Epitope-engineered human hematopoietic stem cells are shielded from CD123-targeted immunotherapy

Targeted eradication of transformed or otherwise dysregulated cells using monoclonal antibodies (mAb), antibody-drug conjugates (ADC), T cell engagers (TCE), or chimeric antigen receptor (CAR) cells is very effective for hematologic diseases. Unlike the breakthrough progress achieved for B cell malignancies, there is a pressing need to find suitable antigens for myeloid malignancies. CD123, the interleukin-3 (IL-3) receptor alpha-chain, is highly expressed in various hematological malignancies, including acute myeloid leukemia (AML). However, shared CD123 expression on healthy hematopoietic stem and progenitor cells (HSPCs) bears the risk for myelotoxicity. We demonstrate that epitope-engineered HSPCs were shielded from CD123-targeted immunotherapy but remained functional, while CD123-deficient HSPCs displayed a competitive disadvantage. Transplantation of genome-edited HSPCs could enable tumor-selective targeted immunotherapy while rebuilding a fully functional hematopoietic system. We envision that this approach is broadly applicable to other targets and cells, could render hitherto undruggable targets accessible to immunotherapy, and will allow continued posttransplant therapy, for instance, to treat minimal residual disease (MRD)

In Silico and Biochemical Analysis of Physcomitrella patens Photosynthetic Antenna: Identification of Subunits which Evolved upon Land Adaptation

Background. In eukaryotes the photosynthetic antenna system is composed of subunits encoded by the light harvesting complex (Lhc) multigene family. These proteins play a key role in photosynthesis and are involved in both light harvesting and photoprotection. The moss Physcomitrella patens is a member of a lineage that diverged from seed plants early after land colonization and therefore by studying this organism, we may gain insight into adaptations to the aerial environment. Principal Findings. In this study, we characterized the antenna protein multigene family in Physcomitrella patens, by sequence analysis as well as biochemical and functional investigations. Sequence identification and analysis showed that some antenna polypeptides, such as Lhcb3 and Lhcb6, are present only in land organisms, suggesting they play a role in adaptation to the sub-aerial environment. Our functional analysis which showed that photo-protective mechanisms in Physcomitrella patens are very similar to those in seed plants fits with this hypothesis. In particular, Physcomitrella patens also activates Non Photochemical Quenching upon illumination, consistent with the detection of an ortholog of the PsbS protein. As a further adaptation to terrestrial conditions, the content of Photosystem I low energy absorbing chlorophylls also increased, as demonstrated by differences in Lhca3 and Lhca4 polypeptide sequences, in vitro reconstitution experiments and low temperature fluorescence spectra. Conclusions. This study highlights the role of Lhc family members in environmental adaptation and allowed proteins associated with mechanisms of stress resistance to be identified within this large family

Optimization of insect cell based protein production processes - online monitoring, expression systems, scale-up

Due to the increasing use of insect cell based expression systems in research and industrial recombinant protein production, the development of efficient and reproducible production processes remains a challenging task. In this context, the application of online monitoring techniques is intended to ensure high and reproducible product qualities already during the early phases of process development. In the following chapter, the most common transient and stable insect cell based expression systems are briefly introduced. Novel applications of insect cell based expression systems for the production of insect derived antimicrobial peptides/proteins (AMPs) are discussed using the example of G. mellonella derived gloverin. Suitable in situ sensor techniques for insect cell culture monitoring in disposable and common bioreactor systems are outlined with respect to optical and capacitive sensor concepts. Since scale-up of production processes is one of the most critical steps in process development, a conclusive overview is given about scale up aspects for industrial insect cell culture processes

Guidelines and Recommendations for Laboratory Analysis in the Diagnosis and Management of Diabetes Mellitus

Background: Multiple laboratory tests are used to diagnose and manage patients with diabetes mellitus. The quality of the scientific evidence supporting the use of these tests varies substantially. Approach: An expert committee compiled evidence-based recommendations for the use of laboratory testing for patients with diabetes. A new system was developed to grade the overall quality of the evidence and the strength of the recommendations. Draft guidelines were posted on the Internet and presented at the 2007 Arnold O. Beckman Conference. The document was modified in response to oral and written comments, and a revised draft was posted in 2010 and again modified in response to written comments. The National Academy of Clinical Biochemistry and the Evidence-Based Laboratory Medicine Committee of the American Association for Clinical Chemistry jointly reviewed the guidelines, which were accepted after revisions by the Professional Practice Committee and subsequently approved by the Executive Committee of the American Diabetes Association. Content: In addition to long-standing criteria based on measurement of plasma glucose, diabetes can be diagnosed by demonstrating increased blood hemoglobin A$_{1c}$ (HbA$_{1c}$) concentrations. Monitoring of glycemic control is performed by self-monitoring of plasma or blood glucose with meters and by laboratory analysis of HbA$_{1c}$. The potential roles of noninvasive glucose monitoring, genetic testing, and measurement of autoantibodies, urine albumin, insulin, proinsulin, C-peptide, and other analytes are addressed. Summary: The guidelines provide specific recommendations that are based on published data or derived from expert consensus. Several analytes have minimal clinical value at present, and their measurement is not recommended