Abstract Introduction: Prostate-specific membrane antigen (PSMA)-targeted radioligand therapy (RLT) with lutetium-177 (177Lu) has demonstrated clinical benefit in prostate cancer (PC). Although alpha-emitters offer superior cytotoxic efficacy, the complex toxicity profile of actinium-225 (225Ac)-PSMA limits its therapeutic application. Lead-212 (212Pb) possesses favorable physical properties, including high linear energy transfer, a 10.6-hour half-life, and a simple decay scheme with a single alpha-emitting daughter nuclide, enabling precise, potent radiation delivery at the cellular level. In this study, we combine in vitro and in vivo models with multiomics and functional assays to elucidate the mechanisms underlying the anti-tumor efficacy of PSMA-targeted RLT. Materials and Methods: 212Pb-ADVC001, a novel 212Pb-based PSMA-targeting RLT in Phase I/II clinical development (NCT05720130) for the treatment of metastatic PC, was used as the 212Pb-PSMA-RLT agent. 177Lu-PSMA-I&T was used as the 177Lu-PSMA-RLT agent. The kinetic mechanisms associated with 212Pb-ADVC001 mediated cell-death were investigated in vitro and ex vivo using transcriptomics and proteomics analyses. To validate the omics findings, functional assays were conducted to assess the effects of 212Pb-PSMA on PC’s cell cycle progression, reactive oxygen species (ROS) generation, and lipid peroxidation in PC cells. Results: 212Pb-ADVC001 displayed potent cytotoxic activity with a mean EC50 of 2.7, 7.2 and 3.3 kBq/mL in PC cell lines PC-3-PIP (PSMAhigh), C4-2 (PSMAint) and LNCaP (PSMAint), respectively. Transcriptomic and proteomic analysis of in vitro treated PC cells and in vivo tumors harvested longitudinally revealed multiple mechanisms of action involving DNA damage, cell cycle arrest and cell death, and immune response modulation. In vitro functional assays confirmed the involvement of ROS production, lipid peroxidation, and cell cycle arrest upon treatment with 212Pb-ADVC001. Particularly, 212Pb-ADVC001 caused significant DNA damage, and reduced DNA content in the S-phase of the cell cycle with a concomitant increase in the G1- and G2/M-phase arrest compared to 177Lu-PSMA-I&T (p < 0.05). Conclusion: Multiomics analyses highlighted the involvement of multiple mechanisms of action in the efficacy of 212Pb-ADVC001 which collectively resulted in effective cancer cell death. Mechanistic studies furthered the understanding of PC radiobiology and cellular responses to beta- and alpha-based RLT with the identification of a cell cycle arrest in PC specifically induced by 212Pb-PSMA. Citation Format: Feifei Liu, Melissa Monterosso, Didier Boucher, Anna Amiss, Stelle Shakti, Kwong Ching Li, Chanwoo Kim, Aimee Horsfall, Kevin Kuan, William Tieu, Stephen Rose, Simon Puttick, Joana Brilhante, Gary Li, Anna Karmann, Thomas Kryza. Mechanistic insights into the anti-tumor activity of 212Pb-PSMA radioligand therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 5275.

Introduction ADVC001 is a patented PSMA-targeting radioligand labelled with 212Pb, an alpha-emitting payload with a short half-life of 10.6 hrs, designed to deliver radiation precisely on a cellular level. Objectives We present results from the Phase 1b part of the TheraPb trial (NCT05720130) evaluating 212Pb-ADVC001 in patients (pts) with metastatic prostate cancer. The feasibility of positron emission tomography (PET) imaging of 212Pb in phantoms and in pts receivin Materials and Methods Open-label, multi-center interval 3+3 dose-escalation and expansion study in pts with progressive PSMA-avid (> liver) mCRPC previously treated with ≥ 1 taxane unless unsuitable/declined and ≥ 1 androgen pathway inhibitor (ARPi). Escalating doses of 60–200 MBq were administered at prespecified intervals every 6, 4, 2 and 1 week(s) for up to 6 cycles in an outpatient setting with minimal restrictions at discharge 3 hours post administration. Primary endpoints were safety and defining the recommended Phase 2 dose and administration schedule.212Pb-ADCV001 point sources in glass capillary tubes in a water phantom were imaged to determine the system spatial resolution, full width at half maximum (FWHM). An IEC phantom was imaged with 98 kBq/mL in the spheres, zero background. Ethics approval for exploratory PET imaging was received and informed consent was obtained by pts.. PET acquisitions were performed using two bed positions, with 15 minutes per bed position, covering the thoracic and abdominal regions including the kidneys, using a Siemens Vision scanner. Results As of the 2 October 2025 data cut-off, dose escalation was completed (n=22). 73% of pts had ≥ 1 prior taxane, 36% had ≥ 2 prior lines of chemotherapy, and all had ≥ 1 prior ARPi. There were no dose-limiting toxicities, dose modifications or treatment discontinuations due to treatment-related adverse events (TRAEs); there were no Gr 4/5 TRAEs, signs of myelosuppression or radiation adverse events of special interest. Xerostomia was predominantly Gr 1, with evidence of reversibility. A PSA50 response was observed in 80% of pts treated at doses ≥ 160 MBq, and a 100% overall response rate (6/6) in pts with RECIST-measurable target lesion(s), including two complete responses.The FWHM of 212Pb-ADVC001 was 4.9 mm and the IEC phantom imaging allowed visualization of the 13 mm spheres, with the recorded count linear with radioactivity. Clinical imaging of 212Pb-ADVC001 PET pair production is feasible. PET maximum intensity projection images (MIP) provide a clear localization of the alpha decay, hence 208Tl and 212Bi, in lesions concordant with diagnostic PSMA PET imaging. Conclusion 212Pb-ADVC001 demonstrated encouraging safety and promising anti-tumor activity in pts with progressive mCRPC. Clinical 212Pb-ADVC001 PET imaging of 208Tl gamma pair production confirmed the alpha emission from 212Pb-ADVC001 within PSMA-avid lesions.Phase 2 expansion will assess 212Pb-ADVC001 with novel dosing strategies to optimize clinical outcomes in 3 indications across mCRPC and metastatic hormone-sensitive prostate cancer. Funding Acknowledgements AdvanCell Pty Ltd

Prostate cancer (PC) is the second leading cause of cancer-related death in men, with metastatic castration-resistant PC (mCRPC) remaining largely incurable. Prostate-specific membrane antigen (PSMA) is a validated target, with PSMA-directed radioligand therapy demonstrating survival benefit in mCRPC. Targeted alpha therapies (TAT) with isotopes Lead-212 (212Pb) and Actinium-225 (225Ac) are in clinical development and deliver high-linear-energy-transfer radiation, inducing potent cytotoxicity and potentially enhancing anti-tumour immunity. Current PSMA-positive preclinical models rely on xenografts or engineered murine cells, limiting evaluation of TAT-induced immune modulation thus far. To investigate the immune-modulatory effects of 212Pb-TAT in metastatic PC, we sought to establish (i) a syngeneic murine model of metastatic PC expressing murine PSMA (mPSMA), and (ii) a PSMA-directed TAT tool compound with high affinity for mPSMA. Murine metastatic PC cells (RM1-BM), which recapitulate late-stage TP53-mutant disease, were engineered to express murine FOLH1 (mPSMA) using a PiggyBac transposon system. Expression of mPSMA was confirmed both in vitro and in vivo by western blotting and immunohistochemistry. The TAT agent 212Pb-PS0001 (AdvanCell) was assessed in biodistribution and efficacy studies in immunocompetent C57Bl/6 mice. RM1-BM-mFolh1 cells expressed mPSMA comparably to human PSMA-positive C4-2 cells. 212Pb-PS0001 showed selective tumour uptake in RM1-BM-mFolh1 models, with minimal off-target accumulation. Therapeutic evaluation demonstrated that a single dose of 212Pb-PS0001 significantly suppressed RM1-BM-mFolh1 tumour growth in immunocompetent mice, while untreated controls exhibited progressive disease. Notably, several animals achieved durable tumour control, suggesting potential activation of an anti-tumour immune response. We established a syngeneic mPSMA-positive metastatic PC model suitable for studying the immune-modulatory effects of PSMA-targeted therapies. This model represents a robust platform for assessing therapeutic efficacy, elucidating immune mechanisms of action, and evaluating rational combination strategies, including integration with immune checkpoint blockade. Preliminary studies with 212Pb-PS0001 demonstrated selective PSMA targeting, rapid clearance, and potent anti-tumour activity, supporting its use as PSMA-TAT tool compound. Melissa Monterosso, Aneesha Jones, Kayden Kwah, Anna Amiss, Abby Sydes, Didier Boucher, Aimee Horsfall, Heather Green, Johannes Koehbach, Ralph Huebner, Yaowu He, Stephen Rose, Gary Li, Feifei Liu, Simon Puttick, Anna Karmann, John Hooper, Thomas Kryza. Development of an immunocompetent murine model of PSMA-positive metastatic prostate cancer to study the impact of PSMA-targeted therapies on the immune response [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Innovations in Prostate Cancer Research and Treatment; 2026 Jan 20-22; Philadelphia PA. Philadelphia (PA): AACR; Cancer Res 2026;86(2_Suppl):Abstract nr B040.

Abstract Introduction: Prostate cancer (PC) is the second leading cause of cancer-related death in men, and metastatic castration-resistant prostate cancer (mCRPC) remains incurable. Prostate-specific membrane antigen (PSMA) is a validated therapeutic target. Targeted alpha therapies (TAT) employing radionuclides such as lead-212 (212Pb) and actinium-225 (225Ac) are in development, delivering high-linear-energy-transfer radiation with potent cytotoxic and potentially immunomodulatory effects. However, currently available PSMA-positive preclinical models rely on human xenografts, which do not allow for the study of radioligand therapy-induced immune modulation. Objective: The immunomodulatory effects of PSMA-targeted alpha therapy are believed to play a key role in the anti-tumor efficacy towards PC. However, the lack of immunocompetent in vivo models of PSMA-positive PC has been a major hurdle in mechanistic and translational studies. To overcome this, we established a syngeneic prostate cancer model expressing murine PSMA (mPSMA) and developed a high-affinity PSMA-directed TAT tool compound specific to mPSMA. Methodology: Murine bone-metastatic prostate cancer cell line, RM1-BM, which recapitulates late-stage TP53-mutant disease, was engineered to express murine FOLH1 (mPSMA) using the PiggyBac transposon system. Expression of mPSMA was confirmed both in vitro and in vivo by western blotting and immunohistochemistry. The affinity and specificity of the TAT agent 212Pb-PS0001 (AdvanCell) for murine and human PSMA were assessed using inhibition and cytotoxicity assays. Biodistribution and therapeutic efficacy were assessed in immunocompetent C57BL/6 mice bearing mPSMA-RM1-BM tumors. Blood and tissue samples collected from treated animals underwent multiomics analyses to characterize immune response modulation induced by 212Pb-PS0001. Results: RM1-BM-mFOLH1 cells expressed mPSMA at levels comparable to human PSMA-positive C4-2 cells. 212Pb-PS0001 exhibited high-affinity, selective binding and robust tumor uptake in RM1-BM-mFOLH1 models, with minimal off-target accumulation. Therapeutic evaluation demonstrated that a single administration of 212Pb-PS0001 significantly inhibited RM1-BM-mFOLH1 tumor growth in immunocompetent mice, whereas untreated controls showed progressive disease. Notably, several treated animals achieved durable tumor control, suggesting potential induction of an anti-tumor immune response. Conclusion: By establishing a syngeneic, mPSMA-positive metastatic prostate cancer model, we enabled the evaluation of PSMA-TAT in an immunocompetent setting. Preliminary studies with 212Pb-PS0001 demonstrated selective PSMA targeting, favorable pharmacokinetics, and potent anti-tumor efficacy, supporting its utility as a PSMA-TAT tool compound for investigating immune-mediated mechanisms of action in prostate cancer. Citation Format: Melissa Monterosso, Aneesha Jones, Kayden Kwah, Anna Amiss, Didier Boucher, Aimee Horsfall, Heather Green, Johannes Koehbach, Ralph Huebner, Yaowu He, Stephen Rose, Gary Li, Feifei Liu, Joana Brilhante, Simon Puttick, Anna Karmann, John Hooper, Thomas Kryza. Evaluation of 212Pb-PSMA radioligand therapy in an immunocompetent prostate cancer model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 2855.

The biodistributions of radiometalated peptides and small molecules are greatly influenced by the charge conferred by the metal-chelator complex. Careful fine-tuning of this charge thus represents an attractive method to optimise pharmacokinetic properties. For this to be an effective strategy, numerous suitable chelators must be available for a given radiometal; each possessing a different net charge. Herein we report the synthesis of 2-(4,7,10-tris(2-amino-2-oxoethyl)-1,4,7,10-tetraazacyclododecan-1-yl)pentanedioic acid (DOTAMGA), a bifunctional chelator for the 203/212Pb theranostic pair containing a single negative charge, previously unattainable with reported radiolead chelators. DOTAMGA was incorporated into a radiopharmaceutical targeting the melanocortin 1 receptor (MC1R), a receptor highly expressed by melanomas, and evaluated in vivo against the commonly used radiolead chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic amide (TCMC). DOTAMGA was synthesised and conjugated to the MC1R targeting peptide MC1RL for subsequent investigation alongside TCMC-MC1RL. DOTAMGA-MC1RL and TCMC-MC1RL exhibited comparable affinity for MC1R in a series of competition binding assays with MC1R expressing cells. DOTAMGA-MC1RL was effectively labelled with 212Pb and 203Pb under standard radiolabelling conditions, and UV–Vis experiments demonstrated more rapid Pb2+ complexation than TCMC-MC1RL. Both 212Pb-labelled compounds remained > 90

BACKGROUND & AIMS:The contribution of common genetic polymorphisms to ulcerative colitis (UC) pathogenesis is modest; however, families with severe colitis may harbor rare variants with large effect sizes that highlight unrecognized pathways. METHODS:A multigenerational family with UC necessitating colectomy was identified. Whole exome sequencing of this kindred was performed, implicating a rare variant in OTUD3. Constitutive knock-out and intestinal specific Otud3 deficient and heterozygous mice were generated. OTUD3 expression in human colonic biopsies and intestinal organoids was assessed using quantitative reverse transcription polymerase chain reaction and immunofluorescence. Prevalence of rare, damaging variants were compared in distinct patient cohorts. Plasmids containing OTUD3 missense variants were introduced into cell lines where OTUD3 was disrupted to determine their effects on cellular response to cytokine stimulation. RESULTS:Constitutive disruption or heterozygosity of Otud3 in mice, or intestinal-specific deletion, resulted in impaired barrier integrity, tight-junction dysregulation, increased endoplasmic reticulum stress, and penetration of luminal bacteria deep into the colonic crypts that preceded a spontaneous progressive colitis. Analysis of distinct UC cohorts demonstrated enrichment of rare, damaging variants in OTUD3. Introduction of OTUD3 variants in intestinal cell lines phenocopied the epithelial immune dysregulation observed in knockout mice. Finally, OTUD3 mRNA and epithelial protein expression were decreased in the quiescent colonic epithelial tissue of genotype-unselected individuals with UC compared with matched non-UC controls. CONCLUSIONS:Our results demonstrate that OTUD3 is required for colonic epithelial barrier function, and plays a role in the pathogenesis of UC.

Prostate-specific membrane antigen (PSMA)-directed radiopharmaceutical therapies continue to improve treatment outcomes in patients with metastatic castration-resistant prostate cancer. Here, we report the in vitro and in vivo characterization of a PSMA-targeted therapy (ADVC001) specifically designed for targeted α-therapy with 212Pb. Methods: The binding affinity to PSMA was determined by PSMA enzymatic assays and by radioligand binding assays using PSMA-high prostate cancer (PC) cells. In vitro cytotoxicity against PC cell lines with high and medium PSMA expression was evaluated using clonogenic, metabolic, and imaging-based cytotoxic assays. Pharmacokinetics and biodistribution were assessed using PSMA-high subcutaneous tumor xenografts. In vivo single-dose and multidose efficacy was assessed in subcutaneous PC xenograft models expressing various levels of PSMA. Results: A high binding affinity to PSMA was observed for ADVC001 with nanomolar inhibitory concentration of 50% values. In cellular assays, [212Pb]Pb-ADVC001 (212Pb-ADVC001 hereafter for simplicity) exhibited specific cytotoxic activity against PSMA-expressing cells with nanomolar effective concentration of 50% values. In vivo biodistribution of 212Pb-ADVC001 in the PC3-PIP xenograft model revealed rapid and persistent tumor uptake, fast renal clearance, and low retention in normal tissues. Single-dose efficacy studies of 212Pb-ADVC001 (0.46 MBq) showed improved survival compared with [177Lu]Lu-PSMA-I&T (177Lu-PSMA-I&T hereafter for simplicity) (20 MBq) treatment. In a multidose experiment, 2 doses of 212Pb-ADVC001 (0.5 MBq) significantly increased median survival (86 d vs. 45.5 d, P < 0.05) compared with 2 doses of 177Lu-PSMA-I&T (15 MBq). Treatment with 212Pb-ADVC001 (0.5 MBq) after initial 177Lu-PSMA-I&T (15 MBq) relapse showed an enhanced survival benefit (59.5 d). In a C4-2 xenograft model with medium-level PSMA expression, single doses of 0.3, 0.8, and 1.1 MBq of 212Pb-ADVC001 significantly extended median survival to 34, 57, and 62.5 d, compared with untreated cohorts (16 d). All treatments were well tolerated. Conclusion: The preclinical results support the clinical development of 212Pb-ADVC001 as a targeted α-therapy for the treatment of patients with PC.

PURPOSE:Receptor CUB domain-containing protein 1 (CDCP1) was evaluated as a target for detection and treatment of breast cancer. EXPERIMENTAL DESIGN:CDCP1 expression was assessed immunohistochemically in tumors from 423 patients [119 triple-negative breast cancer (TNBC); 75 HER2+; and 229 ER+/HER2-, including 228 primary tumors and 229 lymph node and 47 distant metastases). Cell cytotoxicity induced in vitro by a CDCP1-targeting antibody-drug conjugate (ADC), consisting of the human/mouse chimeric antibody ch10D7 and the microtubule disruptor monomethyl auristatin E (MMAE), was quantified, including in combination with HER-targeting ADC trastuzumab emtansine (T-DM1). Detection of CDCP1-expressing primary and metastatic xenografts in mice was examined by PET-CT imaging using zirconium-89-labeled ch10D7. The impact of ch10D7-MMAE on tumor burden and survival in vivo, including in combination with T-DM1, was quantified in cell line and patient-derived xenograft mouse models. RESULTS:CDCP1 is expressed predominantly on the surface of malignant cells of 70% of TNBC, 80% of HER2+ tumors, and increases in ER+/HER2- tumors from 44.9% in primary tumors to 56.4% in lymph node metastases and 74.3% in distant metastases. PET-CT imaging with zirconium-89-labeled ch10D7 is effective for the detection of primary and metastatic CDCP1-expressing TNBC in mice. ADC ch10D7-MMAE kills CDCP1-expressing cells in vitro and controls primary and metastatic TNBC xenografts in mice, conferring significant survival advantages over chemotherapy. It compares favorably to T-DM1 in vivo, and ch10D7-MMAE combined with T-DM1 showed the most potent efficacy, markedly reducing tumor burden of CDCP1+/HER2+ xenografts and prolonging mouse survival, compared with T-DM1 or ch10D7. CONCLUSIONS:CDCP1-directed molecular imaging has the potential to identify aggressive breast cancers for CDCP1-targeted treatment.

TPS275 Background: Prostate cancer (PC) is the second most common cancer and second leading cause of cancer-related death in men. A recent Lancet Commissionarticle predicts a rise in PC cases from 1.4 million in 2020 to 2.9 million by 2040. Despite ten new therapies being available for metastatic PC in the recent decade, the survival advantage from most agents in the setting of metastatic castration-resistant prostate cancer (mCRPC) is measured in months largely due to cross-resistance, and the 5-year survival is approximately 32%. The radioligand therapy 177 Lu-PSMA-617, which uses a beta-emitting isotope linked to a PSMA-targeting small molecule, has shown a median survival advantage in mCRPC of four months, underpinning the critical need for more effective treatments. Excitement has shifted towards the use of alpha-emitting isotopes that deliver a more lethal payload. Unlike beta particles, alpha particles have a short range of tissue penetration (< 0.1 mm) and high linear energy transfer that induces double-stranded DNA breaks and high levels of cytotoxicity to target expressing cancer cells irrespective of cell cycle or oxygenation state. Herein we describe the ongoing clinical trial of [ 212 Pb]Pb-ADVC001 – a novel PSMA-targeted radioligand labelled with the potent alpha-emitting isotope 212 Pb. Methods: This is a prospective, open-label, non-randomized, dose-escalation, dose optimization and expansion study. The study aims to determine the safety and tolerability of escalating doses of [ 212 Pb]Pb-ADVC001 administered every 6, 4 or 2 weeks during the dose-finding phase (Phase 1b) in participants with PSMA-positive mCRPC who have had exposure to at least one androgen receptor pathway inhibitor (ARPi) and taxane-based chemotherapy at any time in the course of their disease. The dose escalation phase will follow an i3+3 design, with each cohort able to backfill up to 20 participants for further characterization of the safety, tolerability and preliminary efficacy of dose and schedules. The expansion phase (Phase 2a) aims to assess the efficacy, safety and tolerability of [ 212 Pb]Pb-ADVC001 at the recommended Phase 2 dose in three groups of participants with PSMA-positive mCRPC. Group 1: Participants who have had exposure to at least one ARPi and have not received a taxane for the treatment of mCRPC. Group 2: Participants who have had exposure to at least one ARPi and received taxane-based chemotherapy for the treatment of mCRPC. Group 3: Participants who have had exposure to 177 Lu-PSMA. If data from Phase 1b demonstrates multiple dosing regimens are reasonably equivalent in terms of safety, tolerability and anti-tumor activity, the Phase 2a expansion study may include adaptive randomization to identify an optimized dose and schedule. The trial is currently enrolling at two clinical sites in Australia. Clinical trial information: NCT05720130 .

177Lu-PSMA-617 is the first FDA-approved targeted radioligand therapy (RLT), and in clinical trials, significantly prolonged survival and improved quality of life in patients with metastatic castration resistant prostate cancer (mCRPC). However, approximately 30% of the patients do not respond to the treatment and most initial responders ultimately relapse. Compared to the beta-emitter 177Lu, the alpha therapy isotope 212Pb may prove to be advantageous due to its short half-life, high linear energy transfer, and efficient induction of immunogenic cell death. Here we used in vitro and in vivo models to elucidate the intrinsic cellular mechanisms of response to 177Lu-PSMA and 212Pb-PSMA treatment. 177Lu-PSMA-I&T and 212Pb-ADVC001 were used as PSMA-targeted 177Lu- and 212Pb-RLT, respectively. The cytotoxic activity in prostate cancer (PC) cell lines was measured by a cell viability assay. In vivo studies were performed in human PC-bearing mice to compare preclinical efficacy of 177Lu- and 212Pb-PSMA therapy. The kinetic mechanisms associated with 177Lu-PSMA-I&T and 212Pb-ADVC001 mediated cell-death were assessed ex vivo using proteomics, and their impact on the PC cell cycle by flow cytometry. 212Pb-ADVC001 displayed potent cytotoxic activity with an average EC50 of 2.7, 7.2 and 3.3 kBq/mL in PC cell lines PC-3-PIP (PSMAhigh), C4-2 (PSMAint) and LNCaP (PSMAint), respectively, compared to 177Lu-PSMA-I&T (430.7, 434.9 and 814.4 kBq/mL, respectively). In PSMAhigh xenograft models, 212Pb-ADVC001 (0.43 MBq) significantly improved median survival to 84 days compared to 48 days for 177Lu-PSMA I&T (20 MBq). Multidose efficacy studies demonstrated superior tumor response with two cycles of 212Pb-ADVC001 (0.46 MBq) compared to two cycles of 177Lu-PSMA I&T (15 MBq). 212Pb-ADVC001 administration in 177Lu-PSMA I&T-relapsed mice resulted in improved survival. In PSMAint xenograft models, 212Pb-ADVC001 (0.9 MBq) also suppressed tumor progression more effectively than 177Lu-PSMA-I&T (30 MBq), resulting in prolonged survival. Proteomics analysis of RLT-treated tumors revealed a specific impact of 212Pb-ADVC001 on cell cycle checkpoints and DNA replication. In vitro cell cycle experiment further confirmed that 212Pb-ADVC001 caused significant DNA damage, and reduced DNA content in the S-phase of the cell cycle with a concomitant increase in the G1 and G2/M-phase arrest compared to 177Lu-PSMA-I&T (p<0.05). Comparison studies of 177Lu- and 212Pb-PSMA treatment on PC cells showed a higher efficacy of 212Pb-PSMA in vitro and in vivo supporting the clinical application of 212Pb-PSMA for mCRPC. Mechanistic studies furthered the understanding of PC radiobiology and cellular responses to beta- and alpha-RLT with the identification of a cell cycle arrest in PC specifically induced by 212Pb-PSMA. Feifei Liu, Melissa E. Monterosso, Didier Boucher, Stelle Shakti, Kwong Ching Li, Aimee Horsfall, Saawan Kumar, Johannes Koehbach, Kevin Kuan, William Tieu, Stephen Rose, Anna Karmann, Simon Puttick, Gary Li, Thomas Kryza. Uncovering the mechanisms of action of 177Lu- and 212Pb- based PSMA radioligand therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1333.

PSMA-targeted radioligand therapy (RLT) with lutetium-177 (177Lu), a beta-emitter, has been clinically proven to be efficacious for the treatment of prostate cancer (PC). Although emerging clinical data indicate that better efficacy can be achieved by alpha-emitters, the toxicities associated with actinium-225 (225Ac) may limit its broad application especially in early lines of treatment. Lead-212 (212Pb), is a promising isotope for alpha therapy with a high linear energy transfer, a half-life of 10.6 hours, and a simple decay scheme. These properties promote efficient targeting of PSMA+ PC cells while limiting off-target toxicities. Pb-ADVC001 is a novel Pb-based PSMA-targeted RLT currently in Phase Ib/IIa clinical trial (NCT05720130) for patients with metastatic castration-resistant prostate cancer (mCRPC). The affinity of ADVC001 and Pb-ADVC001 to PSMA was determined by SPR and radioligand-binding assays in PSMA+ PC cell lines. Cytotoxic activity was measured by cell viability and clonogenic assays. In vivo biodistribution and anti-tumor efficacy studies were performed in human-derived PC-bearing mice. The intrinsic molecular and cellular mechanisms associated with Pb-ADVC001 mediated cell-death were assessed using transcriptomics and proteomics. ADVC001 binds human, mouse and rat PSMA with comparable nanomolar activities, and efficiently internalizes upon binding to PSMA+ PC cells. In vitro, Pb-ADVC001 showed specific and potent cytotoxic activity against multiple PSMA+ human PC cell lines, while sparing PSMA- cells. In vivo biodistribution of Pb-ADVC001 showed tumor uptake at 19.6, 20.9, 9.4 and 5.4 %ID/g at 1, 3, 24 and 48 h, respectively, with retention out to 72 h. High tumor uptake and fast renal clearance resulted in favorable tumor/kidney ratios. Minimal uptake was observed in other normal tissues. Single administration of Pb-ADVC001 in PSMA+ PC xenografts led to dose-dependent anti-tumor response and significant survival benefit. Transcriptomic analysis of in vitro treated PC cells and proteomic analysis of tumors harvested longitudinally revealed multiple mechanisms of action involving DNA damage, cell cycle arrest and cell death, and immune response modulation. Preclinical data demonstrated the potential of Pb-ADVC001 for the treatment of patients with PSMA+ prostate cancer. Transcriptomic and proteomic analyses indicate multiple mechanisms of action collectively resulting in effective cancer therapy. Melissa E. Monterosso, Feifei Liu, Didier Boucher, Stelle Shakti, Kwong Ching Li, Aimee Horsfall, Saawan Kumar, Johannes Koehbach, Kevin Kuan, William Tieu, Stephen Rose, Anna Karmann, Gary Li, Simon Puttick, Thomas Kryza. Preclinical evaluation of a novel Pb-based, PSMA-targeted radioligand, Pb-ADVC001, for prostate cancer treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 588.

Background 5-Fluorouracil (5-FU) remains a core component of systemic therapy for colorectal cancer (CRC). However, response rates remain low, and development of therapy resistance is a primary issue. Combinatorial strategies employing a second agent to augment the therapeutic effect of chemotherapy is predicted to reduce the incidence of treatment resistance and increase the durability of response to therapy. Methods Here, we employed quantitative proteomics approaches to identify novel druggable proteins and molecular pathways that are deregulated in response to 5-FU, which might serve as targets to improve sensitivity to chemotherapy. Drug combinations were evaluated using 2D and 3D CRC cell line models and an ex vivo culture model of a patient-derived tumour. Results Quantitative proteomics identified upregulation of the mitosis-associated protein Aurora B (AURKB), within a network of upregulated proteins, in response to a 24 h 5-FU treatment. In CRC cell lines, AURKB inhibition with the dihydrogen phosphate prodrug AZD1152, markedly improved the potency of 5-FU in 2D and 3D in vitro CRC models. Sequential treatment with 5-FU then AZD1152 also enhanced the response of a patient-derived CRC cells to 5-FU in ex vivo cultures. Conclusions AURKB inhibition may be a rational approach to augment the effectiveness of 5-FU chemotherapy in CRC.

There is significant interest in the development of 212Pb-PSMA–based targeted α-therapy for patients with metastatic castration-resistant prostate cancer. A previous phantom study has shown that 212Pb SPECT is feasible by imaging the 238.6 keV and 75 to 91 keV γ-emissions produced after the β-

Genetic variation at the 19q13.3 KLK locus is linked with prostate cancer susceptibility in men. The non-synonymous KLK3 single nucleotide polymorphism (SNP), rs17632542 (c.536T>C; Ile163Thr-substitution in PSA) is associated with reduced prostate cancer risk, however, the functional relevance is unknown. Here, we identify that the SNP variant-induced change in PSA biochemical activity mediates prostate cancer pathogenesis. The 'Thr' PSA variant leads to small subcutaneous tumours, supporting reduced prostate cancer risk. However, 'Thr' PSA also displays higher metastatic potential with pronounced osteolytic activity in an experimental metastasis in-vivo model. Biochemical characterisation of this PSA variant demonstrates markedly reduced proteolytic activity that correlates with differences in in-vivo tumour burden. The SNP is associated with increased risk for aggressive disease and prostate cancer-specific mortality in three independent cohorts, highlighting its critical function in mediating metastasis. Carriers of this SNP allele have reduced serum total PSA and a higher free/total PSA ratio that could contribute to late biopsy decisions and delay in diagnosis. Our results provide a molecular explanation for the prominent 19q13.3 KLK locus, rs17632542 SNP, association with a spectrum of prostate cancer clinical outcomes. The PSA (KLK3) genetic variant rs17632542 is associated with reduced prostate cancer risk and lower serum PSA levels, although the underlying reasons are unclear. Here, the authors show that this PSA variant reduced proteolytic activity and leads to smaller tumours, but also increases invasion and bone metastasis, indicating its dual risk association depending on tumour context; the variant is associated with both lower risk and poor clinical outcomes.