Siplizumab, a humanized anti-CD2 monoclonal antibody, has been used in conditioning regimens for hematopoietic cell transplantation and tolerance induction with combined kidney-bone marrow transplantation. Siplizumab-based tolerance induction regimens deplete T cells globally while enriching regulatory T cells (Tregs) early posttransplantation. Siplizumab inhibits allogeneic mixed-lymphocyte reactions (MLRs) in vitro. We compared the impact of siplizumab on Tregs versus other T cell subsets in HLA-mismatched allogeneic MLRs using PBMCs. Siplizumab predominantly reduced the percentage of CD4⁺ and CD8⁺ effector memory T cells, which express higher CD2 levels than naïve T cells or resting Tregs. Conversely, siplizumab enriched proliferating CD45RA⁻ FoxP3^HI cells in MLRs. FoxP3 expression was stable over time in siplizumab-containing cultures, consistent with enrichment for bona fide Tregs. Consistently, high-throughput TCRβ CDR3 sequencing of sorted unstimulated and proliferating T cells in MLRs revealed selective expansion of donor-reactive Tregs along with depletion of donor-reactive CD4⁺ effector/memory T cells in siplizumab-containing MLRs. These results indicate that siplizumab may have immunomodulatory functions that may contribute to its success in tolerance-inducing regimens. Our studies also confirm that naïve in addition to effector/memory T cells contribute to the allogeneic MLR and mandate further investigation of the impact of siplizumab on alloreactive naïve T cells.

Background/Aim: DOG1 is a calcium-activated chloride channel that has gained attention as a promising drug target due to its involvement in several processes essential for tumor development and progression. DOG1 is overexpressed in >95% of gastrointestinal stromal tumors (GIST). The aim was to determine DOG1 inhibition antitumoral effects on GIST. Materials and Methods: Human GIST (GIST-T1 and GIST882) cell lines were used to study the effect of DOG1 inhibitors on chloride currents, viability, colony formation, and cell cycle. Results: CaCC_inh-A01 decreased chloride currents. CaCC_inh-A01 and T16_inh-A01 reduced GIST cell viability and CaCC_inh-A01 affected cell cycle distribution leading to G₁ cell-cycle arrest. CaCC_inh-A01 also increased the sub-G₁ phase population, indicative of apoptosis, in GIST882. CaCC_inh-A01 strongly reduced the colony forming ability of the cells, whereas T16_inh-A01 did not. Conclusion: DOG1 inhibition has antitumoral effects in GIST cells in vitro, and could potentially serve as a target for GIST therapy.

In this real-world study, the aims were to prospectively evaluate the expression of inflammatory proteins in serum collected from head and neck cancer patients before and after treatment, and to assess whether there were differences in expression associated with treatment modalities. The mixed study cohort consisted of 180 patients with head and neck cancer. The most common tumor sites were the oropharynx (n = 81), the oral cavity (n = 53), and the larynx (n = 22). Blood tests for proteomics analysis were carried out before treatment, 7 weeks after the start of treatment, and 3 and 12 months after the termination of treatment. Sera were analyzed for 83 proteins using an immuno-oncology biomarker panel (Olink, Uppsala, Sweden). Patients were divided into four treatment groups: surgery alone (Surg group, n = 24), radiotherapy with or without surgery (RT group, n = 94), radiotherapy with concomitant cisplatin (CRT group, n = 47), and radiotherapy with concomitant targeted therapy (RT Cetux group, n = 15). For the overall cohort, the expression levels of 15 of the 83 proteins changed significantly between the pretreatment sample and the sample taken 7 weeks after the start of treatment. At 7 weeks after the start of treatment, 13 proteins showed lower expression in the CRT group compared to the RT group. The majority of the inflammatory proteins had returned to their pretreatment levels after 12 months. It was clearly demonstrated that cisplatin-based chemoradiation has immunological effects in patients with head and neck cancer. This analysis draws attention to several inflammatory proteins that are of interest for further studies.

Combined antigen-specific T cell receptor stimulation and costimulation are needed for complete T cell activation. Belatacept and Abatacept are non-depleting fusion proteins blocking CD28/B7 costimulation, while siplizumab is a depleting anti-CD2 IgG1 monoclonal antibody targeting CD2/CD58 costimulation. Herein, the effect of siplizumab combination therapy with Abatacept or Belatacept on T cell alloreactivity in mixed lymphocyte reaction (MLR) was investigated. In contrast to monotherapy, the combination of siplizumab with Belatacept or Abatacept induced near complete suppression of T cell proliferation and increased the potency of siplizumab-mediated T cell inhibition. Furthermore, dual targeting of CD2 and CD28 costimulation enhanced the selective depletion of memory T cells when compared to monotherapy. While siplizumab monotherapy leads to significant regulatory T cell (Treg) enrichment, high doses of CTLA4-Ig in the combination therapy reduced this effect. These results support the clinical evaluation of dual costimulation blockade, combining siplizumab with Abatacept or Belatacept, for the prophylaxis of organ transplant rejection and improvement of long-term outcomes following transplantation. Ongoing investigative research will elucidate if other forms of siplizumab-based dual costimulatory blockade may be able to induce similarly strong inhibition of T cell activation while still allowing for enrichment of regulatory T cells.