Breast cancer stem cells (BCSCs) resist conventional treatments and cause tumor recurrence. Almost 25% of breast cancers overexpress human epidermal growth factor receptor-2 (HER2). Here we developed a novel multi-targeted nanosystem to specifically eradicate HER2⁺BCSCs. Plasmids containing CXCR1 promoter, PE38 toxin, and 5′UTR of the basic fibroblast growth factor-2 (bFGF 5'UTR) were constructed. Polyamidoamine (PAMAM) dendrimers functionalized with anti-HER2 VHHs were used for plasmid delivery. Stem cell proportion of MDA-MB-231, MDA-MB-231/HER2⁺and MCF-10A were evaluated by mammosphere formation assay. Hanging drop technique was used to produce spheroids. The uptake, gene expression, and killing efficacy of the multi-targeted nanosystem were evaluated in both monolayer and spheroid culture. MDA-MB-231/HER2⁺had higher ability to form mammosphere compared to MCF-10A. Our multi-targeted nanosystem efficiently inhibited the mammosphere formation of MDA-MB-231 and MDA-MB-231/HER2⁺cells, while it was unable to prevent the mammosphere formation of MCF-10A. In the hanging drop culture, MDA-MB-231/HER⁺generated compact well-rounded spheroids, while MCF-10A failed to form compact cellular masses. The multi-targeted nanosystem showed much better uptake, higher PE38 expression, and subsequent cell death in MDA-MB-231/HER2⁺compared to MCF-10A. However, the efficacy of our targeted toxin gene therapy was lower in MDA-MB-231/HER2⁺spheroids compared with that in the monolayer culture. the combination of the cell surface, transcriptional, and translational targeting increased the stringency of the treatment.

Background: Breast cancer stem cells (BCSCs) are cells with a higher ability to metastasis and resistance to conventional treatments. They have a phenotype of (CD44high/CD24low) and the unlimited ability for proliferation. Development of strategies to target the BCSC population may lead to the establishment of more effective cancer therapies. Pseudomonas exotoxin A (PE) is a potent cytotoxic protein. CXCR1 promoter provides BCSC and HER2 specificity on transcription level. 5′UTR of the basic fibroblast growth factor-2 (bFGF 5ʹUTR) provides tumor specificity on translation level. Here, we utilized a mutant form of PE encoding DNA “PE38”, CXCR1 promoter and bFGF 5ʹUTR to target BCSCs. Methods: The stemness of SK-BR-3, MDA-MB-231 and MCF10A cell lines were evaluated based on the expression of the CD44high/CD24low stem cell signature and the ability to form mammospheres. Then, the cell lines were transfected with constructs encoding luciferase/PE38 under the control of the CMV/CXCR1 promoter with or without bFGF 5′UTR. Luciferase protein expression was evaluated using dual-luciferase reporter assay. PE38 transcript expression was measured by real-time PCR, and the cytotoxic effect of PE38 protein expression was determined by MTT assay. Results: The percentage of CD44high/CD24low population did not correlate to mammosphere forming efficiency (MFE). Given that the percentage of CD44 high/CD24 low is not a conclusive BCSC profile, we based our work on the mammosphere assay. However, in comparison with MCF10A, the two tumorigenic cell lines had higher MFE, probably due to their higher BCSC content. Reporter assay and real-time PCR results demonstrated that CXCR1 promoter combined with bFGF 5ʹUTR increased BCSC-specific gene expression. Meanwhile, tightly regulated expression of PE38 using these two gene regulatory elements resulted in high levels of cell death in the two tumorigenic cell lines while having little toxicity toward normal MCF10A. Conclusion: Our data show that PE38, CXCR1 promoter and bFGF 5ʹUTR in combination can be considered as a promising tool for killer gene therapy of breast cancer.

Background /Objectives: The SARS-CoV-2’s high mutations and replication rates contribute to its high infectivity and resistance to current vaccinations and treatments. The primary cause of resistance to most current treatments aligns within the coding regions for the spike S protein of SARS-CoV-2 that has mutated. As a potential novel immunotherapy, we generated a novel fusion protein composed of a soluble ACE2 (sACE2) linked to llama-derived anti-CD16 that targets different variants of spike proteins and enhances natural killer cells to target infected cells. Methods: Here, we generated a novel sACE2-AntiCD16VHH fusion protein using a Gly4Ser linker, synthesized and cloned into the pLVX-EF1alpha-IRES-Puro vector, and further expressed in ExpiCHO-S cells and purified using Ni⁺NTA chromatography. Results: The fusion protein significantly blocked SARS-CoV-2 alpha, beta, delta, gamma, and omicron S-proteins binding and activating angiotensin-converting enzyme receptor-2 (ACE2) on ACE2-expressing RAW-Blue macrophage cells and the secretion of several key inflammatory cytokines, G-CSF, MIP-1A, and MCP-1, implicated in the cytokine release storm (CRS). The sACE2-Anti-CD16VHH fusion protein also bridged NK cells to ACE2-expressing human lung carcinoma A549 cells and significantly activated NK-dependent cytotoxicity. Conclusions: The findings show that a VHH directed against CD16 could be an excellent candidate to be linked to soluble ACE2 to generate a bi-specific molecule (sACE2-AntiCD16VHH) suitable for bridging effector cells and infected target cells to inhibit SARS-CoV-2 variant spike proteins binding to the ACE2 receptor in the RAW-Blue cell line and pro-inflammatory cytokines and to activate natural killer cell cytotoxicity.

Background: Previous studies have shown the altered levels of long-chain polyunsaturated fatty acids (LCPUFAs) in pathological hypoxic conditions. Elevated soluble fms-like tyrosine kinase-1 (sFlt-1) expression in hypoxia plays an important role in the pathogenesis of placental as preeclampsia. Objectives: The eicosapentaenoic acid (EPA; 20:5, n-3) as LCPUFAs (omega-3) might attenuate sFlt-1 and hypoxia-inducible factor-1α (HIF-1α) expressions and secretions. Methods: JEG-3 cells were incubated with dimethyloxalylglycine (DMOG) and EPA. The SFlt-1 gene expression was determined using a real-time polymerase chain reaction. The protein secretion of sFlt-1 and HIF-1α were analyzed using Western blot. Results: The incubation of JEG-3 cells with DMOG significantly elevated messenger ribonucleic acid (mRNA) expression and protein secretion of sFlt-1 (P < 0.05); nevertheless, EPA decreased mRNA expression and protein secretion of sFlt-1 (P < 0.05). Moreover, EPA inhibited the effect of DMOG on sFlt-1 (P = 0.0361) gene expression and protein secretion and HIF-1α (P = 0.0241) protein secretion. Conclusions: The sFlt-1 expression decreased by n-3 fatty acids in trophoblast tumor cell line under induced hypoxia conditions. It seems that changes in sFlt-1 expression are mediated by the transcription factor HIF-1α.

Immune-mediated diseases remain a major global health challenge, while current immunotherapies are often limited by systemic toxicity, poor tissue selectivity, and insufficient control over immune responses. This review examines recent advances in targeted immune modulation through smart biomaterials, programmable biopharmaceuticals, and hybrid biomaterial–biologic systems. A structured literature review was conducted using peer-reviewed studies published between 2022 and 2026 from major biomedical databases. The reviewed evidence indicates that smart biomaterials can respond to disease-associated microenvironmental signals, such as pH changes, hypoxia, oxidative stress, and inflammatory enzymes, enabling localized and context-dependent therapeutic activity. Programmable biopharmaceuticals further enhance precision by directing immune-cell behavior and molecular signaling through engineered biological mechanisms. In addition, hybrid biomaterial–biologic platforms combine environmental responsiveness with the potency of biologic therapeutics, improving immune targeting and reducing unintended systemic effects. Collectively, these technologies support a transition from conventional systemic immunomodulation toward adaptive and site-specific immune control. Despite significant progress, important challenges remain, including biological complexity, manufacturing scalability, long-term safety assessment, and regulatory translation. Nevertheless, the convergence of biomaterials science, synthetic biology, and immunoengineering is creating new opportunities for precision immunotherapy. Targeted immune modulation through programmable and responsive therapeutic systems represents a promising framework for developing safer, more effective, and clinically durable next-generation immunomodulatory therapies.