Cell culture scale translation from a 24-well Single-Use miniature bioreactor and subsequent impact on product and broth quality

To accelerate cell culture process development, most companies have validated scale-down models of their pilot and manufacturing scale bioreactors. Advancing such mimics to even smaller scales requires the large scale engineering environment to be accurately recreated. Here we describe a single-use microwell methodology that accurately reproduces not only cell growth kinetics but also key attributes related to product quality and broth processability. The μ24 miniature bioreactor system enables system level control of agitation (by orbital shaking), with individual well control of pH, DO and temperature. Two distinct plate types are investigated, allowing for either headspace or direct gas sparging. An engineering characterisation was performed evaluating fluid mixing, gas transfer capacity and the dispersed gas phase. Cell culture is investigated using a model CHO DG44 cell line expressing a whole IgG1 mAb [1]. In addition, this work describes scale-up of μ24 results to conventional laboratory scale stirred tank bioreactors (2L) and use of the device for selection of robust and scaleable cell lines through evaluation of product quality. The ‘broth quality’ is also evaluated for primary clarification efficiency using an Ultra Scale-Down (USD) depth filtration rig that requires quantities of material compatible with those available from the miniature bioreactors. Apparent kLa values ranged between 3–22 hr-1 and 4–53 hr-1 for headspace aeration and direct gas sparging respectively. Mixing times were generally in the range 1–13 seconds and decreased with increasing shaking frequency (500–800 rpm). Direct gas sparging also helped to reduce tm values. Cultures performed with headspace aeration showed the highest VCD and antibody titres, whereas those operated with direct gas sparging showed cell growth kinetics and product titres that were more comparable to those found in a conventional 2L stirred bioreactor. Initial results also indicate that key product and broth processability attributes are maintained making the combination of μ24 and USD technologies useful tools in ‘Quality by Design’ driven cell culture process development

Impact of Pre-Existing Treatment with Statins on the Course and Outcome of Tick-Borne Encephalitis

OBJECTIVES: Although statins have anti-inflammatory and potentially also antimicrobial (including antiviral) activity, their therapeutic impact on infectious diseases is controversial. In this study, we evaluated whether pre-existing statin use influenced the course and outcome of tick-borne encephalitis. METHODS: To assess the influence of statin usage on the severity of acute illness and the outcome of tick-borne encephalitis, univariate and multivariable analyses were performed for 700 adult patients with tick-borne encephalitis of whom 77 (11%) were being treated with statins, and for 410 patients of whom 53 (13%) were receiving statins, respectively. RESULTS: Multivariable analyses found no statistically significant association between statin usage and having a milder acute illness. There was also no statistically significant benefit with respect to a favorable outcome defined by the absence of post-encephalitic syndrome (ORs for a favorable outcome at 6 months was 0.96, 95% CI: 0.46-2.04, P = 0.926; at 12 months 0.29, 95% CI: 0.06-1.33, P = 0.111; at 2-7 years after acute illness 0.44, 95% CI: 0.09-2.22, P = 0.321), by a reduction in the frequency of six nonspecific symptoms (fatigue, myalgia/arthralgia memory disturbances, headache, concentration disturbances, irritability) occurring during the 4 week period before the last examination, or by higher SF-36 scores in any of the eight separate domains of health as well as in the physical and mental global overall component. Furthermore, there were no significant differences between patients receiving statins and those who were not in the cerebrospinal fluid or serum levels for any of the 24 cytokines/chemokines measured. CONCLUSIONS: In this observational study, we could not prove that pre-existing use of statins affected either the severity of the acute illness or the long-term outcome of tick-borne encephalitis

Presentations from the annual meeting of the MIT/Industry Cooperative research Program.

Caption titleMay 199

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

Early Lyme Disease (Erythema Migrans) and Its Mimics (Southern Tick-Associated Rash Illness and Tick-Associated Rash Illness)

Erythema migrans, an expanding erythematous skin lesion that develops days to weeks following an Ixodes species tick bite, is the most common clinical manifestation of Lyme disease. Presentations in the United States differ somewhat from that in Europe, presumably because of the different etiologic agents. Diagnosis is based on the appearance of the skin lesion, rather than on laboratory testing. After treatment with an appropriate oral antibiotic for 10 to 14 days, the prognosis is excellent. Two conditions that cause a similar skin lesion following a tick bite, but are of unknown cause, are Southern tick-associated rash illness in the United States and tick-associated rash illness in Japan