only be faced with a combination of surgery and chemotherapy or palliative treatment in the case of advanced patients. Besides, PC tumors are enriched with subpopulations of cancer stem cells (CSCs) that are resistant to the existing chemotherapeutic agents, which raises an important need for the identification of new drugs. To fill this gap, we have tested the anti-tumoral activity of microbial extracts, which chemical diversity offers a broad spectrum of potential new bioactive compounds. Extracts derived from the fungus Onychocola sp. CF-107644 were assayed via high throughput screening followed by bioassay-guided fractionation and resulted in the identification and isolation of six benzophenone derivatives with antitumoral activity: onychocolones A-F (#1-6). The structures of the compounds were established by spectroscopic methods, including ESI-TOF MS, 1D and 2D NMR analyses and X-ray diffraction. Compounds #1-4 significantly inhibited the growth of the pancreas tumoral cell lines, with low-micromolar Median Effective Doses (ED50s). Compound #1 (onychocolone A) was prioritized for further profiling due to its pro-apoptotic effect, which was further validated on 3D spheroids and pancreatic CSCs. Protein expression assays showed that the effect was mechanistically linked to the inhibition of MEK onco-signaling pathway. The efficacy of onychocolone A was also demonstrated in vivo by the reduction of tumor growth in a pancreatic xenograft mouse model generated by CSCs. Altogether, the data support that onychocolone A is a promising new small molecule for hit-to-lead development of a new treatment for PC.

14-3-3σ downregulation sensitizes pancreatic cancer to carbon ions by suppressing the homologous recombination repair pathway.

This study explored the role of 14-3-3σ in carbon ion-irradiated pancreatic adenocarcinoma (PAAD) cells and xenografts and clarified the underlying mechanism. The clinical significance of 14-3-3σ in patients with PAAD was explored using publicly available databases. 14-3-3σ was silenced or overexpressed and combined with carbon ions to measure cell proliferation, cell cycle, and DNA damage repair. Immunoblotting and immunofluorescence (IF) assays were used to determine the underlying mechanisms of 14-3-3σ toward carbon ion radioresistance. We used the BALB/c mice to evaluate the biological behavior of 14-3-3σ in combination with carbon ions. Bioinformatic analysis revealed that PAAD expressed higher 14-3-3σ than normal pancreatic tissues; its overexpression was related to invasive clinicopathological features and a worse prognosis. Knockdown or overexpression of 14-3-3σ demonstrated that 14-3-3σ promoted the survival of PAAD cells after carbon ion irradiation. And 14-3-3σ was upregulated in PAAD cells during DNA damage (carbon ion irradiation, DNA damaging agent) and promotes cell recovery. We found that 14-3-3σ resulted in carbon ion radioresistance by promoting RPA2 and RAD51 accumulation in the nucleus in PAAD cells, thereby increasing homologous recombination repair (HRR) efficiency. Blocking the HR pathway consistently reduced 14-3-3σ overexpression-induced carbon ion radioresistance in PAAD cells. The enhanced radiosensitivity of 14-3-3σ depletion on carbon ion irradiation was also demonstrated in vivo. Altogether, 14-3-3σ functions in tumor progression and can be a potential target for developing biomarkers and treatment strategies for PAAD along with incorporating carbon ion irradiation.

Vascular regional analysis unveils differential responses to anti-angiogenic therapy in pancreatic xenografts through macroscopic photoacoustic imaging.

Pancreatic cancer (PC) is a highly lethal malignancy and the third leading cause of cancer deaths in the U.S. Despite major innovations in imaging technologies, there are limited surrogate radiographic indicators to aid in therapy planning and monitoring. Amongst the various imaging techniques Ultrasound-guided photoacoustic imaging (US-PAI) is a promising modality based on endogenous blood (hemoglobin) and blood oxygen saturation (StO 2 ) contrast to monitor response to anti-angiogenic therapies. Adaptation of US-PAI to the clinical realm requires macroscopic configurations for adequate depth visualization, illuminating the need for surrogate radiographic markers, including the tumoral microvessel density (MVD). In this work, subcutaneous xenografts with PC cell lines AsPC-1 and MIA-PaCa-2 were used to investigate the effects of receptor tyrosine kinase inhibitor (sunitinib) treatment on MVD and StO 2 . Through histological correlation, we have shown that regions of high and low vascular density (HVD and LVD) can be identified through frequency domain filtering of macroscopic PA images which could not be garnered from purely global analysis. We utilized vascular regional analysis (VRA) of treatment-induced StO 2 and total hemoglobin (HbT) changes. VRA as a tool to monitor treatment response allowed us to identify potential timepoints of vascular remodeling, highlighting its ability to provide insights into the TME not only for sunitinib treatment but also other anti-angiogenic therapies.

A phase 1 trial evaluating the safety, tolerability, PK, and preliminary efficacy of QTX3034, an oral G12D-preferring multi-KRAS inhibitor, in patients with solid tumors with KRASG12D mutation.

TPS3172 Background: KRAS is one of the most mutated driver oncogenes across solid tumors, and KRAS G12D missense mutations have high prevalence in pancreatic, colorectal (CRC), and endometrial cancers. QTX3034 is a highly-selective, non-covalent, orally bioavailable, multi-KRAS inhibitor with potent activity across several KRAS variants, particularly G12D. QTX3034 binds to GDP-bound forms of mutant and wild-type KRAS with (sub)nanomolar affinities, allosterically inhibiting its conversion to the active state. QTX3034 inhibits KRAS signaling in in vitro studies and displays synergy with EGFR inhibitors. In vivo evaluation of QTX3034 resulted in significant tumor growth inhibition across various tumor models, with tumor regressions observed in 100% of animals in both HPAC pancreatic and GP2D colorectal KRAS G12D xenograft models. QTX3034 exhibits brain penetration in preclinical studies, including an orthotopic model of intracranial efficacy. This clinical trial is now enrolling patients with advanced solid tumors harboring a KRASG12D mutation. Methods: This is a first-in-human, multicenter, open-label phase 1 study in adult patients with advanced solid tumors with KRASG12D mutations, without prior direct KRAS inhibitor treatment (NCT06227377). The primary objectives are to evaluate the safety and tolerability and to determine maximum tolerated dose (MTD) and/or recommended phase 2 dose (RP2D) of QTX3034 alone, and in combination with cetuximab. Secondary objectives are characterization of the PK profile and preliminary antitumor activity. Part 1 consists of two dose-escalation arms: QTX3034 monotherapy (advanced solid tumors) and QTX3034 plus cetuximab (CRC) and will enroll 3-4 subjects per cohort in a Bayesian Logistic Regression Model (BLRM) design. QTX3034 will be administered orally in 21-day cycles until disease progression, intolerable toxicity, or withdrawal from treatment. Exploratory paired biopsies will be collected in Cycle 2 in a group of patients in the dose-escalation phase. Serial ctDNA will be collected in all patients. Following identification of a RP2D, QTX3034 will be evaluated in dose-expansion cohorts of up to 30 patients with various KRASG12D mutated tumor types to determine preliminary activity: pancreatic cancer, CRC, non-small cell lung cancer and endometrial cancer. To further optimize the recommended dose, patients will be randomized between two dose levels in the pancreatic cancer cohort. QTX3034 plus cetuximab will be explored following identification of a recommended combination dose in patients with CRC. Clinical trial information: NCT06227377 .

Half-life extension of single-domain antibody-drug conjugates by albumin binding moiety enhances antitumor efficacy.

Single-domain antibody-drug conjugates (sdADCs) have been proven to have deeper solid tumor penetration and intratumor accumulation capabilities due to their smaller size compared with traditional IgG format ADCs. However, one of the key challenges for improving clinical outcomes of sdADCs is their abbreviated in vivo half-life. In this study, we innovatively fused an antihuman serum albumin (αHSA) nanobody to a sdADCs targeting oncofetal antigen 5T4, conferring serum albumin binding to enhance the pharmacokinetic profiles of sdADCs. The fusion protein was conjugated with monomethyl auristatin E (MMAE) at s224c site mutation. The conjugate exhibited potent cytotoxicity against various tumor cells. Compared with the nonalbumin-binding counterparts, the conjugate exhibited a 10-fold extended half-life in wild-type mice and fivefold prolonged serum half-life in BxPC-3 xenograft tumor models as well as enhanced tumor accumulation and retention in mice. Consequently, n501-αHSA-MMAE showed potent antitumor effects, which were comparable to n501-MMAE in pancreatic cancer BxPC-3 xenograft tumor models; however, in human ovarian teratoma PA-1 xenograft tumor models, n501-αHSA-MMAE significantly improved antitumor efficacy. Moreover, the conjugate showed mitigated hepatotoxicity. In summary, our results suggested that fusion to albumin-binding moiety as a viable strategy can enhance the therapeutic potential of sdADCs through optimized pharmacokinetics.

Abstract LB053: A novel BRAF-specific allosteric inhibitor that effectively targets acquired RAFi-resistant BRAF(V600E) cancers and RAS-mutated cancers in vitro and in vivo

Abstract Targeting hyperactive RAS-RAF-MEK-ERK signaling for treating cancers has achieved a promising outcomes, and a number of RAS/RAF inhibitors have been applied to clinical cancer treatment. However, these inhibitors have only a short-term efficacy and their effectiveness is abrogated by acquired drug resistance in less than one year. For those resistant/refractory RAS/RAF-mutated cancers, there’s no effective therapeutics available in clinic. In this study, we developed a novel BRAF-specific inhibitor that docks on an allosteric site on BRAF kinase domain. This inhibitor effectively killed both primary and drug-resistant cancer cell lines with RAS mutation or BRAF(V600E) mutation in vitro and inhibited the growth of xenografted cancers derived from these cell lines in vivo without apparent toxicity. Mechanistically, we found that although this inhibitor blocked the catalytic activity of BRAF in vitro kinase assay, it triggered a super high ERK signaling through improving BRAF/CRAF heterodimerization and hence induced cellular apoptosis of cancer cells with RAS mutation or BRAF(V600E) mutation. Surprisingly, there’s no drug resistance occurring when treating cancer cells with RAS mutation or BRAF(V600E) mutation at a low concentration. To further evaluate the efficacy of this BRAF against RAS/RAF-mutated cancers in vivo, we constructed a series of pancreatic patient-derived xenograft cancer models with Kras G12D or G12V mutations, and found that our BRAF inhibitor exhibited a comparable or much better efficacy than Kras G12D-specific inhibitor on theses PDX models. Altogether, our data indicated that our BRAF-specific allosteric inhibitor may serve as a good therapeutic drug for treating BRAF(V600E)/RAS- cancers, particularly those resistant/refractory cancers, though it still need further evaluations via clinical trials. Citation Format: Jiancheng Hu. A novel BRAF-specific allosteric inhibitor that effectively targets acquired RAFi-resistant BRAF(V600E) cancers and RAS-mutated cancers in vitro and in vivo [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 LB053.

Design, synthesis, and evaluation of purine and pyrimidine-based KRAS G12D inhibitors: Towards potential anticancer therapy

Oncogenic RAS mutations, commonly observed in human tumors, affect approximately 30% of cancer cases and pose a significant challenge for effective cancer treatment. Current strategies to inhibit the KRAS G12D mutation have shown limited success, emphasizing the urgent need for new therapeutic approaches. In this study, we designed and synthesized several purine and pyrimidine analogs as inhibitors for the KRAS G12D mutation. Our synthesized compounds demonstrated potent anticancer activity against cell lines with the KRAS G12D mutation, effectively impeding their growth. They also exhibited low toxicity in normal cells, indicating their selective action against cancer cells harboring the KRAS G12D mutation. Notably, the lead compound, PU1-1 induced the programmed cell death of KRAS G12D-mutated cells and reduced the levels of active KRAS and its downstream signaling proteins. Moreover, PU1-1 significantly shrunk the tumor size in a pancreatic xenograft model induced by the KRAS G12D mutation, further validating its potential as a therapeutic agent. These findings highlight the potential of purine-based KRAS G12D inhibitors as candidates for targeted cancer therapy. However, further exploration and optimization of these compounds are essential to meet the unmet clinical needs of patients with KRAS-mutant cancers.

Abstract 447: Anti-tumor efficacy of ompenaclid, a creatine transporter inhibitor, in KRAS mutant tumors, including NSCLC

Abstract Background: Pre-clinical studies identified creatine metabolism to be activated in KRAS mutant metastatic colorectal cancer (CRC) as a mechanism to fuel the increased energy demands of RAS mutant tumors. Metastatic colon cancer cells overexpress and subsequently release creatine kinase-B (CKB) into the extracellular space, where it phosphorylates creatine to generate the high energy metabolite phosphocreatine. Ompenaclid is a small molecule inhibitor of the creatine transporter SLC6A8. Inhibition of SLC6A8 by ompenaclid prevents import of phosphocreatine/creatine, resulting in depletion of intracellular creatine, phosphocreatine and ATP levels. This results in tumor apoptosis and robust anti-tumor efficacy in CRC models with a wide range of KRAS mutant alleles. Ompenaclid is currently in a Phase 2 trial, in combination with FOLFIRI and bevacizumab, in patients with RAS-mutated second-line advanced/metastatic colorectal cancer. Here, we assessed efficacy of ompenaclid in tumors beyond CRC, including KRAS WT and KRAS mutant NSCLC. Methods: Athymic mice were inoculated with CRC, pancreatic, gastric or NSCLC cancer cell lines and treated with ompenaclid once tumors reached ~150 mm3, until termination. Plasma metabolites were quantified by LC-MS/MS and tumoral expression of CKB and SLC6A8 in NSCLC and CRC tumors was measured by qPCR after treatment with a control or ompenaclid-supplemented diet for ~10 days. Combination treatment with MRTX849, a KRAS G12C inhibitor, was conducted in a pancreatic MiaPaca xenograft model. Results: Our studies demonstrate ompenaclid anti-tumor efficacy in NSCLC and pancreatic xenografts. Notably, and consistent with findings in CRC, efficacy in NSCLC was more robust in RAS mutant cancer models, relative to those bearing KRAS WT tumors. Metabolite analysis of the plasma of tumor-bearing mice at baseline revealed that mice harboring RAS mutant tumors had higher levels of p-creatine and lower levels of ATP, indicating that RAS mutant tumors utilize higher amounts of ATP, which may be converted to phosphocreatine by tumoral CKB, as confirmed by qPCR analysis. This finding is consistent with metastatic RAS mutant CRC tumors utilizing extracellular ATP to generate phosphocreatine and thus exhibiting greater dependency on creatine metabolism relative to RAS WT tumors. Treatment with ompenaclid also demonstrated additive efficacy in combination with KRAS G12C inhibitors. In summary, these data support development of ompenaclid in indications beyond CRC, including NSCLC, as well as in combination with KRAS G12C inhibitors. Citation Format: PuiChi Lo, Victor Sanz Chavez, Shugaku Takeda, Masoud Tavazoie, Isabel Kurth. Anti-tumor efficacy of ompenaclid, a creatine transporter inhibitor, in KRAS mutant tumors, including NSCLC [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 447.

Abstract 3141: LCB02A an antibody drug conjugate (ADC) targeting CLDN18.2 expressing solid tumors

Abstract Claudin18.2 (CLDN18.2) is an isoform of CLDN18 with very limited expression in gastric epithelial cells in normal tissues, and aberrant overexpression in gastric, pancreatic, esophageal, ovarian, lung, and other solid tumors. This expression pattern presents a significant hurdle in developing an ADC targeting CLDN18.2 due to potential on-target toxicity to the gastrointestinal (GI) tract. LCB02A is an antibody-drug conjugate (ADC) that targets CLDN18.2, composed of a topoisomerase 1 inhibitor (TOPO1i) as payload (DAR4) linked to the CLDN18.2 specific binding PR301839 hClaudin18.2 (CLDN18.2) is an isoform of CLDN18 with very limited expression in gastric epithelial cells in normal tissues, and aberrant overexpression in gastric, pancreatic, esophageal, ovarian, lung, and other solid tumors. This expression pattern presents a significant hurdle in developing an ADC targeting CLDN18.2 due to potential on-target toxicity to the gastrointestinal (GI) tract. LCB02A is an antibody-drug conjugate (ADC) that targets CLDN18.2, composed of a topoisomerase 1 inhibitor (TOPO1i) as payload (DAR4) linked to the CLDN18.2 specific binding PR301839 humanized IgG1 antibody utilizing LegoChem Biosciences’ ConjuAllTM technology. LCB02A showed potent in vitro cytotoxicity in CLDN18.2 expressing gastric and pancreatic cancer cell lines. LCB02A also showed excellent anti-tumor efficacy in various gastric and pancreatic cell line-derived xenograft (CDX) models and patient-derived xenograft (PDX) models. Preliminary toxicity assessment in cynomolgus monkeys (CLDN18.2 cross-reactive species) demonstrated that LCB02A was well tolerated, with repeat doses of up to 60 mg/kg. In conclusion, LCB02A is a promising ADC for the treatment of CLDN18.2 expressing solid tumors including gastric cancer (GC) and pancreatic ductal adenocarcinoma (PDAC). umanized IgG1 antibody utilizing LegoChem Biosciences’ ConjuAllTM technology. LCB02A showed potent in vitro cytotoxicity in CLDN18.2 expressing gastric and pancreatic cancer cell lines. LCB02A also showed excellent anti-tumor efficacy in various gastric and pancreatic cell line-derived xenograft (CDX) models and patient-derived xenograft (PDX) models. Preliminary toxicity assessment in cynomolgus monkeys (CLDN18.2 cross-reactive species) demonstrated that LCB02A was well tolerated, with repeat doses of up to 60 mg/kg. In conclusion, LCB02A is a promising ADC for the treatment of CLDN18.2 expressing solid tumors including gastric cancer (GC) and pancreatic ductal adenocarcinoma (PDAC). Citation Format: Eun ji Baek, Chul-Woong Chung, Changsik Park, Taeik Jang, Yeojin Jeong, Sang-Ah Lee. LCB02A an antibody drug conjugate (ADC) targeting CLDN18.2 expressing solid tumors [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 3141.

Abstract 1230: MTX-531, a first-in-class pan-PI3K inhibitor spares hyperinsulinemia yielding durable tumor regressions and resilience to adaptive resistance

Abstract Adaptive resistance mechanisms compromise the long-term effectiveness of kinase-targeted agents dictating the need for strategic combination approaches. MTX-531 was computationally designed to selectively target both PI3K and wild type EGFR. MTX-531 exhibits low nanomolar potency against the PI3K isoform family and EGFR with a high degree of specificity predicted by co-crystal structural analyses. Full kinome screening confirmed that MTX-531 is exquisitely selective for its intended targets. We evaluated the pharmacological profile of MTX-531, whereupon single agent treatment with this lead molecule elicited a high incidence of durable tumor regressions in a broad panel of PIK3CA mutant CDX and PDX squamous head and neck (HNSCC) models. The combination of MTX-531 with RAS pathway inhibitors, including agents directed against KRAS G12C, led to durable regressions of KRAS mutant colorectal and pancreatic xenografts, resulting in striking increases in median survival. MTX-531 is exceptionally well tolerated in mice and uniquely does not lead to hyperglycemia and hyperinsulinemia commonly seen with predecessor pan PI3K inhibitors. MTX-531 acts as an agonist of PPARγ, a structural feature of the molecule that is thought to mitigate hyperglycemia induced by PI3K inhibition. This unique attribute of MTX-531 confers a favorable therapeutic index not typically seen with PI3K inhibitors. To the best of our knowledge, MTX-531 is the first reported pan-PI3K inhibitor that does not lead to hyperglycemia. The pharmaceutical profile of MTX-531 includes favorable cross-species oral bioavailability, microsomal stability, and a facile 3 step chemical synthesis. MTX-531, which has the unique capability of concurrently and selectively inhibiting PI3K and EGFR, illustrates the power of rational computational drug design to target multiple adaptive resistance mechanisms in a single molecule. The versatility of MTX-531 in both the single agent and combination settings offers a breadth of development strategies that continue to evolve as new combination candidates become available. MTX-531 is currently undergoing advanced preclinical development for anticipated first in human clinical trials in late 2024. Citation Format: Christopher E. Whitehead, Elizabeth K. Ziemke, Christy L. Frankowski-McGregor, Rachel A. Mumby, June Chung, Jinju Li, Nathaniel Osher, Oluwadara Coker, Michelle Norris, Scott Kopetz, Veera Baladandayuthapani, Melinda Hollingshead, Sharad Verma, Judith S. Sebolt-Leopold. MTX-531, a first-in-class pan-PI3K inhibitor spares hyperinsulinemia yielding durable tumor regressions and resilience to adaptive resistance [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 1230.

Abstract 6512: BPI-501836: A highly potent small molecule inhibitor targeting KRASG12D mutation

Abstract As one of the most notorious un-druggable oncogenes, mutant KRAS impairs the transition of RAS from its active GTP-binding form to the inactive GDP-binding form, leading to sustained activation of downstream MAPK pathway and ultimately tumorigenesis. The recent approved KRASG12C inhibitors have demonstrated clinical benefits in KARSG12C mutant patients. However, the other prevalent KRAS mutations, such as KRASG12D, remain a huge unmet medical challenge. There’s no approved KRASG12D specific inhibitors yet. Here we present BPI-501836, a highly potent and selective small molecular inhibitor for KRASG12D. In vitro, BPI-501836 exhibited very high potency by disrupting the KRASG12D and cRAF interaction in a cell-free assay. In cell-based assays, BPI-501836 inhibited proliferation of KRASG12D mutant cell lines with sub-nanomolar IC50 values, without affecting wide type or non-G12D KRAS mutant cell lines. Moreover, it also attenuated ERK phosphorylation in KRASG12D mutant cells. In vivo, tumor regression was observed following the administration of 6 mg/kg (iv, QW) BPI-501836 in KRASG12D human pancreatic Panc04.03 xenograft mouse models, or in the KRASG12D human pancreatic PK-59 xenograft mouse models with an intraperitoneal injection of 20 mg/kg per week. Noteworthy was the prolonged half-life of BPI-501836 observed across various species, coupled with its high distribution in tumor tissues. Moreover, sustained suppression of ERK phosphorylation in tumor tissues persisted for up to 144 hours after the last dose of BPI-501836 in both xenograft mouse models. This extended pharmacodynamic effect supports the feasibility of a once-weekly dosing schedule in clinical applications. Safety assessments conducted through a 4-week repeated infusion (intravenous, twice weekly) of BPI-501836 in beagle dog demonstrated favorable safety profiles. In conclusion, BPI-501836 emerges as a highly potent and selective KRASG12D inhibitor, exhibiting promising pharmacokinetics, pharmacodynamics, and safety profiles in pre-clinicalstudies. BPI-501836 is currently in IND-enabling stage and holds potential for further development as a therapeutic agent. Citation Format: Xiang Yang, Yuan Lu, Renqi Xu, Xiaoqiang Kong, Yongqing Sun, Xingjie Zhu, Xiaoyun Liu, Zhengyao Zou, Haibo Chen, Yanju Liu, Jialing Sun, Han Han, Cheng Yang, Jing Guo, Hong Lan, Quan Zhou, Lieming Ding, Hao Wu, Jiabing Wang. BPI-501836: A highly potent small molecule inhibitor targeting KRASG12D mutation [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 6512.

Abstract 2618: BCG017, a bispecific ADC targeting PTK7 and EGFR exhibits anti-tumor efficacy in PDX models

Abstract EGFR is a favored target in various cancer therapies. However, EGFR-based treatments have been challenged by drug resistance and cytotoxicity. Targeting EGFR with an antibody-drug conjugate (ADC) is a promising new therapeutic strategy to address the drug resistance, due to the potent killing effects of the payload. Furthermore, we hypothesized that development of a bispecific ADC (bsADC) targeting EGFR and another tumor-associated antigen (TAA) might improve the tumor selectivity, thereby limiting the on-target off-tumor toxicity. One such TAA candidate is PTK7, which belongs to the same receptor tyrosine kinase family as EGFR, and is co-expressed with EGFR in multiple types of solid tumors, including lung, esophageal, head and neck, and colon cancers. Herein, we developed a fully human anti-human PTK7 EGFR bsAb using our proprietary common light chain RenLite® mouse platform and knobs-into-holes technology. The anti-PTK7 EGFR bsAb demonstrates good purity by SEC-HPLC, and can cross-react with human and cynomolgus monkey antigens with favorable affinity. The PTK7 x EGFR bsAb demonstrated higher binding to various types of cancer cell lines, as well as enhanced internalization in vitro compared with PTK7 parental antibody. Next, the bsAb was conjugated with monomethyl auristatin E (MMAE) to generate an anti-PTK7 x EGFR bsADC, BCG017, with a drug-to-antibody ratio of approximately 4. BCG017 demonstrated superior anti-tumor activity compared to benchmarks and its PTK7 parental ADC in cell-derived non-small-cell lung cancer (NSCLC) xenografts, and showed enhanced anti-tumoractivity when compared to benchmark ADCs and parental ADCs in patient-derived breast and pancreatic ductal adenocarcinoma xenograft models. These results suggest that BCG017 have the potential to be a novel therapeutic alternative for PTK7 and EGFR co-expressing tumors. Citation Format: Sufei Yao, Chengzhang Shang, Zhuolin Li, Gao An, Chaoshe Guo, W. Frank An, Yi Yang. BCG017, a bispecific ADC targeting PTK7 and EGFR exhibits anti-tumor efficacy in PDX models [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 2618.

Abstract 6518: BPM 31510: Targeting the tumor microenvironment (TME) via mitochondrial-mediated ROS production

Abstract The tumor microenvironment (TME) consists of an enriched population of immune cells and milieu of factors that modulate key cellular processes such as angiogenesis, metabolism, apoptosis, migration, and proliferation. Previous studies have suggested that BPM 31510 induces ROS through high intra-cellular levels of ubidecarenone in aggressive cancer phenotypes both in vitro and in vivo through induction of mitochondrial ROS. Furthermore, we have previously observed efficacy of BPM 31510 in a highly aggressive in vivo orthotopic rat C6 glioma model where BPM 31510 was administered in combination with radiotherapy. In a pancreatic xenograft mouse model, we previously reported that BPM 31510 impacted the immune cells in the TME by increasing absolute number of infiltrated CD8+ T-cells and decreasing absolute number of infiltrated CD4+ T-cells. Moreover, an ex vivo study of PBMC’s from healthy volunteers suggests that BPM 31510 increases T-cell viability and function while also decreasing T-cell exhaustion. Currently, an ongoing first-line phase 2b study of BPM 31510 in glioblastoma in combination with radiotherapy and temozolomide will assess the clinical efficacy and potential for mechanistic synergy of multiple ROS-inducing mitochondrial mediated regimens on this aggressive cancer phenotype. BPM 31510 will be assessed in this study to provide further evidence for the impact of ROS on immunomodulation. Taken together, BPM 31510 increases mitochondrial ROS/OXPHOS in cancer cells while impacting TME-related immune cell modulation. Citation Format: Stephane Gesta, Maria D. Nastke, Seema Nagpal, Lawrence Recht, Michael A. Kiebish, Niven R. Narain, Vijay R. Modur. BPM 31510: Targeting the tumor microenvironment (TME) via mitochondrial-mediated ROS production [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 6518.

Platinum iodido drugs show potential anti-tumor activity, affecting cancer cell metabolism and inducing ROS and senescence in gastrointestinal cancer cells

Cisplatin-based chemotherapy has associated clinical disadvantages, such as high toxicity and resistance. Thus, the development of new antitumor metallodrugs able to overcome different clinical barriers is a public healthcare priority. Here, we studied the mechanism of action of the isomers trans and cis-[PtI2(isopropylamine)2] (I5 and I6, respectively) against gastrointestinal cancer cells. We demonstrate that I5 and I6 modulate mitochondrial metabolism, decreasing OXPHOS activity and negatively affecting ATP-linked oxygen consumption rate. Consequently, I5 and I6 generated Reactive Oxygen Species (ROS), provoking oxidative damage and eventually the induction of senescence. Thus, herein we propose a loop with three interconnected processes modulated by these iodido agents: (i) mitochondrial dysfunction and metabolic disruptions; (ii) ROS generation and oxidative damage; and (iii) cellular senescence. Functionally, I5 reduces cancer cell clonogenicity and tumor growth in a pancreatic xenograft model without systemic toxicity, highlighting a potential anticancer complex that warrants additional pre-clinical studies.