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Introduction: Atypical hemolytic uremic syndrome (aHUS) poses a significant health challenge due to its rarity and severity within the spectrum of thrombotic microangiopathy. Despite efforts to optimize and personalize health care for patients with aHUS, understanding the individual experiences, needs, and desires of patients with aHUS and their relatives remains limited., Methods: Here, we present a nationwide, exploratory, qualitative interview study with a direct content analysis approach. In-depth interviews and a 6-week evaluation were audio-recorded and conducted using a semistructured topic guide, based on the Institute for Positive Health (IPH) model., Results: Analysis of 10 interviews involving 6 patients with aHUS and 13 relatives revealed the prevalence of long-term disease symptoms in adult patients, notably fatigue, which significantly impacted daily functioning. Moreover, the resilience demonstrated by patients and their relatives was noteworthy; however, the acute phase of aHUS and the unpredictable nature of disease recurrence could profoundly affect mental well-being. The emotional toll of aHUS is pervasive, with feelings of fear, guilt, and trauma persisting across disease phases in both patients and relatives. Challenges in medical care, including delays in diagnosis and the need for personalized and uniform protocols, were highlighted. Support was deemed crucial, indicating the necessity for enhancements in the accessibility to comprehensible disease information and psychological counseling. Finally, complexities surrounding genetic testing and carriership were discussed., Conclusion: This study underscores the profound, enduring, and multifaced impact of aHUS. The insights gleaned from the experiences and needs of patients with aHUS and their relatives could lay the foundation for development and implementation of more personalized innovations in aHUS health care., (© 2024 International Society of Nephrology. Published by Elsevier Inc.)

Introduction: The introduction of eculizumab has improved the outcome in patients with atypical hemolytic uremic syndrome (aHUS). The optimal treatment strategy is debated. Here, we report the results of the CUREiHUS study, a 4-year prospective, observational study monitoring unbiased eculizumab discontinuation in Dutch patients with aHUS after 3 months of therapy., Methods: All pediatric and adult patients with aHUS in native kidneys and a first-time eculizumab treatment were evaluated. In addition, an extensive cost-consequence analysis was conducted., Results: A total of 21 patients were included in the study from January 2016 to October 2020. In 17 patients (81%), a complement genetic variant or antibodies against factor H were identified. All patients showed full recovery of hematological thrombotic microangiopathy (TMA) parameters after the start of eculizumab. A renal response was noted in 18 patients. After a median treatment duration of 13.6 weeks (range 2.1-43.9), eculizumab was withdrawn in all patients. During follow-up (80.7 weeks [0.0-236.9]), relapses occurred in 4 patients. Median time to first relapse was 19.5 (14.3-53.6) weeks. Eculizumab was reinitiated within 24 hours in all relapsing patients. At last follow-up, there were no chronic sequelae, i.e., no clinically relevant increase in serum creatinine (sCr), proteinuria, and/or hypertension in relapsing patients. The low sample size and event rate did not allow to determine predictors of relapse. However, relapses only occurred in patients with a likely pathogenic variant. The cost-effectiveness analysis revealed that the total medical expenses of our population were only 30% of the fictive expenses that would have been made when patients received eculizumab every fortnight., Conclusion: It is safe and cost-effective to discontinue eculizumab after 3 months of therapy in patients with aHUS in native kidneys. Larger data registries are needed to determine factors associated with suboptimal kidney function recovery during eculizumab treatment, factors to predict relapses, and long-term outcomes of eculizumab discontinuation., (© 2022 Published by Elsevier Inc. on behalf of the International Society of Nephrology.)

Complex cellular targets such as G protein-coupled receptors (GPCRs), ion channels, and other multi-transmembrane proteins represent a significant challenge for therapeutic antibody discovery, primarily because of poor stability of the target protein upon extraction from cell membranes. To assess whether a limited set of membrane-bound antigen formats could be exploited to identify functional antibodies directed against such targets, we selected a GPCR of therapeutic relevance (CCR1) and identified target binders using an in vitro yeast-based antibody discovery platform (Adimab TM ) to expedite hit identification. Initially, we compared two different biotinylated antigen formats overexpressing human CCR1 in a 'scouting' approach using a subset of the antibody library. Binders were isolated using streptavidin-coated beads, expressed as yeast supernatants, and screened using a high-throughput binding assay and flow cytometry on appropriate cell lines. The most suitable antigen was then selected to isolate target binders using the full library diversity. This approach identified a combined total of 183 mAbs with diverse heavy chain sequences. A subset of clones exhibited high potencies in primary cell chemotaxis assays, with IC 50 values in the low nM/high pM range. To assess the feasibility of any further affinity enhancement, full-length hCCR1 protein was purified, complementary-determining region diversified libraries were constructed from a high and lower affinity mAb, and improved binders were isolated by fluorescence-activated cell sorting selections. A significant affinity enhancement was observed for the lower affinity parental mAb, but not the high affinity mAb. These data exemplify a methodology to generate potent human mAbs for challenging targets rapidly using whole cells as antigen and define a route to the identification of affinity-matured variants if required.

Recombinant antibodies are becoming increasingly important in the field of proteomics. Recent advances include the development of large phage-antibody libraries that contain high-affinity binders to almost any target protein, and new methods for high-throughput selection of antibody-antigen interactions. Coupled with a range of new screening technologies that use high-density antibody arrays to identify differentially expressed proteins, these antibody libraries can be applied to whole proteome analysis.

We have developed a novel technique for high-throughput screening of recombinant antibodies, based on the creation of antibody arrays. Our method uses robotic picking and high-density gridding of bacteria containing antibody genes followed by filter-based enzyme-linked immunosorbent assay (ELISA) screening to identify clones that express binding antibody fragments. By eliminating the need for liquid handling, we can thereby screen up to 18,342 different antibody clones at a time and, because the clones are arrayed from master stocks, the same antibodies can be double spotted and screened simultaneously against 15 different antigens. We have used our technique in several different applications, including isolating antibodies against impure proteins and complex antigens, where several rounds of phage display often fail. Our results indicate that antibody arrays can be used to identify differentially expressed proteins.