Antitumor Modalities Research Articles

GSH-responsive polymeric micelles-based augmented photoimmunotherapy synergized with PD-1 blockade for eliciting robust antitumor immunity against colon tumor

Phototherapy is a promising antitumor modality, which consists of photothermal therapy (PTT) and photodynamic therapy (PDT). However, the efficacy of phototherapy is dramatically hampered by local hypoxia in tumors, overexpression of indoleamine 2,3-dioxygenase (IDO) and programmed cell death ligand-1 (PD-L1) on tumor cells. To address these issues, self-assembled multifunctional polymeric micelles (RIMNA) were developed to co-deliver photosensitizer indocyanine green (ICG), oxygenator MnO2, IDO inhibitor NLG919, and toll-like receptor 4 agonist monophosphoryl lipid A (MPLA). It is worth noting that RIMNA polymeric micelles had good stability, uniform morphology, superior biocompatibility, and intensified PTT/PDT effect. What’s more, RIMNA-mediated IDO inhibition combined with programmed death receptor-1 (PD-1)/PD-L1 blockade considerably improved immunosuppression and promoted immune activation. RIMNA-based photoimmunotherapy synergized with PD-1 antibody could remarkably inhibit primary tumor proliferation, as well as stimulate the immunity to greatly suppress lung metastasis and distant tumor growth. This study offers an efficient method to reinforce the efficacy of phototherapy and alleviate immunosuppression, thereby bringing clinical benefits to cancer treatment.Graphical

Multimodal enhancement of ferroptosis for synergistic cascade colorectal cancer therapy

Emerging research positions ferroptosis as a formidable anti-tumor modality for colorectal cancer (CRC). However, the current pathways to potentiate ferroptosis through a singular or dual approach have fallen short in maximizing treatment efficacy. Herein, a multimodal enhancement strategy of ferroptosis is achieved for synergistic cascade colorectal cancer therapy, via clinical-related drug repurposing and prodrug self-assembly of SN38 and sulfasalazine (SAS). In this process, the SN38 or SAS prodrug is synthesized by conjugating ferrocene via disulfide bonds to SN38 or SAS, then the two prodrugs self-assemble into a hybrid nanomedicine (FSS). FSS exhibited pronounced stability under physiological conditions, while demonstrating rapid disassembly via glutathione-responsive break of disulfide bonds in tumor microenvironments. Then, a multimodal enhancement strategy of cancer ferroptosis is achieved via the SAS-induced regulation of amino acid metabolism, SN38-resulted regulation of glutathione synthesis, accumulation of ferroptosis-related lipid signaling, and ferrocene-directed iron overload in tumors. The potent ferroptosis-based CRC anticancer effects of FSS were systematically explored and confirmed by comprehensive in vitro and in vivo analyses, including cellular experiments, subcutaneous and orthotopic tumor-bearing mouse models, and RNA-sequencing transcriptomic studies. This study lays the groundwork for the clinical translation of multimodal ferroptosis inducers, offering an effective and safe approach to cancer treatment.

Nanocatalytic NO gas therapy against orthotopic oral squamous cell carcinoma by single iron atomic nanocatalysts

ABSTRACT Oral squamous cell carcinoma (OSCC) has been being one of the most malignant carcinomas featuring high metastatic and recurrence rates. The current OSCC treatment modalities in clinics severely deteriorate the quality of life of patients due to the impaired oral and maxillofacial functions. In the present work, we have engineered the single-atom Fe nanocatalysts (SAF NCs) with a NO donor (S-nitrosothiol, SNO) via surface modification to achieve synergistic nanocatalytic NO gas therapy against orthotopic OSCC. Upon near-infrared laser irradiation, the photonic hyperthermia could effectively augment the heterogeneous Fenton catalytic activity, meanwhile trigger the thermal decomposition of the engineered NO donor, thus producing toxic hydroxyl radicals (•OH) and antitumor therapeutic NO gas at tumor lesion simultaneously, and consequently inducing the apoptotic cell death of tumors via mitochondrial apoptosis pathway. This therapeutic paradigm presents an effective local OSCC therapeutics in a synergistic manner based on the nanocatalytic NO gas therapy, providing a promising antitumor modality with high biocompatibility.

P53-Armed Oncolytic Virotherapy Improves Radiosensitivity in Soft-Tissue Sarcoma by Suppressing BCL-xL Expression.

Soft-tissue sarcoma (STS) is a heterogeneous group of rare tumors originating predominantly from the embryonic mesoderm. Despite the development of combined modalities including radiotherapy, STSs are often refractory to antitumor modalities, and novel strategies that improve the prognosis of STS patients are needed. We previously demonstrated the therapeutic potential of two telomerase-specific replication-competent oncolytic adenoviruses, OBP-301 and tumor suppressor p53-armed OBP-702, in human STS cells. Here, we demonstrate in vitro and in vivo antitumor effects of OBP-702 in combination with ionizing radiation against human STS cells (HT1080, NMS-2, SYO-1). OBP-702 synergistically promoted the antitumor effect of ionizing radiation in the STS cells by suppressing the expression of B-cell lymphoma-X large (BCL-xL) and enhancing ionizing radiation-induced apoptosis. The in vivo experiments demonstrated that this combination therapy significantly suppressed STS tumors' growth. Our results suggest that OBP-702 is a promising antitumor reagent for promoting the radiosensitivity of STS tumors.

Risk of HBV reactivation in HCC patients undergoing combination therapy of PD-1 inhibitors and angiogenesis inhibitors in the antiviral era

BackgroundAlthough routine antiviral therapy has been implemented in HCC patients, the risk of HBV reactivation (HBVr) remains with the use of programmed cell death-1(PD-1) blockade‐based combination immunotherapy and the relevant risk factors are also unclear. Therefore, we aimed to identify the incidence and risk factors of HBVr in HCC patients undergoing combination therapy of PD-1 inhibitors and angiogenesis inhibitors and concurrent first-line antivirals.MethodsWe included a total of 218 HBV-related HCC patients with first-line antivirals who received PD-1 inhibitors alone or together with angiogenesis inhibitors. According to the anti-tumor therapy modalities, patients were divided into PD-1 inhibitors monotherapy group (anti-PD-1 group) and combination therapy group (anti-PD-1 plus angiogenesis inhibitors group). The primary study endpoint was the incidence of HBVr.ResultsHBVr occurred in 16 (7.3%) of the 218 patients, 2 cases were found in the anti-PD-1 group and the remaining 14 cases were in the combination group. The Cox proportional hazard model identified 2 independent risk factors for HBVr: combination therapy (hazard ratio [HR], 4.608, 95%CI 1.010–21.016, P = 0.048) and hepatitis B e antigen (HBeAg) positive (HR, 3.695, 95%CI 1.246–10.957, P = 0.018). Based on the above results, we developed a simple risk-scoring system and found that the high-risk group (score = 2) developed HBVr more frequently than the low-risk group (score = 0) (Odds ratio [OR], 17.000, 95%CI 1.946–148.526, P = 0.01). The area under the ROC curve (AUC-ROC) was 7.06 (95%CI 0.581–0.831, P = 0.006).ConclusionHBeAg-positive patients receiving combination therapy have a 17-fold higher risk of HBVr than HBeAg-negative patients with PD-1 inhibitors monotherapy.

Abstract 2374: Bispecific and trispecific GPRC5D antibodies with potent cell-killing activity against multiple myeloma

Abstract Background: GPRC5D is a G protein-coupled receptor that is expressed on multiple myeloma cells but absent from most healthy tissues except for hair follicles. It is an attractive target for anti-tumor modalities including T cell-engaging bispecific antibodies. Recent clinical data demonstrate that combination therapy of T cell-engaging molecules individually targeting GPRC5D and BCMA on tumor cells offers unprecedented therapeutic benefits in relapsed/refractory multiple myeloma patients. A trispecific antibody that simultaneously engages these molecules while activating T cells is expected to provide comparable and additional benefits as a monotherapy. Methods: Multipass membrane proteins are valuable therapeutic targets but are largely inaccessible for antibody discovery due to structural complexity and high conservation. We developed an antibody discovery platform (MPS) tailored for membrane proteins that utilizes advanced immunization techniques including DNA, mRNA, and Lipoparticles (virus-like particles) and evolutionarily divergent host species. From the parental antibodies isolated, we engineered panels of GPRC5DxCD3 bispecific and GPRC5DxBCMAxCD3 trispecific antibodies using multiple formats and CD3 arms that encompass different geometries and binding stoichiometries. GPRC5D multispecific antibodies were tested for target specificity using Integral Molecular’s Membrane Proteome Array (MPA), and epitope residues were defined using alanine scanning mutagenesis. Both bispecific and trispecific antibodies were tested for in vitro potency and cytokine release using MM.1R and RPMI 8226 multiple myeloma cell lines. In vivo efficacy for a subset of antibodies was determined using xenograft models in PBMC-engrafted mice. Results: We immunized chickens with GPRC5D to obtain high-titer immune responses and configured resulting antibodies as T cell-engaging bispecific molecules with a CD3 binding arm. A subset of GPRC5DxCD3 bispecific antibodies displayed potent T cell-mediated cytotoxicity with picomolar potency on multiple myeloma cell lines. Antibody specificity profiling showed high specificity binding, and epitope mapping helped to explain why only some GPRC5D antibodies were active as multispecific antibodies. Bispecific molecules were then re-engineered as trispecific antibodies with additional binding to BCMA, a well-validated multiple myeloma target. GPRC5DxBCMAxCD3 trispecifics showed improved cytotoxic activity against multiple myeloma cells. Both bispecific and trispecific antibodies demonstrated a large window between cell killing and cytokine release. We will present in vivo efficacy data for a panel of bi- and trispecific molecules against MM.1R xenografts in PBMC-engrafted mice. Citation Format: Hayley Roth, Daniel Rogers, Ileine Sanchez, Breanna Tyrell, Trevor Barnes, Ami Snyder, Nenna Nowgu, Kyle Doolan, Benjamin J. Doranz, Ross Chambers, Joseph B. Rucker. Bispecific and trispecific GPRC5D antibodies with potent cell-killing activity against multiple myeloma [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 2374.

Emerging Translational Approaches for Lung Cancer Therapeutics

Selective inactivation of tumour genes and immunogenic activation of host anti-tumor surveillance as curement modalities are becoming more common in lung cancer due to genomics-driven comprehensive molecular profiling's identification of advanced chemically and immunologically noticeable vulnerabilities. However, recurrence occurs as a result of persistent disease states that are resistant to these specified therapies after an initially dramatic response. Although various mechanisms and conceptual frameworks for therapeutic resistance have been put forth, it has proven difficult to account for and forecast resistance trajectories in advance. In this review, we cover the present state of genomics-guided individualized therapy against both oncogenic changes and post-therapy resistance pathways in both NSCLC and SCLC. We include the most prominent oncogenic changes found in NSCLC. We list a variety of innovative and developing precision medicine strategies for SCLC, which is currently very resistant to specified therapy. In addition, we review a number of immunotherapy strategies that are being studied clinically for lung cancer, either individually or in combination. This review not only analyses tumour heterogeneity as a source of residual disease and emphasizes common molecular themes underlying many classes of resistance but also highlights potential solutions to these obstacles. We underline how a thorough understanding of these themes helps forecast and enhance therapeutic approaches, including poly-therapy approaches, to prevent tumour growth in the first place.

Current progress in the regulation of endogenous molecules for enhanced chemodynamic therapy.

Chemodynamic therapy (CDT) is a potential cancer treatment strategy, which relies on Fenton chemistry to transform hydrogen peroxide (H2O2) into highly cytotoxic reactive oxygen species (ROS) for tumor growth suppression. Although overproduced H2O2 in cancerous tissues makes CDT a feasible and specific tumor therapeutic modality, the treatment outcomes of traditional chemodynamic agents still fall short of expectations. Reprogramming cellular metabolism is one of the hallmarks of tumors, which not only supports unrestricted proliferative demands in cancer cells, but also mediates the resistance of tumor cells against many antitumor modalities. Recent discoveries have revealed that various cellular metabolites including H2O2, iron, lactate, glutathione, and lipids have distinct effects on CDT efficiency. In this perspective, we intend to provide a comprehensive summary of how different endogenous molecules impact Fenton chemistry for a deep understanding of mechanisms underlying endogenous regulation-enhanced CDT. Moreover, we point out the current challenges and offer our outlook on the future research directions in this field. We anticipate that exploring CDT through manipulating metabolism will yield significant advancements in tumor treatment.

Treatment evaluation by volumetric segmentation in pediatric optic pathway glioma: evaluation of the effect of bevacizumab on intra-tumor components.

Progressive pediatric optic pathway gliomas (OPGs) are treated by diverse systemic antitumor modalities. Refined insights on the course of intra-tumoral components are limited. We performed an exploratory study on the longitudinal volumetric course of different (intra-)tumor components by manual segmentation of MRI at the start and after 3, 6 and 12months of bevacizumab (BVZ) treatment. Thirty-one patients were treated with BVZ (median 12months, range: 2-39months). During treatment the total tumor volume decreased with median 19.9% (range: -62.3 to + 29.7%; n = 30) within the first 3 months, decreased 19.0% (range: -68.8 to + 96.1%; n = 28) between start and 6 months and 27.2% (range: -73.4 to + 36.0%; n = 21) between start and 12months. Intra-tumoral cysts were present in 12 OPGs, all showed a decrease of volume during treatment. The relative contrast enhanced volume of NF1 associated OPG (n = 11) showed an significant reduction compared to OPG with a KIAA1549-BRAF fusion (p < 0.01). Three OPGs progressed during treatment, but were not preceded by an increase of relative contrast enhancement. Treatment with BVZ of progressive pediatric OPGs leads to a decrease of both total tumor volume and cystic volume for the majority of OPGs with emphasis on the first three months. NF1 and KIAA1549-BRAF fusion related OPGs showed a different (early) treatment effect regarding the tumor enhancing component on MRI, which did not correlate with tumor volume changes. Future research is necessary to further evaluate these findings and its relevance to clinical outcome parameters.

Hollow Cavity CaO2 @Polydopamine Nanocomposites for pH-Responsive Ca2+ -Enhanced Efficient Mild Hyperthermia in the NIR-II Region.

Second near-infrared (NIR-II) mild photothermal therapy with higher tissue penetration depth and less damage to healthy tissues is emerging as an attractive antitumor modality, but its therapeutic efficiency is dramatically suppressed by the resistance of heat shock proteins (HSPs). As a widely explored photothermal agent, the application of polydopamine (PDA) in the NIR-II region is hampered by low photothermal conversion efficiency (PCE). Herein, its PCE in the NIR-II region is improved by developing novel hollow cavity CaO2 @PDA nanocomposites through chelation-induced diffusion of inner core Ca2+ to the shell PDA to facilitate multiple reflections of laser in the cavity. Upon pH-responsive degradation of CaO2 , its structure is transformed into a stacked "nano-mesh" with excellent light absorption and an enlarged effective irradiation area. Overloading of Ca2+ ions not only induces downregulation of HSPs but also enhances interference of light on membrane potential, which further aggravate mitochondrial dysfunction and reduce the thermotolerance of tumor cells, promoting efficient mild hyperthermia of PDA in the NIR-II region.

Metal organic frameworks based intelligent nanoadjuvants for boosting tumor immunotherapy through enhanced ICD and lactic acid regulation

Immunotherapy has been considered as a potent antitumor modality while its efficacy is frequently hampered by the insufficient immunogenicity and immunosuppressive tumor microenvironment (TME) with overexpressed lactic acid (LA). Herein, an intelligent nanoadjuvant MOF@LOD-siRNA@Lips-FA (MLSLF) was developed by loading lactate oxidase (LOD) and monocarboxylate transporter 4 (MCT4)-inhibiting siRNA into Fe-MOF and further coated with folate-liposomes (FA-lip) for enhanced immunotherapy. The MLSLF could response to endogenous glutathione (GSH), resulting in the depletion of GSH and production of •OH for effective chemodynamic therapy (CDT). On one hand, the death of cancer cells would arouse immunogenic cell death (ICD) to release antigens and enhance tumor immunogenicity. Meanwhile, LOD could decompose intracellular LA to H2O2, which could further promote ICD and boost anti-tumor immune response. On the other hand, the siRNA could restrict the expression of MCT4, which would hinder the intracellular LA efflux. The combination of LOD and siRNA could reduce the LA content in TME and relieve immunosuppression TME. The in vivo results revealed that MLSLF could effectually suppress the growth of primary and distant tumors in tumor-bearing mice. Therefore, the smart MLSLF could be a synergistic anti-tumor strategy for efficient immunotherapy through the combination of enhanced ICD and LA metabolism blockade.

Clinicopathological features of a rare cancer: Intrahepatic lymphoepithelioma-like cholangiocarcinoma with Epstein-Barr virus infection

PurposeEpstein-Barr virus-related lymphoepithelioma-like cholangiocarcinoma (EBV-LELCC), a subtype of intrahepatic cholangiocarcinoma (IHCC), is an extremely rare cancer. To date, only few cases have been reported. Therefore, more studies are needed to provide new insights into its clinicopathological characteristics and treatment. MethodsWe retrospectively collected data from 16 EBV-LELCC patients admitted to our hospital between January 2013 and February 2022. We summarized their clinical characteristics and analyzed the genomic features of 5 patients by whole-exon sequencing. In addition, the Kaplan-Meier method was used to assess the prognostic differences between EBV-LELCC and EBV-negative IHCC. ResultsA total of 16 EBV-LELCC patients aged between 35 and 70 were included in this study and were characterized by female predominance. Eight genetic mutations including KMT2C, ARID1B, BAZ1A, NPM1, POLE, PER3, TOPBP1, USP1 were identified from 5 patients. There were 11 stage I, 2 stage III and 3 stage IV patients in this study. The overall survival of stage I and stage III EBV-LELCC patients after radical surgery was significantly better than that of EBV-negative IHCC patients with matched stage (p = 0.0119). Notably, a stage IV patient treated with a variety of antitumor modalities including surgery, interventional therapy, radiotherapy, chemotherapy, targeted therapy and immunotherapy achieved long-term survival of more than seven years. ConclusionAltogether, EBV-LELCC presents a more favorable prognosis than IHCC. This study suggests that patients with early EBV-LELCC have a good prognosis after radical surgery, and even patients with advanced EBV-LELCC are expected to have a longer survival under appropriate and timely treatment. For such a rare cancer with unique clinicopathological features and molecular patterns, more research is needed to facilitate its diagnosis and treatment.

Ce6 nanoassemblies: Molecular mechanism and strategies for combinational anticancer therapy

AbstractNowadays, cancer has become the leading cause of death worldwide, driving the need for effective therapeutics to improve patient prognosis. Photodynamic therapy (PDT) has been widely applied as an antitumor modality, owing to its minimal invasiveness, localized tumor damage, and high safety profile. However, its efficacy is limited by poor stability of photosensitizers, inadequate tumor accumulation, and a complex tumor microenvironment. To overcome these challenges, extensive endeavors have been made to explore the co‐assembly of the widely used photosensitizer chlorin e6 (Ce6) with various functional small molecules to enhance pharmacodynamic activity. This review provides a comprehensive overview of current studies on Ce6‐based nanoparticles for effective PDT and precise delivery of functional molecules. The self‐assembly mechanism will be discussed in detail, with a focus on potential strategies for combinational therapy with PDT.

Injectable microenvironment-responsive hydrogels with redox-activatable supramolecular prodrugs mediate ferroptosis-immunotherapy for postoperative tumor treatment.

Immunotherapy is an emerging antitumor modality with high specificity and persistence, but its application for resected tumor treatment is impeded by the low availability of tumor-specific antigens and strong immunosuppression in the wound margin. Here a nanoengineered hydrogel is developed for eliciting robust cooperative ferroptosis-immunotherapeutic effect on resected tumors. Specifically, β-cyclodextrin (β-CD) is first grafted onto oxidized sodium alginate (OSA) through Schiff base ligation, which could trap cRGD-modified redox-responsive Withaferin prodrugs (WA-cRGD) to obtain the hydrogel building blocks (Gel@WA-cRGD). Under Ca2+-mediated crosslinking, Gel@WA-cRGD rapidly forms physiologically stable hydrogels, of which the porous network is used to deliver programmed cell death ligand 1 antibodies (aPD-L1). After injection into the post-surgical wound cavity, the β-CD-entrapped WA-cRGD is detached by the local acidity and specifically internalized by residual tumor cells to trigger ferroptosis, thus releasing abundant damage-associated molecular patterns (DAMPs) and tumor-derived antigens for activating the antigen-presenting cell-mediated cross-presentation and downstream cytotoxic T cell (CTL)-mediated antitumor responses. Furthermore, aPD-L1 could block PD-1/PD-L1 interaction and enhance the effector function of CTLs to overcome tumor cell-mediated immunosuppression. This cooperative hydrogel-based antitumor strategy for ferroptosis-immunotherapy may serve as a generally-applicable approach for postoperative tumor management. STATEMENT OF SIGNIFICANCE: To overcome the immunosuppressive microenvironment in resected solid tumors for enhanced patient survival, here we report a nanoengineered hydrogel incorporated supramolecular redox-activatable Withaferin prodrug and PD-L1 antibody, which could elicit robust cooperative ferroptosis-immunotherapeutic effect against residual tumor cells in the surgical bed to prevent tumor relapse, thus offering a generally-applicable approach for postoperative tumor management.

The interaction of chlorin photosensitizers for photodynamic therapy with blood transport proteins

Photodynamic therapy (PDT) is still considered to be a promising antitumor modality whose potential is not fully disclosed. This unique combination of visible light, molecular oxygen and a light-sensitive molecule, i.e. a photosensitizer (PS), leads to a generation of reactive oxygen species inducing direct tumor cell death, damage to tumor microvasculature and induction of a local inflammatory reaction. A much higher selectivity of tumor targeting during PDT can be achieved by the PS binding to appropriate delivery vehicles with pronounced affinity to tumor tissue. Blood lipoproteins are often considered to be such agents enhancing PS tumor accumulation. Here, we focus on the interaction between a series of charged chlorin PSs synthesized on a chlorophyll a platform with blood transport proteins. The ability of PS molecules to form molecular complexes with potential carries – biocompatible polymers or surfactants is also considered and briefly discussed. Our experimental studies do indicate that a charge sign, number and relative position of charged groups in the macrocyclic molecule strongly influence the PS-protein interaction. The monocationic chlorin PS with a pronounced hydrophobic surface is found to be delivered by lipoproteins, while trianionic chlorin e6 is preferentially associated with serum albumin. The addition of biocompatible micellar or polymeric carriers widely used to improve biocompatibility of many drugs remains an elution profile almost unchanged despite strong PS-carrier binding. It is important that both di- and tricationic chlorins are not associated with any transport proteins and the mechanism of their accumulation in tumors must be different from other PSs. Taking into account that lipoproteins are highly important carriers for PS molecules in antitumor photodynamic therapy, we can make an important conclusion that chlorins bearing one cationic group at a certain position of a macrocycle are more efficient photosensitizing agents compared to anionic or polycationic macrocycles.

Nanoengineered sonosensitive platelets for synergistically augmented sonodynamic tumor therapy by glutamine deprivation and cascading thrombosis.

Ultrasound (US)-activated sonodynamic therapy (SDT) stands for a distinct antitumor modality because of its attractive characteristics including intriguing noninvasiveness, desirable safety, and high tissue penetration depth, which, unfortunately, suffers from compromised therapeutic efficacy due to cancer cell-inherent adaptive mechanisms, such as glutathione (GSH) neutralization response to reactive oxygen species (ROS), and glutamine addictive properties of tumors. In this work, we developed a biological sonosensitive platelet (PLT) pharmacytes for favoring US/GSH-responsive combinational therapeutic of glutamine deprivation and augmented SDT. The amino acid transporter SLC6A14 blockade agent α-methyl-DL-tryptophan (α-MT)-loaded and MnO2-coated porphyrinic metal-organic framework (MOF) nanoparticles were encapsulated in the PLTs through the physical adsorption of electrostatic attraction and the intrinsic endocytosis of PLTs. When the sonosensitive PLT pharmacytes reached tumor sites through their natural tendencies to TME, US stimulated the PLTs-loaded porphyrinic MOF to generate ROS, resulting in morphological changes of the PLTs and the release of nanoparticles. Subsequently, intracellular high concentration of GSH and extracellular spatio-temporal controlled US irradiation programmatically triggered the release of α-MT, which enabled the synergistically amplified SDT by inducing amino acid starvation, inhibiting mTOR, and mediating ferroptosis. In addition, US stimulation achieved the targeted activation of PLTs at tumor vascular site, which evolved from circulating PLTs to dendritic PLTs, effectively blocking the blood supply of tumors through thrombus formation, and revealing the encouraging potential to facilitate tumor therapeutics.

Bioengineered nanogenerator with sustainable reactive oxygen species storm for self-reinforcing sono-chemodynamic oncotherapy

Oxidative stress-based antitumor modalities derived from reactive oxygen species (ROS) storms have attracted increasing attention. Nevertheless, low delivery efficiency, poor selectivity, hypoxia and overexpressed glutathione (GSH) have severely restricted the sustainable generation of the ROS storm in tumor cells. Herein, we design a bioengineered nanogenerator by coordination-driven co-assembly of sonosensitizer indocyanine green (ICG), Fenton-like agent copper ion (CuⅡ) and mitochondrial respiratory inhibitor metformin (MET), which is then camouflaged by a cancer cytomembrane to induce a sustainable intracellular ROS storm for on-demand self-reinforcing sono-chemodynamic oncotherapy. Such a nanogenerator with a core–shell structure, suitable diameter and outstanding stability can efficiently accumulate in tumor regions and then internalize into tumor cells through the camouflaging and homologous targeting strategy of the cancer cytomembrane. The nanogenerator shows an exceptional instability under the triple stimulations of acidic lysosomes, overexpressed GSH and ultrasound (US) radiation, thereby resulting in the rapid disassembly and burst drug release. Interestingly, the released MET significantly enhances the sonodynamic therapy (SDT) efficacy of the released ICG by inhibiting mitochondrial respiration and meanwhile the released CuⅡ obviously reduces ROS elimination by downregulating overexpressed GSH for self-amplifying and self-protecting the intracellular ROS storm. Moreover, such a nanogenerator almost completely achieves the tumor ablation in vivo in a single therapy cycle. Taken together, our bioengineered nanogenerator with a sustainable ROS storm can provide a promising strategy for ROS storm-based oncotherapy.

Abstract 6305: Bispecific claudin 18.2 and GPRC5D antibodies with potent cell-killing activity for cancer therapeutics

Abstract Tumor-associated antigens with little or no expression in healthy tissue are attractive targets for anti-tumor modalities including T cell-engaging bispecific antibodies. CLDN18.2 is a transmembrane adhesion protein undetectable in most healthy adult tissues but highly expressed in gastric, pancreatic, esophageal, and lung cancers. GPRC5D is a G protein-coupled receptor that is absent from most healthy tissues except for hair follicles but expressed on multiple myeloma cells. Both CLND18.2 and GPRC5D are novel oncology targets with no clinically approved therapies against them. Using our discovery platform tailored to complex transmembrane proteins, we have developed bispecific antibodies with the ability to potently kill CLDN18.2-positive or GPRC5D-positive cells. Multipass membrane proteins are valuable therapeutic targets in oncology and other disease areas but are largely inaccessible as antibody targets due to their poor expression, membrane-dependent structure, small extracellular regions, and poor immunogenicity due to sequence conservation. We developed an antibody discovery platform (MPS) that specifically addresses each of these challenges. This platform utilizes advanced immunization techniques including DNA, mRNA, and Lipoparticles (virus-like particles). It also employs chickens as an evolutionarily divergent host species for robust immune responses against conserved targets. Antibodies raised in chickens are directly humanized prior to isolation, reducing the need for downstream engineering. In two separate campaigns, we immunized chickens with CLDN18.2 and GPRC5D and obtained high-titer immune responses. We were successful in isolating high-affinity antibodies specific to each of these targets. Our CLDN18.2 antibodies showed specificity for their target and did not bind the closely related splice variant CLND18.1, which is highly expressed in healthy lung tissue. Additionally, the antibodies showed no binding to ~6,000 other proteins that were screened using a Membrane Proteome Array. A subset of antibodies from both discovery programs were configured as bispecific molecules using a CD3-targeting arm to bring tumor cells into close proximity with cytotoxic T cells that mediate cell killing. The panels of molecules encompassed multiple bispecific formats bearing different stoichiometries, geometries, and sizes to enable identification of lead molecules with favorable activities and safety profiles. Both CLDN18.2 and GPRC5D bispecifics showed potent T cell-mediated cytotoxicity with picomolar potency. They also showed good cytokine release and developability profiles. GPRC5D x CD3 and CLDN18.2 x CD3 bispecifics hold promise as potent and safe therapeutics for different cancer types. Citation Format: Ileine Sanchez, Hayley Roth, Brad Screnci, David Tucker, Nick Molino, Trevor Barnes, Paige Murphy, Kyle Guldner, Tim Phillips, Kristen Shema, Thomas Charpentier, Alyssa Cunningham, Janae Latta, Breanna Tyrell, Meghan Pitts, Carmen Navia, Charles Azuelos, Anna Lobley, Jawhara Karam, Valerie Fiers, Daniela Reyes, Kate Slovik, Alison Snyder, Marianne Assogba, Kai-Ti Chang, Riley Payne, Kyle Doolan, Ross Chambers, Joe Rucker, Benjamin Doranz. Bispecific claudin 18.2 and GPRC5D antibodies with potent cell-killing activity for cancer therapeutics. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6305.

CRISPR/Cas9 and Chlorophyll Coordination Micelles for Cancer Treatment by Genome Editing and Photodynamic Therapy.

CRISPR/Cas9-based gene therapy and photodynamic therapy both show promise for cancer treatment but still have their drawbacks limited by tumor microenvironment and long treatment duration. Herein, CRISPR/Cas9 genome editing and photodynamic strategy for a synergistic anti-tumor therapeutic modality is merged. Chlorophyll (Chl) extracted from natural green vegetables is encapsulated in Pluronic F127 (F127) micelles and Histidine-tagged Cas9 can be effectively chelated onto micelles via metal coordination by simple incubation, affording Cas9-Chl@F127 micelles. Mg2+ acts as an enzyme cofactor to correlatively enhance Cas9 gene-editing activity. Upon laser irradiation, Chl as an effective photosensitizer generates reactive oxygen species (ROS) to kill tumor cells. Meanwhile, CRISPR/Cas9, mediated by dual deliberately designed gRNAsof APE1 and NRF2, can reprogram the tumor microenvironment by increasing the intracellular oxygen accumulation and impairing the oxidative defense system of tumor cells. Cas9-Chl@F127 micelles can responsively release Cas9 in the presence of abundant ATP or low pH in tumor cells. In a murine tumor model, Cas9-Chl@F127 complexed with dual gRNAs including APE1 and NRF2 significantly inhibits the tumor growth. Taken together, Cas9-Chl@F127 micelles, representing the first Chl-based green biomaterial for the delivery of Cas9, show great promise for the synergistic anti-tumor treatment by PDT and gene editing.

TME-triggered copper-coordinated engineered programmable nanogenerators for on-demand cascade-amplifying oxidative stress.

Although oxidative stress-based antitumor modality derived from reactive oxygen species (ROS) storm has attracted considerable attention in copper-based nanomaterials, its efficiency is still weakened by the insufficient hydrogen peroxide (H2O2) and overexpressed glutathione (GSH) in a tumor microenvironment (TME). In view of this, we designed an engineered programmable spike-like nanogenerator via the coordination-driven co-assembly of Evans Blue (EB), copper ions (CuII), and 5-hydroxy-p-naphthoquinone (HND). For programmable nanogenerators, the introduction of EB as a stabilizer-like component can not only adjust its morphology but also achieve its visual tracking. Interestingly, such programmable nanogenerators can be efficiently enriched in tumor regions and then internalized into tumor cells due to ECH with spike-like morphology. Notably, once the nanogenerator is disintegrated and burst to release the drug upon acidic lysosome and endogenous GSH triggering, the released HND can not only efficiently amplify endogenous H2O2 by intracellular oxidoreductases but also down-regulate the peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (Pin 1) activity. In addition, the released CuII ions can efficiently catalyze the degradation of the endogenous H2O2 to amplify hydroxyl radicals (˙OH) and down-regulate the overexpressed GSH to reduce ˙OH elimination for on-demand cascade-amplifying oxidative stress. Importantly, such programmable nanogenerators show an excellent antitumor effect via down-regulating the Pin 1 activity and cascade-amplifying oxidative stress. In this study, we propose a spatiotemporally programmable cascade nanogenerator for oxidative stress-based antitumor therapy.