Orthotopic Pancreatic Tumor Model Research Articles

Knocking down of Xkr8 enhances chemotherapy efficacy through modulating tumor immune microenvironment

Scramblase Xk-related protein 8 (Xkr8) regulates the externalization of phosphatidylserine (PS) during apoptosis and holds a pivotal role in fostering tumor immunosuppression. Targeting Xkr8 in conjunction with chemotherapy demonstrated a novel avenue for amplifying antitumor immune response and overcoming chemo-immune resistance. Here we further evaluated this strategy by using a clinically relevant orthotopic model and elucidated the mechanism through in-depth single-cell RNA sequencing (scRNA-seq). We found that Xkr8 knockdown exhibited the potential to lead to immunogenic cell death (ICD) by impeding the normal clearance of apoptotic cells. Co-delivery of Xkr8 small interference RNA (siRNA) and a prodrug conjugate of 5-fluorouracil (5-Fu) and oxoplatin (FuOXP) showed remarkable therapeutic efficacy in an orthotopic pancreatic tumor model with increased infiltration of proliferative NK cells and activated macrophages in the tumor microenvironment (TME). Single-cell trajectory analysis further unveiled that tumor infiltrating CD8+ T cells are differentiated favorably to cytotoxic over exhausted phenotype after combination treatment. Our study sheds new light on the impact of Xkr8 knockdown on TME and solidifies the rationale of combining Xkr8 knockdown with chemotherapy to treat various types of cancers.

Tumor-penetrating iRGD facilitates penetration of poly(floxuridine-ketal)-based nanomedicine for enhanced pancreatic cancer therapy

Efficient intratumoral penetration is essential for nanomedicine to eradicate pancreatic tumors. Although nanomedicine can enter the perivascular space of pancreatic tumors, their access to distal tumor cells, aloof from the vessels, remains a formidable challenge. Here, we synthesized an acid-activatable macromolecular prodrug of floxuridine (FUDR)—poly(FUDR-ketal), engineered a micellar nanomedicine of FUDR, and intravenously co-administered the nanomedicine with the tumor-penetrating peptide iRGD for enhanced treatment of pancreatic tumor. A FUDR-derived mono-isopropenyl ether was synthesized and underwent self-addition polymerization to afford the hydrophobic poly(FUDR-ketal), which was subsequently co-assembled with amphiphilic DSPE-mPEG into the micellar nanomedicine with size of 12 nm and drug content of 56.8 wt% using nanoprecipitation technique. The acetone-based ketal-linked poly(FUDR-ketal) was triggered by acid to release FUDR to inhibit cell proliferation. In an orthotopic pancreatic tumor model derived from KPC (KrasLSL-G12D/+; Trp53LSL-R172H/+; Pdx1-Cre) cells that overexpress neuropilin-1 (NRP-1) receptor, iRGD improved penetration of FUDR nanomedicine into tumor parenchyma and potentiated the therapeutic efficacy. Our nanoplatform, along with iRGD, thus appears to be promising for efficient penetration and activation of acid-responsive nanomedicines for enhanced pancreatic cancer therapy.

Inoculation of Pan02 cells produces tumor nodules in mouse pancreas: Characterization of a novel orthotopic pancreatic ductal adenocarcinoma tumor model for interventional studies.

Preclinical models of cancer are vital for assessing and predicting efficacies and toxicities of novel treatments prior to testing in human subjects. Current pancreatic tumor models exhibit variable growth rates, unpredictable tumor size after implantation in non-native tissues, or require surgical implantation. Surgical implantation in the pancreas may produce not only unpredictable tumor uptake but could also elicit additional inflammatory responses. In searching for a pancreatic carcinoma cell that can be introduced into a mouse via simple injection, we found that Pan02, a murine ductal pancreatic adenocarcinoma derived from a pancreatic lesion of a C57BL/6 mouse, inoculated peritoneally can consistently produce pancreatic tumors. This intraperitoneal, but not intravenous, introduction of Pan02 cells leads to the attachment and growth of Pan02 in the pancreas before spreading to other tissues. Time-course tissue analysis indicates that the Pan02 cells first find, infiltrate, and grow within the pancreas, producing a pancreatic tumor model. This model appears to mimic pancreatic cancer development in humans and is the first reported use of Pan02 cells to produce orthotopic pancreatic and metastatic neoplasms in a mouse model without the need for tumor implantation within matrices or survival surgeries. This orthotopic pancreatic tumor model, with consistent tumor uptake, synchronized tumor development and survival, and predictable outcomes may enable and accelerate the preclinical evaluation of treatment candidates for pancreatic cancer.

Knocking down of Xkr8 enhances chemotherapy efficacy through modulating tumor immune microenvironment

Scramblase Xkr8 regulates the externalization of phosphatidylserine (PS) during apoptosis and holds a pivotal role in fostering tumor immunosuppression. Targeting Xkr8 in conjunction with chemotherapy demonstrated a novel avenue for amplifying antitumor immune response and overcoming chemo-immune resistance. Here we further evaluated this strategy by using a clinically relevant orthotopic model and elucidated the mechanism through in-depth single-cell RNA sequencing (scRNA-seq). We found that Xkr8 knockdown exhibited the potential to lead to immunogenic cell death (ICD) by impeding the normal clearance of apoptotic cells. Co-delivery of Xkr8 small interference RNA (siRNA) and chemo prodrug FuOXP showed remarkable therapeutic efficacy in an orthotopic pancreatic tumor model with an increase of proliferative NK cells and activated macrophages infiltration in the tumor microenvironment (TME). Single-cell trajectory analysis further unveiled that tumor infiltrating CD8+ T cells are differentiated favorably to cytotoxic over exhausted phenotype after combination treatment. Our study sheds new light on the impact of Xkr8 knockdown on TME and solidifies the rationale of combining Xkr8 knockdown with chemotherapy to treat various types of cancers.

Endoscopic Hyperspectral Imaging System to Discriminate Tissue Characteristics in Tissue Phantom and Orthotopic Mouse Pancreatic Tumor Model.

In this study, we developed an endoscopic hyperspectral imaging (eHSI) system and evaluated its performance in analyzing tissues within tissue phantoms and orthotopic mouse pancreatic tumor models. Our custom-built eHSI system incorporated a liquid crystal tunable filter. To assess its tissue discrimination capabilities, we acquired images of tissue phantoms, distinguishing between fat and muscle regions. The system underwent supervised training using labeled samples, and this classification model was then applied to other tissue phantom images for evaluation. In the tissue phantom experiment, the eHSI effectively differentiated muscle from fat and background tissues. The precision scores regarding fat tissue classification were 98.3% for the support vector machine, 97.7% for the neural network, and 96.0% with a light gradient-boosting machine algorithm, respectively. Furthermore, we applied the eHSI system to identify tumors within an orthotopic mouse pancreatic tumor model. The F-score of each pancreatic tumor-bearing model reached 73.1% for the KPC tumor model and 63.1% for the Pan02 tumor models. The refined imaging conditions and optimization of the fine-tuning of classification algorithms enhance the versatility and diagnostic efficacy of eHSI in biomedical applications.

Statins abrogate gemcitabine-induced PD-L1 expression in pancreatic cancer-associated fibroblasts and cancer cells with improved therapeutic outcome.

Acombination of chemotherapy with immunotherapy has been proposed to have better clinical outcomes in Pancreatic Ductal Adenocarcinoma (PDAC). On the other hand, chemotherapeutics is known to have certain unwanted effects on the tumor microenvironment that may mask the expected beneficial effects of immunotherapy. Here, we have investigated the effect of gemcitabine (GEM), on two immune checkpoint proteins (PD-L1 and PD-L2) expression in cancer associated fibroblasts (CAFs) and pancreatic cancer cells (PCCs). Findings of in vitro studies conducted by using in-culture activated mouse pancreatic stellate cells (mPSCs) and human PDAC patients derived CAFs demonstrated that GEM significantly induces PD-L1 and PD-L2 expression in these cells. Moreover, GEM induced phosphorylation of STAT1 and production of multiple known PD-L1-inducing secretory proteins including IFN-γ in CAFs. Upregulation of PD-L1 in PSCs/CAFs upon GEM treatment caused T cell inactivation and apoptosis in vitro. Importantly, Statins suppressed GEM-induced PD-L1 expression both in CAFs and PCCs while abrogating the inactivation of T-cells caused by GEM-treated PSCs/CAFs. Finally, in an immunocompetent syngeneic orthotopic mouse pancreatic tumor model, simvastatin and GEM combination therapy significantly reduced intra-tumor PD-L1 expression and noticeably reduced the overall tumor burden and metastasis incidence. Together, the findings of this study have provided experimental evidence that illustrates potential unwanted side effects of GEM that could hamper the effectiveness of this drug as mono and/or combination therapy. At the same time the findings also suggest use of statins along with GEM will help in overcoming these shortcomings and warrant further clinical investigation.

Tumor-repopulating cell-derived microparticles elicit cascade amplification of chemotherapy-induced antitumor immunity to boost anti-PD-1 therapy

Immune checkpoint blockade (ICB) therapy, particularly antibodies targeting the programmed death receptor 1 (PD-1) and its ligand (PD-L1), has revolutionized cancer treatment. However, its efficacy as a standalone therapy remains limited. Although ICB therapy in combination with chemotherapy shows promising therapeutic responses, the challenge lies in amplifying chemotherapy-induced antitumor immunity effectively. This relies on efficient drug delivery to tumor cells and robust antigen presentation by dendritic cells (DCs). Here, we developed tumor-repopulating cell (TRC)-derived microparticles with exceptional tumor targeting to deliver doxorubicin (DOX@3D-MPs) for improve anti-PD-1 therapy. DOX@3D-MPs effectively elicit immunogenic tumor cell death to release sufficient tumor antigens. Heat shock protein 70 (HSP70) overexpressed in DOX@3D-MPs contributes to capturing tumor antigens, promoting their phagocytosis by DCs, and facilitating DCs maturation, leading to the activation of CD8+ T cells. DOX@3D-MPs significantly enhance the curative response of anti-PD-1 treatment in large subcutaneous H22 hepatoma, orthotopic 4T1 breast tumor and Panc02 pancreatic tumor models. These results demonstrate that DOX@3D-MPs hold promise as agents to improve the response rate to ICB therapy and generate long-lasting immune memory to prevent tumor relapse.

Novel GRPR-Targeting Peptide for Pancreatic Cancer Molecular Imaging in Orthotopic and Liver Metastasis Mouse Models.

Despite advancements in pancreatic cancer treatment, it remains one of the most lethal malignancies with extremely poor diagnosis and prognosis. Herein, we demonstrated the efficiency of a novel peptide GB-6 labeled with a near-infrared (NIR) fluorescent dye 3H-indolium, 2-[2-[2-[(2-carboxyethyl)thio]-3-[2-[1,3-dihydro-3,3-dimethyl-5-sulfo-1-(3-sulfopropyl)-2H-indol-2-ylidene]ethylidene]-1-cyclohexen-1-yl]ethenyl]-3,3-dimethyl-5-sulfo-1-(3-sulfopropyl)-, inner salt (MPA) and radionuclide technetium-99m (99mTc) as targeting probes using the gastrin-releasing peptide receptor (GRPR) that is overexpressed in pancreatic cancer as the target. A short linear peptide with excellent in vivo stability was identified, and its radiotracer [99mTc]Tc-HYNIC-PEG4-GB-6 and the NIR probe MPA-PEG4-GB-6 exhibited selective and specific uptake by tumors in an SW1990 pancreatic cancer xenograft mouse model. The favorable biodistribution of the tracer [99mTc]Tc-HYNIC-PEG4-GB-6 in vivo afforded tumor-specific accumulation with high tumor-to-muscle and -bone contrasts and renal body clearance at 1 h after injection. The biodistribution analysis revealed that the tumor-to-pancreas and -intestine fluorescence signal ratios were 5.2 ± 0.3 and 6.3 ± 1.5, respectively, in the SW1990 subcutaneous xenograft model. Furthermore, the high signal accumulation in the orthotopic pancreatic and liver metastasis tumor models with tumor-to-pancreas and -liver fluorescence signal ratios of 7.66 ± 0.48 and 3.94 ± 0.47, respectively, enabled clear tumor visualization for intraoperative navigation. The rapid tumor targeting, precise tumor boundary delineation, chemical versatility, and high potency of the novel GB-6 peptide established it as a high-contrast imaging probe for the clinical detection of GRPR, with compelling additional potential in molecular-targeted therapy.

Co-delivery of three synergistic chemotherapeutics in a core-shell nanoscale coordination polymer for the treatment of pancreatic cancer

The combination chemotherapy regimen FOLFIRINOX comprising folinic acid, 5-fluorouracil, irinotecan, and oxaliplatin is the first-line treatment for patients with advanced pancreatic cancer, but its use remains prohibitive for the majority of patients due to severe side effects. Here, we report a core-shell nanoscale coordination polymer (NCP) nanoparticle co-delivering a potent and synergistic combination of oxaliplatin, gemcitabine, and SN38 (OGS), for the treatment of pancreatic cancer in mouse models. OGS contains key synergistic components of FOLFIRINOX in a controllable drug ratio., It exhibited particle stability in blood circulation and enhanced deposition of the drugs in acidic tumor environments. In vitro, OGS showed superior cytotoxicity over free drug combinations and robust cytotoxic synergism among its three components. In vivo, OGS improved drug circulation, increased tumor deposition, and exhibited superior antitumor efficacy over the free drug combination in both subcutaneous and orthotopic pancreatic tumor models. OGS treatment achieved 75–91% tumor growth inhibition and prolonged mouse survival by 1.6- to 2.8-folds while minimizing systemic toxicities such as neutropenia, hepatotoxicity, and renal toxicity. This work uncovers a novel and clinically relevant nanomedicine strategy to co-deliver synergistic combination chemotherapies for difficult-to-treat cancers.

Pancreatic Cancer Presents Distinct Nanomechanical Properties During Progression.

Cancer progression is closely related to changes in the structure and mechanical properties of the tumor microenvironment (TME). In many solid tumors, including pancreatic cancer, the interplay among the different components of the TME leads to a desmoplastic reaction mainly due to collagen overproduction. Desmoplasia is responsible for the stiffening of the tumor, poses a major barrier to effective drug delivery and has been associated with poor prognosis. The understanding of the involved mechanisms in desmoplasia and the identification of nanomechanical and collagen-based properties that characterize the state of a particular tumor can lead to the development of novel diagnostic and prognostic biomarkers. In this study, in vitro experiments were conducted using two human pancreatic cell lines. Morphological and cytoskeleton characteristics, cells' stiffness and invasive properties were assessed using optical and atomic force microscopy techniques and cell spheroid invasion assay. Subsequently, the two cell lines were used to develop orthotopic pancreatic tumor models. Tissue biopsies were collected at different times of tumor growth for the study of the nanomechanical and collagen-based optical properties of the tissue using Atomic Force Microscopy (AFM) and picrosirius red polarization microscopy, respectively. The results from the in vitro experiments demonstrated that the more invasive cells are softer and present a more elongated shape with more oriented F-actin stress fibers. Furthermore, ex vivo studies of orthotopic tumor biopsies on MIAPaCa-2 and BxPC-3 murine tumor models highlighted that pancreatic cancer presents distinct nanomechanical and collagen-based optical properties relevant to cancer progression. The stiffness spectrums (in terms of Young's modulus values) showed that the higher elasticity distributions were increasing during cancer progression mainly due desmoplasia (collagen overproduction), while a lower elasticity peak was evident -due to cancer cells softening- on both tumor models. Optical microscopy studies highlighted that collagen content increases while collagen fibers tend to form align patterns. Consequently, during cancer progression nanomechanical and collagen-based optical properties alter in relation to changes in collagen content. Therefore, they have the potential to be used as novel biomarkers for assessing and monitoring tumor progression and treatment outcomes.

Intraperitoneal Delivery of Iopamidol to Assess Extracellular pH of Orthotopic Pancreatic Tumor Model by CEST-MRI.

The extracellular pH (pHe) of solid tumors is often acidic, as a consequence of the Warburg effect, and an altered metabolic state is often associated with malignancy. It has been shown that acidosis can promote tumor progression; thus, many therapeutic strategies have been adopted against tumor metabolism; one of these involves alkalinization therapies to raise tumor pH to inhibit tumor progression, improve immune surveillance, and overcome resistance to chemotherapies. Chemical exchange saturation transfer-magnetic resonance imaging (CEST-MRI) is a noninvasive technique that can measure pH in vivo using pH-sensitive contrast agents. Iopamidol, an iodinated contrast agent, clinically used for computed tomography (CT), contains amide group protons with pH-dependent exchange rates that can reveal the pHe of the tumor microenvironment. In this study, we optimized intraperitoneal (IP) delivery of iopamidol to facilitate longitudinal assessments of orthotopic pancreatic tumor pHe by CEST-MRI. Following IV-infusion and IP-bolus injections, we compared the two protocols for assessing tumor pH. Time-resolved CT imaging was used to evaluate the uptake of iopamidol in the tumor, revealing that IP-bolus delivered a high amount of contrast agent 40 min postinjection, which was similar to the amounts reached with the IV-infusion protocol. As expected, both IP and IV injection protocols produced comparable measurements of tumor pHe, showing no statistically significant difference between groups (p=0.16). In addition, we showed the ability to conduct longitudinal monitoring of tumor pHe using CEST-MRI with the IP injection protocol, revealing a statistically significant increase in tumor pHe following bicarbonate administration (p < 0.001). In conclusion, this study shows the capability to measure pHe using an IP delivery of iopamidol into orthotopic pancreatic tumors, which is important to conduct longitudinal studies.

Dynamic ginsenoside-sheltered nanocatalysts for safe ferroptosis-apoptosis combined therapy

Chemodynamic therapy (CDT)-activated apoptosis is a potential anticancer strategy. However, CDT encounters a bottleneck in clinical translation due to its serious side effects and low efficacy. Here, we first reveal that surface engineering of ginsenoside Rg3 dramatically alters the organ distribution and tumor enrichment of systematically administered nanocatalysts using the orthotopic pancreatic tumor model while avoiding toxicity and increasing efficacy in vivo to address the key and universal toxicity problems encountered in nanomedicine. Compared with nanocatalysts alone, Rg3-sheltered dynamic nanocatalysts form hydrophilic nanoclusters, prolonging their circulation lifespan in the blood, protecting the internal nanocatalysts from leakage while allowing their specific release at the tumor site. Moreover, the nanoclusters provide a drug-loading platform for Rg3 so that more Rg3 reaches the tumor site to achieve obvious synergistic effect with nanocatalysts. Rg3-sheltered dynamic nanocatalysts can simultaneously activate ferroptosis and apoptosis to significantly improve anticancer efficacy. Systematic administration of ginsenoside Rg3-sheltered nanocatalysts inhibited 86.6% of tumor growth without toxicity and prolonged the survival time of mice. This study provides a promising approach of nanomedicine with high biosafety and a new outlook for catalytic ferroptosis-apoptosis combined antitumor therapies. Statement of significanceChemodynamic therapy (CDT) has limited clinical efficacy in cancer. In this study, we developed Rg3-sheltered dynamic nanocatalysts, which could simultaneously activate ferroptosis based on CDT-activated apoptosis, and ultimately form a combined therapy of ferroptosis-apoptosis to kill tumors. Studies have shown that the nanocatalysts after Rg3 surface engineering dramatically alters the pharmacokinetics and organ distribution of the nanocatalysts after being systematically administered, resulting in avoiding the toxicity of the nanocatalysts. Nanocatalysts also act as a drug-loading platform, guiding more Rg3 into the tumor site. This study emphasizes that nanocatalysts after Rg3 surface engineering improve the safety and effectiveness of ferroptosis-apoptosis combined therapy, providing an effective idea for clinical practices.

Nanocarrier Co-formulation for Delivery of a TLR7 Agonist plus an Immunogenic Cell Death Stimulus Triggers Effective Pancreatic Cancer Chemo-immunotherapy.

Although toll-like receptor (TLR) agonists hold great promise as immune modulators for reprogramming the suppressive immune landscape in pancreatic ductal adenocarcinoma (PDAC), their use is limited by poor pharmacokinetics (PK) and off-target systemic inflammatory effects. To overcome these challenges as well as to attain drug synergy, we developed a lipid bilayer (LB)-coated mesoporous silica nanoparticle (silicasome) platform for co-delivery of the TLR7/8 agonist 3M-052 with the immunogenic chemotherapeutic agent irinotecan. This was accomplished by incorporating the C18 lipid tail of 3M-052 in the coated LB, also useful for irinotecan remote loading in the porous interior. Not only did the co-formulated carrier improve PK, but it strengthened the irinotecan-induced immunogenic cell death response by 3M-052-mediated dendritic cell activation at the tumor site as well as participating lymph nodes. The accompanying increase in CD8+ T-cell infiltration along with a reduced number of regulatory T-cells was associated with tumor shrinkage and metastasis disappearance in subcutaneous and orthotopic KRAS-mediated pancreatic carcinoma tumor models. Moreover, this therapeutic outcome was accomplished without drug or nanocarrier toxicity. All considered, dual-delivery strategies that combine chemo-immunotherapy with co-formulated TLR agonists or other lipid-soluble immune modulators predict successful intervention in heterogeneous PDAC immune landscapes.

Abstract 4231: Hypoxia reduction in tandem with anti-angiogenic therapy remodels the pancreatic tumor microenvironment

Abstract Pancreatic cancer patients remain largely unresponsive to single agent immune checkpoint blockade (ICB) therapy. This refractory state originates from PDAC’s unique immunosuppressive microenvironment that is densely populated by suppressive myeloid cells, exhibits poor vascularity, and is highly hypoxic. We previously showed that the hypoxia-activated prodrug TH-302 (Evofosfamide) reduces intratumoral hypoxia through a tissue remodeling process, initiates tumor vasculature reorganization, and sensitizes aggressive, spontaneous murine models of prostate cancer to ICB. In a clinical trial testing the combination of Evofosfamide with cytotoxic T-lymphocyte-associated protein (CTLA-4) blockade (NCT03098160) a subset of metastatic, immune checkpoint blockade refractory patients showed prolonged progression free survival. While these studies highlight hypoxia reduction as therapeutically tractable, we lack complete understanding of the contribution of the tumor vasculature to hypoxia reduction therapy, as well as the downstream consequences of hypoxia reduction on the cellular composition of the tumor microenvironment and the responsiveness to immunotherapy. We used a transplantable, orthotopic pancreatic tumor model derived from Kras+/LSL-G12D; Trp53+/LSL-R172H; Pdx1-Cre mice and a syngeneic prostate tumor model to study tumor responses to hypoxia-reduction, anti-angiogenic therapy, and combination immunotherapy. We find that Evofosfamide with anti-VEGFR-2 (DC101) significantly extends mouse survival. Combination therapy reduces intratumoral hypoxia, improves vessel integrity, and increases intratumoral DNA damage. In response to the improved metabolic microenvironment, CD8 T cells gain enhanced effector function and lose expression of exhaustion-associated features. Like other anti-angiogenic regimens, combination Evofosfamide and DC101 leads to an increased frequency of PD-L1 expressing cells within the tumor, however blockade of PD-1 failed to prolong survival. Bulk-tumor RNA sequencing and tumor infiltrating lymphocyte analysis implicates immature myeloid cells as mediators of resistance. Re-education of the infiltrating myeloid stroma with CD40 agonist could improve responses to hypoxia-reducing, anti-angiogenic therapy. Put together, these data indicate that targeted hypoxic reduction with anti-angiogenic therapy remodels the pancreatic tumor microenvironment. In this setting, CD40 agonist over PD-1 blockade provides an additive benefit in prolonging mouse survival. Citation Format: Arthur J. Liu, Michael A. Curran. Hypoxia reduction in tandem with anti-angiogenic therapy remodels the pancreatic tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4231.

Nanodrug simultaneously regulates stromal extracellular matrix and glucose metabolism for effective immunotherapy against orthotopic pancreatic cancer

The extremely dense stromal extracellular matrix (ECM) and low glucose level affect the infiltration and effector functions of antitumor immune cells, respectively, which remain two concurrent main hurdles for achieving effective immunotherapy in pancreatic tumor. It has been found that the C-X-C motif chemokine ligand 1 (CXCL1) gene negatively regulates the stromal ECM formation, while the glucose transporter 1 (GLUT1) gene mediates aberrant glucose metabolism to restrict the glucose supply to antitumor immune cells. Here, in order to restore the anti-tumor immunity for effective immunotherapy against the pancreatic tumor, a pH-sensitive and mesothelin-targeting nanodrug incorporating CXCL1 siRNA and PI3K signaling inhibitor wortmannin (WT) was prepared. Owing to its unique structural design, the nanodrug showed advantages to accumulate in and penetrate the orthotopic pancreatic tumor. Consequently, silencing the CXCL1 gene inhibited the stromal ECM formation to promote tumor infiltration of antitumor immune cells, and suppressing GLUT1 through PI3K inhibition blocked the glycolysis of cancer cells and tumor-associated macrophages (TAMs) to increase the intratumor glucose level for sustained effector functions of antitumor immune cells. In murine orthotopic pancreatic tumor model, this multifunctional nanodrug mediated strong synergistic effect of CXCL1 siRNA and WT to remodel the unfavorable tumor immunosuppressive microenvironment (TME), which dramatically boosted the therapeutic efficacy of immune checkpoint blockade in this highly malignant and immunologically “cold” tumor.

Fe2P nanorods based photothermal therapy combined with immune checkpoint inhibitors for pancreatic cancer

Abstract Treatment of pancreatic cancer is faced with great difficulties and challenges due to high lethality and metastasis. Synergism of targeted therapy and immunotherapy has been considered as ideal strategy to both eliminate primary tumors and control metastases. For the treatment of advanced pancreatic cancer, we demonstrated a local photothermal therapy (PTT) following administration of monoclonal antibody of programmed death ligand 1 (αPD-L1). Fe2P nanorods were employed as a Fenton agent and photothermal agent, which modified with DSPE-PEG2000-Mal for improved biocompatibility and Mal mediated-antigen presentation. Under a low dose laser irradiation at 980 nm, Fe2P-PEG-Mal nanorods (NRs) mediated PTT could induce immunogenic tumor cell death that can cause dendritic cells (DCs) infiltration and maturation. In a bilateral pancreatic tumor model, the local treatment of NRs-PTT on primary tumor could cause the increased infiltration of cytotoxic T lymphocytes (CTLs) and decreased residential of M2 macrophages in untreated distal tumors. Furthermore, subsequently intervened αPD-L1 could enhance cell death triggered by CTLs in distal tumors through reversing immunosuppression. An orthotopic pancreatic tumor model was used to further confirm the therapeutic outcome. Finally, the combination of NRs based PTT and αPD-L1 based immunotherapy was able to significantly eliminate orthotopic pancreatic tumors and reduce mesentery metastases. Thus, the strategy may provide a more effective treatment for pancreatic cancer.

Ultrasound‐Responsive Microfluidic Microbubbles for Combination Tumor Treatment

AbstractAnticancer treatment is a worldwide challenge, and there are constant endeavors in this field to develop intelligent drug delivery systems to improve the therapeutic effect. This paper presents novel ultrasound‐responsive delivery microbubbles (UDMs), with chemotherapeutic‐encapsulated hydrogel shells and therapeutic gas cores. These are developed using microfluidic electrospray to treat pancreatic tumors. As the hydrogel shells are composed of thermo‐sensitive poly(N‐isopropylacrylamide) (PNIPAM), they can contract under mild hyperthermia generated by the acoustic‐wave‐induced oscillation of the encapsulated gas. Thus, by triggering with ultrasound, the chemotherapeutic Gemcitabine (Gem) encapsulated in the hydrogel shells and the hydrogen sulfide (H2S) encased in the cores can be controllably released. Based on the UDMs, it is demonstrated that the multiple drug‐loaded microcarriers exhibit an excellent tumor‐killing effect through combination therapy. The Gem and H2S coloaded UDMs have good therapeutic efficacy and adaptability in an orthotopic pancreatic implantation tumor model established in mice. These results indicate that the proposed UDMs have a large potential for codelivery of multiple drugs for treating deep‐seated tumors and for realizing combination drug treatment to avoid untoward reactions.

Enriched pharmacokinetic behavior and antitumor efficacy of thymoquinone by liposomal delivery.

Background: Thymoquinone (TQ) has potential anti-inflammatory, immunomodulatory and anticancer effects but its clinical use is limited by its low solubility, poor bioavailability and rapid clearance. Aim: To enhance systemic bioavailability and tumor-specific toxicity of TQ. Materials & methods: Cationic liposomal formulation of TQ (D1T) was prepared via ethanol injection method and their physicochemical properties, anticancer effects in orthotopic xenograft pancreatic tumor model and pharmacokinetic behavior of D1T relative to TQ were evaluated. Results: D1T showed prominent inhibition of pancreatic tumor progression, significantly greater in vivo absorption, approximately 1.5-fold higher plasma concentration, higher bioavailability, reduced volume of distribution and improved clearance relative to TQ. Conclusion: Encapsulation of TQ in cationic liposomal formulation enhanced its bioavailability and anticancer efficacy against xenograft pancreatic tumor.

Ablative Radiotherapy Reprograms the Tumor Microenvironment of a Pancreatic Tumor in Favoring the Immune Checkpoint Blockade Therapy.

The low overall survival rate of patients with pancreatic cancer has driven research to seek a new therapeutic protocol. Radiotherapy (RT) is frequently an option in the neoadjuvant or palliative settings for pancreatic cancer treatment. This study explored the effect of RT protocols on the tumor microenvironment (TME) and their consequent impact on anti-programmed cell death ligand-1 (PD-L1) therapy. Using a murine orthotopic pancreatic tumor model, UN-KC-6141, RT-disturbed TME was examined by immunohistochemical staining. The results showed that ablative RT is more effective than fractionated RT at recruiting T cells. On the other hand, fractionated RT induces more myeloid-derived suppressor cell infiltration than ablative RT. The RT-disturbed TME presents a higher perfusion rate per vessel. The increase in vessel perfusion is associated with a higher amount of anti-PD-L1 antibody being delivered to the tumor. Animal survival is increased by anti-PD-L1 therapy after ablative RT, with 67% of treated animals surviving more than 30 days after tumor inoculation compared to a median survival time of 16.5 days for the control group. Splenocytes isolated from surviving animals were specifically cytotoxic for UN-KC-6141 cells. We conclude that the ablative RT-induced TME is more suited than conventional RT-induced TME to combination therapy with immune checkpoint blockade.

Metallothionein-1G suppresses pancreatic cancer cell stemness by limiting activin A secretion via NF-κB inhibition.

Resistance to chemotherapy is a long-standing problem in the management of cancer, and cancer stem cells are regarded as the main source of this resistance. This study aimed to investigate metallothionein (MT)-1G involvement in the regulation of cancer stemness and provide a strategy to overcome chemoresistance in pancreatic ductal adenocarcinoma (PDAC).Methods: MT1G was identified as a critical factor related with gemcitabine resistance in PDAC cells by mRNA microarray. Its effects on PDAC stemness were evaluated through sphere formation and tumorigenicity. LC-MS/MS analysis of conditional medium revealed that activin A, a NF-κB target, was a major protein secreted from gemcitabine resistant PDAC cells. Both loss-of-function and gain-of-function approaches were used to validate that MT1G inhibited NF-κB-activin A pathway. Orthotopic pancreatic tumor model was employed to explore the effects on gemcitabine resistance with recombinant follistatin to block activin A.Results: Downregulation of MT1G due to hypermethylation of its promoter is related with pancreatic cancer stemness. Secretome analysis revealed that activin A, a NF-κB target, was highly secreted by drug resistant cells. It promotes pancreatic cancer stemness in Smad4-dependent or independent manners. Mechanistically, MT1G negatively regulates NF-κB signaling and promotes the degradation of NF-κB p65 subunit by enhancing the expression of E3 ligase TRAF7. Blockade of activin A signaling with follistatin could overcome gemcitabine resistance.Conclusions: MT1G suppresses PDAC stemness by limiting activin A secretion via NF-κB inhibition. The blockade of the activin A signaling with follistatin may provide a promising therapeutic strategy for overcoming gemcitabine resistance in PDAC.