Personalized RNA mutanome vaccines mobilize poly-specific therapeutic immunity against cancer.

T cells directed against mutant neo-epitopes drive cancer immunity. However, spontaneous immune recognition of mutations is inefficient. We recently introduced the concept of individualized mutanome vaccines and implemented an RNA-based poly-neo-epitope approach to mobilize immunity against a spectrum of cancer mutations. Here we report the first-in-human application of this concept in melanoma. We set up a process comprising comprehensive identification of individual mutations, computational prediction of neo-epitopes, and design and manufacturing of a vaccine unique for each patient. All patients developed T cell responses against multiple vaccine neo-epitopes at up to high single-digit percentages. Vaccine-induced T cell infiltration and neo-epitope-specific killing of autologous tumour cells were shown in post-vaccination resected metastases from two patients. The cumulative rate of metastatic events was highly significantly reduced after the start of vaccination, resulting in a sustained progression-free survival. Two of the five patients with metastatic disease experienced vaccine-related objective responses. One of these patients had a late relapse owing to outgrowth of ß2-microglobulin-deficient melanoma cells as an acquired resistance mechanism. A third patient developed a complete response to vaccination in combination with PD-1 blockade therapy. Our study demonstrates that individual mutations can be exploited, thereby opening a path to personalized immunotherapy for patients with cancer.

Dynamic QoS Management for a Flexible 5G/6G Network Core: A Step toward a Higher Programmability.

The academic and professional community has recently started to develop the concept of 6G networks. The scientists have defined key performance indicators and pursued large-scale automation, ambient sensing intelligence, and pervasive artificial intelligence. They put great efforts into implementing new network access and edge computing solutions. However, further progress depends on developing a more flexible core infrastructure according to more complex QoS requirements. Our research aims to provide 5G/6G core flexibility by customizing and optimizing network slices and introducing a higher level of programmability. We bind similar services in a group, manage them as a single slice, and enable a higher level of programmability as a prerequisite for dynamic QoS. The current 5G solutions primarily use predefined queues, so we have developed highly flexible, dynamic queue management software and moved it entirely to the application layer (reducing dependence on the physical network infrastructure). Further, we have emulated a testbed environment as realistically as possible to verify the proposed model capabilities. Obtained results confirm the validity of the proposed dynamic QoS management model for configuring queues' parameters according to the service management requirements. Moreover, the proposed solution can also be applied efficiently to 5G core networks to resolve complex service requirements.

Novel basic-region helix-loop-helix transcription factor (AnBH1) of Aspergillus nidulans counteracts the CCAAT-binding complex AnCF in the promoter of a penicillin biosynthesis gene.

Cis-acting CCAAT elements are found frequently in eukaryotic promoter regions. Many of the genes containing such elements in their promoters are regulated by a conserved multimeric CCAAT-binding complex. In the fungus Emericella (Aspergillus) nidulans, this complex was designated AnCF (A.nidulans CCAAT-binding factor). AnCF regulates several genes, including the penicillin biosynthesis genes ipnA and aatA. Since it is estimated that the CCAAT-binding complex regulates more than 200 genes, an important question concerns the regulation mechanism that allows so many genes to be regulated by a single complex in a gene-specific manner. One of the answers to this question appears to lie in the interaction of AnCF with other transcription factors. Here, a novel transcription factor designated AnBH1 was isolated. The corresponding anbH1 gene was cloned and found to be located on chromosome IV. The deduced AnBH1 protein belongs to the family of basic-region helix-loop-helix (bHLH) transcription factors. AnBH1 binds in vitro as a homodimer to an, not previously described, asymmetric E-box within the aatA promoter that overlaps with the AnCF-binding site. This is the first report demonstrating that the CCAAT-binding complex and a bHLH transcription factor bind to overlapping sites. Since deletion of anbH1 appears to be lethal, the anbH1 gene was replaced by a regulatable alcAp-anbH1 gene fusion. The analysis of aatAp-lacZ expression in such a strain indicated that AnBH1 acts as a repressor of aatA gene expression and therefore counteracts the positive action of AnCF.

Prothymosin alpha associates with the oncoprotein SET and is involved in chromatin decondensation.

Prothymosin alpha (ProTalpha) is a histone H1-binding protein that interacts with the transcription coactivator CREB-binding protein and potentiates transcription. Based on coimmunoprecipitation and mammalian two-hybrid assays, we show here that ProTalpha forms a complex with the oncoprotein SET. ProTalpha efficiently decondenses human sperm chromatin, while overexpression of GFP-ProTalpha in mammalian cells results in global chromatin decondensation. These results indicate that decondensation of compacted chromatin fibers is an important step in the mechanism of ProTalpha function.

The histone chaperone SET/TAF-Ibeta interacts functionally with the CREB-binding protein.

The oncoprotein SET/TAF-Ibeta is a histone chaperone which is involved in cell-cycle control and chromatin remodeling. Confocal laser scanning microscopy reveals that SET is localized in distinct foci of variable size throughout the nucleoplasm of interphase cells. We report here that SET interacts directly with the acetyltransferase CREB-binding protein (CBP) and enhances the transactivation potential of the transcription coactivator. Our data suggest that the histone chaperone SET regulates the CBP-mediated transcription and may indicate a general principle by which transcriptional regulators cooperate with histone chaperones for gene activation.

Parathymosin affects the binding of linker histone H1 to nucleosomes and remodels chromatin structure.

Linker histone H1 is the major factor that stabilizes higher order chromatin structure and modulates the action of chromatin-remodeling enzymes. We have previously shown that parathymosin, an acidic, nuclear protein binds to histone H1 in vitro and in vivo. Confocal laser scanning microscopy reveals a nuclear punctuate staining of the endogenous protein in interphase cells, which is excluded from dense heterochromatic regions. Using an in vitro chromatin reconstitution system under physiological conditions, we show here that parathymosin (ParaT) inhibits the binding of H1 to chromatin in a dose-dependent manner. Consistent with these findings, H1-containing chromatin assembled in the presence of ParaT has reduced nucleosome spacing. These observations suggest that interaction of the two proteins might result in a conformational change of H1. Fluorescence spectroscopy and circular dichroism-based measurements on mixtures of H1 and ParaT confirm this hypothesis. Human sperm nuclei challenged with ParaT become highly decondensed, whereas overexpression of green fluorescent protein- or FLAG-tagged protein in HeLa cells induces global chromatin decondensation and increases the accessibility of chromatin to micrococcal nuclease digestion. Our data suggest a role of parathymosin in the remodeling of higher order chromatin structure through modulation of H1 interaction with nucleosomes and point to its involvement in chromatin-dependent functions.

A basic-region helix-loop-helix protein-encoding gene (devR) involved in the development of Aspergillus nidulans.

Basic-region helix-loop-helix (bHLH) proteins form an interesting class of eukaryotic transcription factors often involved in developmental processes. Here, a so far unknown bHLH protein-encoding gene of the filamentous ascomycete Aspergillus nidulans was isolated and designated devR for regulator of development. Deletion of devR revealed that the gene is non-essential for vegetative growth. However, the deletion mutant produced wrinkled colonies, a yellow pigment and did not form conidia on minimal agar plates. Conidiophore development was initiated normally, and colonies produced conidiophores with metulae and phialides. However, the phialides continued to grow filamentously and produced a second conidiophore with a vesicle at its end. The addition of KCl (0.6 M) to the medium suppressed the knock-out phenotype. The DeltadevR phenotype resembled that of a mutation in the tcsA gene encoding a histidine kinase domain and a response regulator domain. Here, we generated a tcsA deletion mutant. In a DeltatcsA strain, a DevR-Egfp protein fusion was detected in the cytoplasm, whereas in the wild type, the protein fusion was exclusively located in the nuclei, indicating that TcsA is required for nuclear localization of DevR. devR mRNA steady-state levels were similar in sporulating and vegetatively growing mycelia, and independent of a functional brlA gene. Moreover, under all conditions tested, self-crossing of the DeltadevR mutant strain was never observed. Taken together, devR encodes a bHLH regulatory protein that is part of the tcsA signal transduction network and required for development under standard growth conditions.