JUN mediates the senescence associated secretory phenotype and immune cell recruitment to prevent prostate cancer progression.

BACKGROUND: Prostate cancer develops through malignant transformation of the prostate epithelium in a stepwise, mutation-driven process. Although activator protein-1 transcription factors such as JUN have been implicated as potential oncogenic drivers, the molecular programs contributing to prostate cancer progression are not fully understood. METHODS: We analyzed JUN expression in clinical prostate cancer samples across different stages and investigated its functional role in a Pten-deficient mouse model. We performed histopathological examinations, transcriptomic analyses and explored the senescence-associated secretory phenotype in the tumor microenvironment. RESULTS: Elevated JUN levels characterized early-stage prostate cancer and predicted improved survival in human and murine samples. Immune-phenotyping of Pten-deficient prostates revealed high accumulation of tumor-infiltrating leukocytes, particularly innate immune cells, neutrophils and macrophages as well as high levels of STAT3 activation and IL-1ß production. Jun depletion in a Pten-deficient background prevented immune cell attraction which was accompanied by significant reduction of active STAT3 and IL-1ß and accelerated prostate tumor growth. Comparative transcriptome profiling of prostate epithelial cells revealed a senescence-associated gene signature, upregulation of pro-inflammatory processes involved in immune cell attraction and of chemokines such as IL-1ß, TNF-α, CCL3 and CCL8 in Pten-deficient prostates. Strikingly, JUN depletion reversed both the senescence-associated secretory phenotype and senescence-associated immune cell infiltration but had no impact on cell cycle arrest. As a result, JUN depletion in Pten-deficient prostates interfered with the senescence-associated immune clearance and accelerated tumor growth. CONCLUSIONS: Our results suggest that JUN acts as tumor-suppressor and decelerates the progression of prostate cancer by transcriptional regulation of senescence- and inflammation-associated genes. This study opens avenues for novel treatment strategies that could impede disease progression and improve patient outcomes.

Preclinical and Clinical Observations Implying Combination Therapy to Enhance the Efficacy of the Her-2/neu B-Cell Peptide-Based Vaccine HER-Vaxx and to Prevent Immune Evasion.

Her-2/neu-targeting therapy by passive application with trastuzumab is associated with acquired resistance and subsequent metastasis development, which is attributed to the upregulation of tumoral PD-L1 expression and the downregulation of Her-2/neu. We aimed to investigate this association, following active immunization with our recently constructed B-cell peptide-based Her-2/neu vaccines in both preclinical and clinical settings. Immunohistochemistry (IHC), fluorescence in situ hybridization (FISH), and combined positive score (CPS) were applied to evaluate Her-2/neu and PD-L1 expression using a murine syngeneic tumor model for Her-2/neu lung metastases and tumor biopsies from a gastric cancer patient with disease progression. A significant and concomitant reduction in Her-2/neu and the upregulation of PD-L1 expression was observed in vaccinated mice after 45 days, but not after 30 days, of metastases development. A significant increase in tumor-infiltrating B lymphocytes was observed at both time points. The downregulation of Her-2/neu and the upregulation of PD-L1 were observed in a patient's primary tumor at the disease progression time point but not prior to vaccination (Her-2/neu IHC: 3 to 0, FISH: 4.98 to 1.63; PD-L1 CPS: 0% to 5%). Our results further underline the need for combination therapy by targeting PD-L1 to prevent metastasis formation and immune evasion of Her-2/neu-positive and PD-L1-negative tumor cells.

Vaccine Based on Recombinant Fusion Protein Combining Hepatitis B Virus PreS with SARS-CoV-2 Wild-Type- and Omicron-Derived Receptor Binding Domain Strongly Induces Omicron-Neutralizing Antibodies in a Murine Model.

BACKGROUND: COVID-19, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a recurrent endemic disease affecting the whole world. Since November 2021, Omicron and its subvariants have dominated in the spread of the disease. In order to prevent severe courses of disease, vaccines are needed to boost and maintain antibody levels capable of neutralizing Omicron. Recently, we produced and characterized a SARS-CoV-2 vaccine based on a recombinant fusion protein consisting of hepatitis B virus (HBV)-derived PreS and two SARS-CoV-2 wild-type RBDs. OBJECTIVES: To develop a PreS-RBD vaccine which induces high levels of Omicron-specific neutralizing antibodies. METHODS: We designed, produced, characterized and compared strain-specific (wild-type: W-PreS-W; Omicron: O-PreS-O), bivalent (mix of W-PreS-W and O-PreS-O) and chimeric (i.e., W-PreS-O) SARS-CoV-2 protein subunit vaccines. Immunogens were characterized in vitro using protein chemical methods, mass spectrometry, and circular dichroism in combination with thermal denaturation and immunological methods. In addition, BALB/c mice were immunized with aluminum-hydroxide-adsorbed proteins and aluminum hydroxide alone (i.e., placebo) to study the specific antibody and cytokine responses, safety and Omicron neutralization. RESULTS: Defined and pure immunogens could be produced in significant quantities as secreted and folded proteins in mammalian cells. The antibodies induced after vaccination with different doses of strain-specific, bivalent and chimeric PreS-RBD fusion proteins reacted with wild-type and Omicron RBD in a dose-dependent manner and resulted in a mixed Th1/Th2 immune response. Interestingly, the RBD-specific IgG levels induced with the different vaccines were comparable, but the W-PreS-O-induced virus neutralization titers against Omicron (median VNT50: 5000) were seven- and twofold higher than the W-PreS-W- and O-PreS-O-specific ones, respectively, and they were six-fold higher than those of the bivalent vaccine. CONCLUSION: Among the tested immunogens, the chimeric PreS-RBD subunit vaccine, W-PreS-O, induced the highest neutralizing antibody titers against Omicron. Thus, W-PreS-O seems to be a highly promising COVID-19 vaccine candidate for further preclinical and clinical evaluation.

Development and In Vivo Evaluation of Small-Molecule Ligands for Positron Emission Tomography of Immune Checkpoint Modulation Targeting Programmed Cell Death 1 Ligand 1.

A substantial portion of patients do not benefit from programmed cell death protein 1/programmed cell death 1 ligand 1 (PD-1/PD-L1) checkpoint inhibition therapies, necessitating a deeper understanding of predictive biomarkers. Immunohistochemistry (IHC) has played a pivotal role in assessing PD-L1 expression, but small-molecule positron emission tomography (PET) tracers could offer a promising avenue to address IHC-associated limitations, i.e., invasiveness and PD-L1 expression heterogeneity. PET tracers would allow for improved quantification of PD-L1 through noninvasive whole-body imaging, thereby enhancing patient stratification. Here, a large series of PD-L1 targeting small molecules were synthesized, leveraging advantageous substructures to achieve exceptionally low nanomolar affinities. Compound 5c emerged as a promising candidate (IC50 = 10.2 nM) and underwent successful carbon-11 radiolabeling. However, a lack of in vivo tracer uptake in xenografts and notable accumulation in excretory organs was observed, underscoring the challenges encountered in small-molecule PD-L1 PET tracer development. The findings, including structure-activity relationships and in vivo biodistribution data, stand to illuminate the path forward for refining small-molecule PD-L1 PET tracers.