The introduction of checkpoint immunotherapeutic agents in the last decade has revolutionized cancer treatment. Although anti-PD-1, anti-PD-L1 and anti-CTLA4 are promising therapies, many patients fail to respond or relapse due to drug resistance potentially due to redundancy of immune checkpoints. One of the ways to improve the efficacy of this cancer treatment is to target two or even three immune checkpoints. To date, the benefit of combined anti-VISTA/anti-PD-L1 therapy has been confirmed, but no one has investigated the efficacy of blocking these negative immune checkpoints with a bispecific anti-VISTA/anti-PD-L1 antibody.
In this study, the bispecific antibodies (bsAbs) were produced in three formats: symmetric (IgG-HC-scFv), asymmetric (Fab-scFv-Fc(KIH)) and 2 x scFv. The binding and blocking properties of these bispecific antibodies (bsAbs) and their efficacy compared to monotherapy and combination therapy were then determined using endometrial (RL95-2), pancreatic (PANC-1) and breast (BT-20) cancer cell lines.
The bsAbs generated in this study showed weaker binding properties to PD-1 and VISTA in ELISA (EC50) than the parent antibodies (atezolizumab and onvatilimab). Blockade of VISTA/VSIG-3 binding was also weaker with bsAbs compared to onvatilimab, but the ability to block the PD-1/PD-L1 pathway was slightly better than with atezolizumab. The Fc-based bsAbs showed statistically significant higher levels of lysis of endometrial, breast and pancreatic cancer cells. The symmetric bsAbs (IgG-HC-scFv) showed the most promising therapeutic potential. Higher levels of cancer cell lysis were associated with higher levels of pro-inflammatory cytokines. Both the asymmetric and symmetric bsAbs resulted in higher secretion levels of IFN-γ, TNFα and Granzyme B than anti-VISTA, anti-PD-L1 monotherapy and anti-VISTA/anti-PD-L1 combination therapy.
The high level of tumor cell lysis and increased expression of pro-inflammatory cytokines induced by the Fc-based bsAbs suggest a novel approach for the treatment of pancreatic, endometrial and breast cancer.
Copyright © 2025 Bielski, Barczyński, Mikitiuk, Myrcha, Rykała, Boon, Gąsior, Hec-Gałązka, Holak and Sitar.

The adoptive transfer of T cells expressing chimeric antigen receptors (CARs) is effective in B cell malignancies. However, the persistence of cancer cells with low levels or complete absence of the target antigen, thereby evading detection by CAR T cells, leads to relapse. These evasion mechanisms highlight the need for receptors with enhanced sensitivity and multispecificity. We introduce a synthetic chimeric T cell receptor (ChTCR) that confers superior antigen sensitivity compared with CARS and previous hybrid TCR designs and is readily adapted for bispecific targeting. ChTCRs replicate the structure of natural TCRs, form classical immune synapses and demonstrate TCR-like signaling. T cells expressing bispecific ChTCRs (Bi-ChTCRs) are more effective than bispecific CAR T cells in eradicating tumors with heterogeneous antigen expression in vivo in female mice. The Bi-ChTCR architecture is resilient and can be designed to target pairs of B cell and multiple myeloma antigens. These findings provide a widely applicable strategy to combat tumor heterogeneity and prevent relapse.
© 2025. The Author(s).

BRAF-mutated colorectal cancer correlates with poor prognosis and limited response to standard treatments. Combining immune checkpoint inhibitors with BRAF/MEK inhibitors shows promise against BRAF-mutant melanoma in both preclinical and clinical trials. Therefore, we hypothesized that the treatment would be effective against BRAF-mutant colorectal cancer. In this study, we assessed the efficacy of combining immune checkpoint inhibitors with BRAF and/or MEK inhibitors in BRAF-mutant colorectal cancers. We treated BRAF V600E colorectal cancer cells HT-29 and SNU-1235 with encorafenib (BRAF inhibitor) and binimetinib (MEK inhibitor) and assessed the degrees of MAPK inhibition, JAK/STAT inhibition, cell viability, apoptosis, and the expression of antigen presenting machinery. We also inoculated HT-29 cells into mice and treated them with an immune checkpoint inhibitor (durvalumab), encorafenib, and binimetinib for 4 weeks. We found that treatment with BRAF inhibitor, MEK inhibitor, or their combination led to significant tumor growth reduction, along with the MAPK and JAK/STAT pathway inhibition, antigen presenting machinery induction, and cytotoxic T cell activation. Our study demonstrates the potential effectiveness of combining immune checkpoint inhibitors with BRAF or MEK inhibitors for BRAF-mutated colorectal cancers.
© 2025. The Author(s).

The aim of this study was to investigate the regenerative effect of lyophilized dental follicle mesenchymal stem cells (DF-MSCs) combined with rat platelet-rich fibrin (PRF) on geriatric skin wounds. Human DF-MSCs which were isolated from the wisdom teeth of healthy donors and PRF were mixed and incubated in a 37 °C incubator for 1-2 h containing 1 million cells in 150 mg PRF. The mixture was suspended in a freeze-drying solution and then lyophilized. Wounds were created on the back skin of Wistar albino rats using a 6 mm punch. Lyophilized DF-MSCs, PRF, or PRF + DF-MSCs were applied to the wounds of rats. On the 15th day, the wound area was histopathologically evaluated in rats. Blood samples from rats were analyzed for total antioxidant status (TAOS), and inflammatory cytokine levels using ELISA. In both young and geriatric rats treated with lyophilized PRF + DF-MSCs, wound area began to significantly decrease from the 10th day compared to the untreated group (p < 0.05). Histopathological examination revealed that in the lyophilized PRF + DF-MSCs treated groups, epithelial integrity and scarless healing significantly increased compared to the untreated groups (p < 0.05). There were no significant differences in TAOS, total oxidant status (TOS), tumor necrosis factor (TNF), interleukin-6 (IL6), and hydroxyproline levels in serum samples from young rats on the 15th day. In geriatric rats, hydroxyproline (HYPS) levels were increased in the DF-MSC and PRF + DF-MSC groups (p < 0.01), TNF was significantly elevated in PRF geriatric group and IL6 was increased in the PRF group compared to the control group (p = 0.01). Lyophilized PRF + DF-MSCs, which is a shelf-stable and ready-to-use product, hold promise, especially for traumatic wounds in geriatric individuals with longer healing times.
© 2025. The Author(s).

Pseudomonas aeruginosa is the predominant pathogen causing chronic infection in the airway of patients with bronchiectasis (BE), a chronic respiratory disease with high prevalence worldwide. Environmental factors are vital for bacterial successful colonization. Here, with sputa and bronchoalveolar lavage fluids, we determined that the concentration of airway antimicrobial peptide LL-37 and lactate was elevated in BE patients, especially in those infected with P. aeruginosa. The in vitro antibacterial assay revealed the bactericidal activity of LL-37 against the clinical P. aeruginosa isolates, which were dampened in the acidic condition. P. aeruginosa production of outer membrane vesicles (OMVs) enhanced in the lactate-adjusted acidic condition. Transcriptomic analysis suggested that OMVs induce the hyperproduction of the chemical compound 2-heptyl-4-quinolone (HHQ) in the bacterial population, which was verified by high-performance liquid chromatography. The positively charged HHQ interfered with the binding of LL-37 to bacterial cell membrane, potentiating the P. aeruginosa resistance to LL-37. To our knowledge, this is a new resistance mechanism of P. aeruginosa against antimicrobial peptides and may provide theoretical support for the development of new antibacterial therapies.
© 2025 The Author(s). MedComm published by Sichuan International Medical Exchange & Promotion Association (SCIMEA) and John Wiley & Sons Australia, Ltd.