ABSTRACT
The Warburg effect is ubiquitous in proliferative mammalian cells, including cancer cells, but poses challenges for biopharmaceutical production, as lactate accumulation inhibits cell growth and protein production. Previous efforts to eliminate lactate production via knockout have failed in mammalian bioprocessing since lactate dehydrogenase has proven essential. However, here we eliminated the Warburg effect in Chinese hamster ovary (CHO) and HEK293 cells by simultaneously knocking out lactate dehydrogenase and regulators involved in a negative feedback loop that typically inhibits pyruvate conversion to acetyl-CoA. In contrast to long-standing assumptions about the role of aerobic glycolysis, Warburg-null cells maintain wildtype growth rate while producing negligible lactate. Further characterization of Warburg-null CHO cells showed a compensatory increase in oxygen consumption, a near total reliance on oxidative metabolism, and higher cell densities in fed-batch cell culture. These cells remained amenable for production of diverse biotherapeutic proteins, reaching industrially relevant titers and maintaining product glycosylation. Thus, the ability to eliminate the Warburg effect is an important development for biotherapeutic production and provides a tool for investigating a near-universal metabolic phenomenon.
ABSTRACT
Objectives: During their domestic quarantine, Covid-19 patients face major physical, psychological and social challenges. The description of support needs and specific topics brought to supportive conversations will be used to add to the body of knowledge about stressors and resources. Methods: A total of 109 telephone conversations with 69 quarantined Corona patients were documented by psychotherapists and physicians at Heidelberg University Hospital from November 2020 to April 2021. Subsequently, clinical documentations were analyzed according to a qualitative content analysis. Results: Most physical complaints related to cardio-respiratory symptoms (29 %), previous illnesses (24 %), and exhaustion or fatigue (16 %). On the psychological level, patients reported mainly anxiety (31 %) and depressive symptoms (16 %). On a social level, patients described stress related to family (56 %), work (20 %), and time in quarantine (16 %). Social support, individual coping strategies, a positive prognosis on the course of the corona disease, psychotherapy, and satisfactory medical care were mentioned as relieving factors. Therapeutic interventions aimed at stabilization and consisted of psychoeducation, relaxation techniques, and general counseling. Conclusions: The study shows that physical complaints, psychological symptoms, and social factors are brought into telephone support conversations. The support offer met a high demand and was well accepted.
Subject(s)
COVID-19 , Quarantine , Anxiety/psychology , COVID-19/epidemiology , Humans , Psychophysiologic Disorders , Quarantine/psychology , TelephoneABSTRACT
Despite the essential role secretory IgAs play in the defense against pathogenic invasion and the proposed value of recombinant secretory IgAs as novel therapeutics, currently there are no IgA-based therapies in clinics. Secretory IgAs are complex molecules and the major bottleneck limiting their therapeutic potential is a reliable recombinant production system. In this report, we addressed this issue and established a fast and robust production method for secretory IgAs in CHO-K1 cells using BAC-based expression vectors. As a proof of principle, we produced IgAs against Clostridium difficile toxins TcdA and TcdB. Recombinant secretory IgAs produced using our expression system showed comparable titers to IgGs, widely used as therapeutic biologicals. Importantly, secretory IgAs produced using our method were functional and could efficiently neutralize Clostridium difficile toxins TcdA and TcdB. These results show that recombinant secretory IgAs can be efficiently produced, thus opening the possibility to use them as therapeutic agents in clinics.