Analysis of Ethylene Signal Regulating Sucrose Metabolism in Strawberry Fruits Based on RNA-seq.

Ethylene is a key hormone that regulates the maturation and quality formation of horticultural crops, but its effects on non-respiratory climacteric fruits such as strawberries are not yet clear. In this study, strawberry fruits were treated with exogenous ethephon (ETH) and 1-methylcyclopropene (1-MCP). It was found that ETH treatment increased the soluble solids and anthocyanin content of the fruits, reduced hardness, and decreased organic acid content, while 1-MCP treatment inhibited these processes. Transcriptome analysis revealed that differentially expressed genes (DEGs) were enriched in the starch-sucrose metabolism pathway. qRT-PCR results further showed significant changes in the expression levels of sucrose metabolism genes, confirming the influence of ethylene signals on soluble sugar accumulation during strawberry fruit development. This study elucidates the quality changes and molecular mechanisms of ethylene signal in the development of strawberry fruits, providing some key targets and theoretical support for guiding strawberry cultivation and variety improvement.

Nanodrug-bacteria conjugates-mediated oncogenic collagen depletion enhances immune checkpoint blockade therapy against pancreatic cancer.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) cancer cells specifically produce abnormal oncogenic collagen to bind with integrin α3ß1 receptor and activate the downstream focal adhesion kinase (FAK), protein kinase B (AKT), and mitogen-activated protein kinase (MAPK) signaling pathway. Collectively, this promotes immunosuppression and tumor proliferation and restricts the response rate of clinical cancer immunotherapies. METHODS: Here, by leveraging the hypoxia tropism and excellent motility of the probiotic Escherichia coli strain Nissle 1917 (ECN), we developed nanodrug-bacteria conjugates to penetrate the extracellular matrix (ECM) and shuttle the surface-conjugated protein cages composed of collagenases and anti-programmed death-ligand 1 (PD-L1) antibodies to PDAC tumor parenchyma. FINDINGS: We found the oncogenic collagen expression in human pancreatic cancer patients and demonstrated its interaction with integrin α3ß1. We proved that reactive oxygen species (ROS) in the microenvironment of PDAC triggered collagenase release to degrade oncogenic collagen and block integrin α3ß1-FAK signaling pathway, thus overcoming the immunosuppression and synergizing with anti-PD-L1 immunotherapy. CONCLUSIONS: Collectively, our study highlights the significance of oncogenic collagen in PDAC immunotherapy, and consequently, we developed a therapeutic strategy that can deplete oncogenic collagen to synergize with immune checkpoint blockade for enhanced PDAC treatment efficacy. FUNDING: This work was supported by the University of Wisconsin Carbone Cancer Center Research Collaborative and Pancreas Cancer Research Task Force, UWCCC Transdisciplinary Cancer Immunology-Immunotherapy Pilot Project, and the start-up package from the University of Wisconsin-Madison (to Q.H.).

Estrogen receptor-negative/progesterone receptor-positive breast cancer has distinct characteristics and pathologic complete response rate after neoadjuvant chemotherapy.

PURPOSE: The status of hormone receptors (HR) is an independent factor affecting survival and chemotherapy sensitivity in breast cancer (BC) patients, with estrogen receptor (ER) and progesterone receptor (PR) having the most significant effects. The ER-/PR + phenotype has been controversial in BC, and experts will face many challenges in determining treatment strategies. Herein, we systematically analyzed the clinicopathological characteristics of the ER-/PR + phenotype in BC patients and the response to chemotherapy. PATIENTS AND METHODS: We included two cohorts. The first cohort counted the relationship between clinicopathologic data and survival outcomes for 72,666 female patients in the Surveillance, Epidemiology, and End Results (SEER) database. The second cohort analyzed the relationship between clinicopathological data and pathologic complete response (pCR) rate in 879 patients at the Harbin Medical University Cancer Hospital. The classification data were compared by the chi-square test and Fister's exact test of the Logistic regression model, and predictor variables with P < 0.05 in the univariate analysis were included in the multivariate regression analysis. The Kaplan-Meier method evaluated breast cancer-specific survival (BCSS) and overall survival (OS) to investigate the relationship between different HR typing and survival and pCR. RESULTS: In the two cohorts, 704 (0.9%) and 11 (1.3%) patients had the ER-/PR + phenotype, respectively. The clinicopathologic features of patients with the ER-/PR + phenotype are more similar to those of the ER-/PR- phenotype. The ER-/PR + phenotype is more common in younger and premenopausal women, and most ER-/PR + phenotypes exhibit higher histological grades. Survival analysis showed that there were significant differences in OS and BCSS among patients with different HR states (P < 0.001). The survival results of patients with the ER + /PR + phenotype were the best. The prognosis of the ER-/PR + phenotype was similar to that of the ER-/PR- phenotype. On the other hand, we found that HR status was also an independent predictor of post-NAC pCR rate in BC patients. The ER + /PR- and ER-/PR- phenotypes were more sensitive to chemotherapy than the ER + /PR + phenotypes. CONCLUSION: HR status is the main factor affecting BC's survival outcome and pCR rate. Patients with the ER-/PR + phenotype possess more aggressive biological factors and can benefit significantly from chemotherapy. We need to pay more attention to this group and achieve individualized treatment, which will help us treat BC better and provide new targets and blueprints for our clinical treatment.

Redox modulation with a perfluorocarbon nanoparticle to reverse Treg-mediated immunosuppression and enhance anti-tumor immunity.

Tumor hypoxia and high glutathione (GSH) expression promote regulatory T cell (Treg) infiltration and maintain its immunosuppressive function, which significantly reduces the response rate of cancer immunotherapy. Here, we developed an immunomodulatory nano-formulation (FEM@PFC) to reverse Treg-mediated immunosuppression by redox regulation in the tumor microenvironment (TME). Oxygen carried in perfluorocarbon (PFC) was delivered to the TME, thus relieving the hypoxic condition and inhibiting Treg infiltration. More importantly, GSH depletion by the prodrug efficiently restricted the Foxp3 expression and immunosuppressive function of Tregs, thus breaking the shackles of tumor immunosuppression. Additionally, the supplement of oxygen cooperated with the consumption of GSH to enhance the irradiation-induced immunogenic cell death and subsequent dendritic cell (DC) maturation, thereby efficiently promoting the activation of effector T cells and restricting the immunosuppression of Tregs. Collectively, the FEM@PFC nano-formulation reverses Treg-mediated immunosuppression and regulates the redox balance in the TME to boost anti-tumor immunity and prolong the survival of tumor-bearing mice, which provides a new immunoregulatory strategy from the perspective of redox modulation.

Small-Molecule Ferritin Degrader as a Pyroptosis Inducer.

Exploring the response of malignant cells to intracellular metabolic stress is critical for understanding pathologic processes and developing anticancer therapies. Herein, we developed ferritin-targeting proteolysis targeting chimeras (PROTACs) to establish the iron excess stress inside cancer cells and investigated subsequent cellular behaviors. We conjugated oleic acid that binds to the ferritin dimer to the ligand of von Hippel-Lindau (VHL) E3 ligase through an alkyl linker. The screened chimera, DeFer-2, degraded ferritin and then rapidly elevated the free iron content, thereby initiating the caspase 3-GSDME-mediated pyroptosis in cancer cells rather than typical ferroptosis that is always associated with iron ion overload. According to its structural and physicochemical characteristics, DeFer-2 was loaded into a tailored albumin-based nano-formulation, which substantially inhibited tumor growth and prolonged the survival time of mice bearing B16F10 subcutaneous tumors with negligible adverse effects. This study developed a ferritin-targeting PROTAC for iron overload stress, revealed iron metabolic dysregulation-mediated pyroptosis, and provided a PROTAC-based pyroptosis inducer for anticancer treatment.

Active recruitment of anti-PD-1-conjugated platelets through tumor-selective thrombosis for enhanced anticancer immunotherapy.

Immune checkpoint inhibitors (ICIs) can reinvigorate T cells to eradicate tumor cells, showing great potential in combating various types of tumors. We propose a delivery strategy to enhance tumor-selective ICI accumulation, which leverages the responsiveness of platelets and platelet-derivatives to coagulation cascade signals. A fused protein tTF-RGD targets tumor angiogenic blood vessel endothelial cells and initiates the coagulation locoregionally at the tumor site, forming a "cellular hive" to recruit anti-PD-1 antibody (aPD-1)-conjugated platelets to the tumor site and subsequently activating platelets to release aPD-1 antibody to reactivate T cells for improved immunotherapy. Moreover, on a patient-derived xenograft breast cancer model, the platelet membrane-coated nanoparticles can also respond to the coagulation signals initiated by tTF-RGD, thus enhancing the accumulation and antitumor efficacy of the loaded chemotherapeutics. Our study illustrates a versatile platform technology to enhance the local accumulation of ICIs and chemodrugs by taking advantage of the responsiveness of platelets and platelet derivatives to thrombosis.

Chemically engineering cells for precision medicine.

Cell-based therapy holds great potential to address unmet medical needs and revolutionize the healthcare industry, as demonstrated by several therapeutics such as CAR-T cell therapy and stem cell transplantation that have achieved great success clinically. Nevertheless, natural cells are often restricted by their unsatisfactory in vivo trafficking and lack of therapeutic payloads. Chemical engineering offers a cost-effective, easy-to-implement engineering tool that allows for strengthening the inherent favorable features of cells and confers them new functionalities. Moreover, in accordance with the trend of precision medicine, leveraging chemical engineering tools to tailor cells to accommodate patients individual needs has become important for the development of cell-based treatment modalities. This review presents a comprehensive summary of the currently available chemically engineered tools, introduces their application in advanced diagnosis and precision therapy, and discusses the current challenges and future opportunities.

Enhancing Gasdermin-induced tumor pyroptosis through preventing ESCRT-dependent cell membrane repair augments antitumor immune response.

Pore-forming Gasdermin protein-induced pyroptosis in tumor cells promotes anti-tumor immune response through the release of pro-inflammatory cytokines and immunogenic substances after cell rupture. However, endosomal sorting complexes required for transport (ESCRT) III-mediated cell membrane repair significantly diminishes the tumor cell pyroptosis by repairing and subsequently removing gasdermin pores. Here, we show that blocking calcium influx-triggered ESCRT III-dependent membrane repair through a biodegradable nanoparticle-mediated sustained release of calcium chelator (EI-NP) strongly enhances the intracellularly delivered GSDMD-induced tumor pyroptosis via a bacteria-based delivery system (VNP-GD). An injectable hydrogel and a lyophilized hydrogel-based cell patch are developed for peritumoral administration for treating primary and metastatic tumors, and implantation for treating inoperable tumors respectively. The hydrogels, functioning as the local therapeutic reservoirs, can sustainedly release VNP-GD to effectively trigger tumor pyroptosis and EI-NP to prevent the ESCRT III-induced plasma membrane repair to boost the pyroptosis effects, working synergistically to augment the anti-tumor immune response.

Harnessing DNA for immunotherapy: Cancer, infectious diseases, and beyond.

Despite the rapid development of immunotherapy, low response rates, poor therapeutic outcomes and severe side effects still limit their implementation, making the augmentation of immunotherapy an important goal for current research. DNA, which has principally been recognized for its functions of encoding genetic information, has recently attracted research interest due to its emerging role in immune modulation. Inspired by the intrinsic DNA-sensing signaling that triggers the host defense in response to foreign DNA, DNA or nucleic acid-based immune stimulators have been used in the prevention and treatment of various diseases. Besides that, DNA vaccines allow the synthesis of target proteins in host cells, subsequently inducing recognition of these antigens to provoke immune responses. On this basis, researchers have designed numerous vehicles for DNA and nucleic acid delivery to regulate immune systems. Additionally, DNA nanostructures have also been implemented as vaccine delivery systems to elicit strong immune responses against pathogens and diseased cells. This review will introduce the mechanism of harnessing DNA-mediated immunity for the prevention and treatment of diseases, summarize recent progress, and envisage their future applications and challenges.

Site-Specific Polyplex on CCR7 Down-Regulation and T Cell Elevation for Lymphatic Metastasis Blocking on Breast Cancer.

Tumor metastasis contributes to high cancer mortality. Tumor cells in lymph nodes (LNs) are difficult to eliminate but underlie uncontrollable systemic metastasis. The CC chemokine receptor 7 (CCR7) is overexpressed in tumor cells and interacts with CC chemokine ligand 21 (CCL21) secreted from LNs, potentiating their lymphatic migration. Here, a site-specific polyplex is developed to block the CCR7-CCL21 signal and kill tumor cells toward LNs, greatly limiting their lymphatic infiltration. A CCR7-targeting small interfering RNA (siCCR7) is condensed by mPEG-poly-(lysine) with chlorin e6 (Ce6) modification (PPLC) to form PPLC/siCCR7. The knockdown of CCR7 by siCCR7 in tumor cells significantly reduced their response on CCL21 and LN tropism. Additionally, photodynamic therapy-mediated immune activation precisely targets and kills tumor cells released from the primary foci before they reaches the LNs, reducing the number of tumor cells entering the LNs. Consequently, the PPLC/siCCR7 polyplexes inhibited up to 92% of lung metastasis in 4T1 tumor bearing mice and reduced tumor cell migration to LNs by up to 80%. This site-specific strategy optimized anti-metastasis efficacy and promotes the clinical translational development of anti-metastatic therapy.

Depletion of tumor associated macrophages enhances local and systemic platelet-mediated anti-PD-1 delivery for post-surgery tumor recurrence treatment.

Immunosuppressive cells residing in the tumor microenvironment, especially tumor associated macrophages (TAMs), hinder the infiltration and activation of T cells, limiting the anti-cancer outcomes of immune checkpoint blockade. Here, we report a biocompatible alginate-based hydrogel loaded with Pexidartinib (PLX)-encapsulated nanoparticles that gradually release PLX at the tumor site to block colony-stimulating factor 1 receptors (CSF1R) for depleting TAMs. The controlled TAM depletion creates a favorable milieu for facilitating local and systemic delivery of anti-programmed cell death protein 1 (aPD-1) antibody-conjugated platelets to inhibit post-surgery tumor recurrence. The tumor immunosuppressive microenvironment is also reprogrammed by TAM elimination, further promoting the infiltration of T cells into tumor tissues. Moreover, the inflammatory environment after surgery could trigger the activation of platelets to facilitate the release of aPD-1 accompanied with platelet-derived microparticles binding to PD-1 receptors for re-activating T cells. All these results collectively indicate that the immunotherapeutic efficacy against tumor recurrence of both local and systemic administration of aPD-1 antibody-conjugated platelets could be strengthened by local depletion of TAMs through the hydrogel reservoir.

Chemically and Biologically Engineered Bacteria-Based Delivery Systems for Emerging Diagnosis and Advanced Therapy.

Bacteria are one of the main groups of organisms, which dynamically and closely participate in human health and disease development. With the integration of chemical biotechnology, bacteria have been utilized as an emerging delivery system for various biomedical applications. Given the unique features of bacteria such as their intrinsic biocompatibility and motility, bacteria-based delivery systems have drawn wide interest in the diagnosis and treatment of various diseases, including cancer, infectious diseases, kidney failure, and hyperammonemia. Notably, at the interface of chemical biotechnology and bacteria, many research opportunities have been initiated, opening a promising frontier in biomedical application. Herein, the current synergy of chemical biotechnology and bacteria, the design principles for bacteria-based delivery systems, the microbial modulation, and the clinical translation are reviewed, with a special focus on the emerging advances in diagnosis and therapy.

In situ self-assembled peptide nanofibers for cancer theranostics.

Self-assembled nanofibers hold tremendous promise for cancer theranostics owing to their in situ assembly, spatiotemporal responsiveness, and diverse bioactivity. Herein, this review summarizes the recent advances of self-assembled peptide nanofibers and their applications in biological systems, focusing on the dynamic process of capturing cancer cells from the outside-in. (1) In situ self-assembly in response to pathological or physiological changes. (2) Diverse functions at different locations of tumors, such as forming thrombus in tumor vasculature, constructing a barrier on the cancer cell membrane, and disrupting the cancer organelles. Of note, with the assembly/aggregation induced residence (AIR) effect, the nanofibers could form a drug depot in situ for sustained release of chemotherapeutic drugs to increase their local concentration and prolong the residence time. Finally, perspectives toward future directions and challenges are presented to further understand and expand this exciting field.

Fluorine assembly nanocluster breaks the shackles of immunosuppression to turn the cold tumor hot.

Clinical investigations have shown that a nonimmunogenic "cold" tumor is usually accompanied by few immunopositive cells and more immunosuppressive cells in the tumor microenvironment (TME), which is still the bottleneck of immune activation. Here, a fluorine assembly nanocluster was explored to break the shackles of immunosuppression, reawaken the immune system, and turn the cold tumor "hot." Once under laser irradiation, FS@PMPt produces sufficient reactive oxygen species (ROS) to fracture the ROS-sensitive linker, thus releasing the cisplatin conjugated PMPt to penetrate into the tumors and kill the regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs). Meanwhile, ROS will induce potent immunogenic cell death (ICD) and further promote the accumulation of dendritic cells (DCs) and T cells, therefore not only increasing the infiltration of immunopositive cells from the outside but also reducing the immunosuppressive cells from the inside to break through the bottleneck of immune activation. The FS@PMPt nanocluster regulates the immune process in TME from negative to positive, from shallow to deep, to turn the cold tumor into a hot tumor and provoke a robust antitumor immune response.

Targeting pulmonary tumor microenvironment with CXCR4-inhibiting nanocomplex to enhance anti-PD-L1 immunotherapy.

Anti-programmed cell death 1 ligand 1 (PD-L1) therapy is extraordinarily effective in select patients with cancer. However, insufficient lymphocytic infiltration, weak T cell-induced inflammation, and immunosuppressive cell accumulation in the tumor microenvironment (TME) may greatly diminish the efficacy. Here, we report development of the FX@HP nanocomplex composed of fluorinated polymerized CXCR4 antagonism (FX) and paclitaxel-loaded human serum albumin (HP) for pulmonary delivery of anti-PD-L1 small interfering RNA (siPD-L1) to treat orthotopic lung tumors. FX@HP induced T cell infiltration, increased expression of calreticulin on tumor cells, and reduced the myeloid-derived suppressor cells/regulatory T cells in the TME, thereby acting synergistically with siPD-L1 for effective immunotherapy. Our work suggests that the CXCR4-inhibiting nanocomplex decreases tumor fibrosis, facilitates T cell infiltration and relieves immunosuppression to modulate the immune process to improve the objective response rate of anti-PD-L1 immunotherapy.

Treatment of acute lung injury and early- and late-stage pulmonary fibrosis with combination emulsion siRNA polyplexes.

Acute lung injury (ALI) and idiopathic pulmonary fibrosis (IPF) are severe lung diseases causing irreversible lung damage and premature death. Both diseases share multiple pathological features, including overexpression of C-X-C chemokine receptor type 4 (CXCR4) and upregulation of plasminogen activator inhibitor-1 (PAI-1). The goal of the present study was to evaluate therapeutic potential of pulmonary treatment with combined inhibition of CXCR4 and PAI-1 in ALI and various disease stages of IPF. We report preparation of perfluorocarbon emulsion polyplexes containing a fluorinated polymeric CXCR4 antagonist (F-PAMD) as an siRNA carrier suitable for pulmonary delivery. In vitro testing of the emulsion polyplexes in primary lung fibroblasts from IPF mice showed high cellular uptake and promising antifibrotic effect as indicated by the decreased expression of α smooth muscle actin, when compared with conventional siRNA polyplexes. Biodistribution analysis in mice with IPF showed prolonged lung retention and widespread lung distribution following intratracheal administration of the formulations. The emulsion polyplexes showed promising therapeutic efficacy in ALI and in early fibrinogenic stage of IPF. Increased survival was observed in the model of late-stage IPF. The use of perfluorocarbon emulsion polyplexes to achieve combined CXCR4 antagonism and PAI-1 inhibition is a promising strategy for treatment of ALI and IPF.

Increased Survival by Pulmonary Treatment of Established Lung Metastases with Dual STAT3/CXCR4 Inhibition by siRNA Nanoemulsions.

Lung metastasis is a common and deadly occurrence in many types of solid tumors. Chemokine receptor CXCR4 and transcription factor signal transducer and activator of transcription 3 (STAT3) are among potential therapeutic targets in lung metastatic cancer. Both CXCR4 and STAT3 play important roles in the proliferation, angiogenesis, and metastasis of cancer cells. Here, we report on the development of a pulmonary delivery (p.d.) system based on perfluorocarbon (PFC) nanoemulsions for combined delivery of a partially fluorinated polymeric CXCR4 antagonist (FM) and anti-STAT3 small interfering RNA (siRNA). We have prepared FM-stabilized PFC (FM@PFC) as a delivery system of therapeutic siRNA adsorbed on the surface of the emulsion. These FM@PFC/siRNA nanoemulsions inhibited both CXCR4 and STAT3, as demonstrated by effective anti-invasive ability in vitro and related antimetastatic activity in vivo. The combined nanoemulsions provided a comprehensive anticancer effect in the model of established lung metastasis of breast carcinoma, which was dependent on induction of cancer cell apoptosis, anti-angiogenic effect, anti-invasive activity, and overcoming of the immunosuppressive tumor microenvironment. Direct comparison with intravenous (i.v.) injection showed superior activity of pulmonary administration as indicated by significantly increased animal survival. Overall, this work established the suitability of the PFC nanoemulsions for p.d. of combination anticancer treatments and as a promising method to treat lung metastasis.

Reversibly Stabilized Polycation Nanoparticles for Combination Treatment of Early- and Late-Stage Metastatic Breast Cancer.

Metastatic breast cancer is a major cause of cancer-related female mortality worldwide. The signal transducer and activator of transcription 3 (STAT3) and the chemokine receptor CXCR4 are involved in the metastatic spread of breast cancer. The goal of this study was to develop nanomedicine treatment based on combined inhibition of STAT3 and CXCR4. We synthesized a library of CXCR4-inhibiting polymers with a combination of beneficial features that included PEGylation, fluorination, and bioreducibility to achieve systemic delivery of siRNA to silence STAT3 expression in the tumors. An in vivo structure-activity relationship study in an experimental lung metastasis model revealed superior antimetastatic activity of bioreducible fluorinated polyplexes when compared with nonreducible controls despite similar CXCR4 antagonism and the ability to inhibit in vitro cancer cell invasion. When compared with nonreducible and nonfluorinated polyplexes, improved siRNA delivery was observed with the bioreducible fluorinated polyplexes. The improvement was ascribed to a combination of enhanced physical stability, decreased serum destabilization, and improved intracellular trafficking. Pharmacokinetic analysis showed that fluorination decreased the rate of renal clearance of the polyplexes and contributed to enhanced accumulation in the tumors. Therapeutic efficacy of the polyplexes with STAT3 siRNA was assessed in early stage breast cancer and late-stage metastatic breast cancer with primary tumor resection. Strong inhibition of the primary tumor growth and pronounced antimetastatic effects were observed in both models of metastatic breast cancer. Mechanistic studies revealed multifaceted mechanism of action of the combined STAT3 and CXCR4 inhibition by the developed polyplexes relying both on local and systemic effects.

Engineering a high-affinity humanized anti-CD24 antibody to target hepatocellular carcinoma by a novel CDR grafting design.

Cluster of differentiation 24 (CD24) is a specific surface marker involved in the tumorigenesis and progression of hepatocellular carcinoma (HCC). However, all reported anti-CD24 antibodies are murine ones with inevitable immunogenicity. To address this, a method using both molecular structure and docking-based complementarity determining region (CDR) grafting was employed for humanization. After xenogeneic CDR grafting into a human antibody, three types of canonical residues (in the VL/VH interface core, in the loop foundation, and interaction with loop residues) that support loop conformation and residues involved in the antigen-binding interface were back-mutated. Four engineered antibodies were produced, among which hG7-BM3 has virtually identical 3-D structure and affinity parameters with the parental chimeric antibody cG7. In vitro, hG7-BM3 demonstrated superior immunogenicity and serum stability to cG7. Antibody-dependent cellular cytotoxicity (ADCC), tumor cell internalization and in vivo targeting assays indicate that hG7-BM3 has the potential for development as an antibody-drug conjugate (ADC). We therefore generated the hG7-BM3-VcMMAE conjugate, which was shown to induce tumor cell apoptosis and effectively suppress nude mice bearing HCC xenografts. In conclusion, our study provides new inspiration for antibody humanization and an ADC candidate for laboratory study and clinical applications.

Anti-VEGFR2-interferon-α2 regulates the tumor microenvironment and exhibits potent antitumor efficacy against colorectal cancer.

Interferon-α (IFNα) has multiple antitumor effects including direct antitumor toxicity and the ability to potently stimulate both innate and adaptive immunity. However, its clinical applications in the treatment of malignancies have been limited because of short half-life and serious adverse reactions when attempting to deliver therapeutically effective doses. To address these issues, we fused IFNα2a to the anti-vascular endothelial growth factor and receptor 2 (VEGFR2) antibody JZA00 with the goal of targeting it to the tumor microenvironment where it can stimulate the antitumor immune response. The fusion protein, JZA01, is effective against colorectal cancer by inhibiting angiogenesis, exhibiting direct cytotoxicity, and activating the antitumor immune response. Although JZA01 exhibited reduced IFNα2 activity in vitro compared with native IFNα2, VEGFR2 targeting permitted efficient antiproliferative, proapoptotic, antiangiogenesis, and immune-stimulating effects against the colorectal tumors HCT-116 and SW620. JZA01 showed in vivo efficacy in NOD-SCID mice-bearing established HCT-116 tumors. In conclusion, this study describes an antitumor immunotherapy that is highly promising for the treatment of colorectal cancer.