Non-invasive monitoring of neoadjuvant radiation therapy response in soft tissue sarcomas by multiparametric MRI and quantification of circulating tumor DNA-A study protocol.

BACKGROUND: Wide resection remains the cornerstone of localized soft-tissue sarcomas (STS) treatment. Neoadjuvant radiation therapy (NRT) may decrease the risk of local recurrences; however, its effectiveness for different histological STS subtypes has not been systematically investigated. The proposed prospective study evaluates the NRT response in STS using liquid biopsies and the correlation of multiparametric magnetic resonance imaging (mpMRI) with histopathology and immunohistochemistry. METHODS: Patients with localized high-grade STS, who qualify for NRT, are included in this study. LIQUID BIOPSIES: Quantification of circulating tumor DNA (ctDNA) in patient blood samples is performed by targeted next-generation sequencing. Soft-tissue sarcoma subtype-specific panel sequencing in combination with patient-specific exome sequencing allows the detection of individual structural variants and point mutations. Circulating free DNA is isolated from peritherapeutically collected patient plasma samples and ctDNA quantified therein. Identification of breakpoints is carried out using FACTERA. Bioinformatic analysis is performed using samtools, picard, fgbio, and the MIRACUM Pipeline. MPMRI: Combination of conventional MRI sequences with diffusion-weighted imaging, intravoxel-incoherent motion, and dynamic contrast enhancement. Multiparametric MRI is performed before, during, and after NRT. We aim to correlate mpMRI data with the resected specimen's macroscopical, histological, and immunohistochemical findings. RESULTS: Preliminary data support the notion that quantification of ctDNA in combination with tumor mass characterization through co-registration of mpMRI and histopathology can predict NRT response of STS. CLINICAL RELEVANCE: The methods presented in this prospective study are necessary to assess therapy response in heterogeneous tumors and lay the foundation of future patient- and tumor-specific therapy concepts. These methods can be applied to various tumor entities. Thus, the participation and support of a wider group of oncologic surgeons are needed to validate these findings on a larger patient cohort.

CRP Enhances the Innate Killing Mechanisms Phagocytosis and ROS Formation in a Conformation and Complement-Dependent Manner.

Phagocytosis and the formation of reactive oxygen species (ROS) in phagocytic leukocytes are an effective killing mechanism of the innate host defense. These cellular processes of innate immunity function in a complex interplay with humoral factors. C-reactive protein (CRP) in its activated, monomeric isoform (mCRP) has been shown to activate immune cells via the classical complement pathway. We investigated the complement-dependent effects of monomeric CRP (mCRP) on neutrophils and monocyte subtypes using complement-specific inhibitors by both flow cytometry and confocal fluorescence microscopy. We demonstrate that CRP-induced ROS generation is a conformation-specific and complement-dependent process in leukocyte subsets with classical monocytes as the primary source of ROS amongst human monocyte subsets. Elucidation of this complex interplay of CRP and complement in inflammation pathophysiology might help to improve anti-inflammatory therapeutic strategies.

Dissociation of C-Reactive Protein Localizes and Amplifies Inflammation: Evidence for a Direct Biological Role of C-Reactive Protein and Its Conformational Changes.

C-reactive protein (CRP) is a member of the pentraxin superfamily that is widely recognized as a marker of inflammatory reactions and cardiovascular risk in humans. Recently, a growing body of data is emerging, which demonstrates that CRP is not only a marker of inflammation but also acts as a direct mediator of inflammatory reactions and the innate immune response. Here, we critically review the various lines of evidence supporting the concept of a pro-inflammatory "CRP system." The CRP system consists of a functionally inert circulating pentameric form (pCRP), which is transformed to its highly pro-inflammatory structural isoforms, pCRP* and ultimately to monomeric CRP (mCRP). While retaining an overall pentameric structure, pCRP* is structurally more relaxed than pCRP, thus exposing neoepitopes important for immune activation and complement fixation. Thereby, pCRP* shares its pro-inflammatory properties with the fully dissociated structural isoform mCRP. The dissociation of pCRP into its pro-inflammatory structural isoforms and thus activation of the CRP system occur on necrotic, apoptotic, and ischemic cells, regular ß-sheet structures such as ß-amyloid, the membranes of activated cells (e.g., platelets, monocytes, and endothelial cells), and/or the surface of microparticles, the latter by binding to phosphocholine. Both pCRP* and mCRP can cause activation of platelets, leukocytes, endothelial cells, and complement. The localization and deposition of these pro-inflammatory structural isoforms of CRP in inflamed tissue appear to be important mediators for a range of clinical conditions, including ischemia/reperfusion (I/R) injury of various organs, cardiovascular disease, transplant rejection, Alzheimer's disease, and age-related macular degeneration. These findings provide the impetus to tackle the vexing problem of innate immunity response by targeting CRP. Understanding the "activation process" of CRP will also likely allow the development of novel anti-inflammatory drugs, thereby providing potential new immunomodulatory therapeutics in a broad range of inflammatory diseases.