Transitioning from manual to stirred-tank bioreactor manufacturing of IDCT, An allogeneiccell therapy to treat lumbar degenerative disc disease

DiscGenics is a clinical stage regenerative medicine company focused on developing cell therapies that alleviate pain and restore function in patients with degenerative disc disease (DDD), a major cause of low back pain which is a driver of disability worldwide. The Company’s lead product candidate, IDCT, is a homologous, allogeneic, off-the-shelf, injectable cell therapy under investigational use in the US (ClinicalTrials.gov Identifier: NCT03347708). The manufacturing process for IDCT involves isolating cells from donated intervertebral disc tissue and expanding them into proprietary progenitor cells known as discogenic cells. For preclinical and early clinical testing, cell production was a manual process which relied on pooling individual flasks to achieve the desired lot size. For successful scale-up and commercial production, DiscGenics seeks to modify the IDCT manufacturing process to utilize one large, single vessel per lot, while also applying bioprocess controls and more robust analytical methods to ensure consistent and optimal production of drug product. Once these changes are implemented, the product critical quality attributes (CQAs) must be maintained. DiscGenics has engaged GE Healthcare (GEHC) and the Centre for Commercialization of Regenerative Medicine (CCRM) for assay, media, and process development at the Centre for Advanced Therapeutic Cell Technologies (CATCT) in Toronto, ON., Canada. In partnership with the Federal Economic Development Agency for Southern Ontario (FedDev Ontario), CATCT accelerates the development, industrialization, and adoption of cell manufacturing technologies to improve patient access to cell and gene therapies. In this collaborative project, discogenic cells were generated in traditional static culture using CellStacks (Corning), in PBS-MINI bioreactor systems (PBS Biotech), and in stirred-tank reactors (STRs) (Eppendorf), which was led by the GEHC/CCRM team. Parameters such as cell viability, fold growth, and identity via flow cytometry were compared across modalities. For the STRs, multiple control parameters were evaluated to improve cell growth and assess successful maintenance of a consistent environment for cell quality. In this study, we found that we are able to maintain CQAs between the production modalities, with cell growth being significantly improved in the STR platform. In the STRs, in-process measurements of metabolites aligned with cell growth found using a custom sampling method. Increased cell expansion was facilitated by modified agitation, inoculation, and perfusion feeding strategies. Additionally, the process-controlled STRs provide non-invasive, continuous process data monitoring which allow for development of specified control ranges of manufacturing parameters. The quality by design (QbD) approach taken for the STR process development and improvement has allowed an increase in the lot size, process knowledge, and data-driven process definition. This presentation describes the approach and benefits of transitioning from a manual process to a suspension-based, process-controlled, stirred-tank reactor to produce allogeneic cell therapies

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

A Th1-inducing Adenoviral Vaccine for Boosting Adoptively Transferred T Cells

Although the benefits of adoptive T-cell therapy can be increased by prior lymphodepletion of the recipient, this process usually requires chemotherapy or radiation. Vaccination with antigens to which the transferred T cells respond should be a less toxic means of enhancing their activity, but to date such vaccines have not been effective. We, therefore, determined which characteristics an adenoviral vaccine has to fulfill to optimally activate and expand adoptively transferred antigen-specific T cells in vivo. We evaluated (i) antigen, (ii) flagellin, a Toll-like receptor (TLR) 5 ligand, and (iii) an inhibitor of the antigen-presenting attenuator A20. Vaccination of mice before T-cell transfer with a vaccine that contained all three components dramatically enhanced the effector function of ovalbumin (OVA)-specific T cells as judged by the regression of established B16-OVA tumors compared to one- and two-component vaccines. Immunization with the three-component vaccine induced a strong Th1 environment, which was critical for the observed synergy and proved as effective as cytoxan-induced lymphodepletion in enhancing in vivo T-cell expansion. Thus, the combination of our vaccine with T-cell therapy has the potential to enhance and broaden adoptive cellular immunotherapy

Immunotherapy for Osteosarcoma: Genetic Modification of T cells Overcomes Low Levels of Tumor Antigen Expression

Human epidermal growth factor receptor 2 (HER2) is expressed by the majority of human osteosarcomas and is a risk factor for poor outcome. Unlike breast cancer, osteosarcoma cells express HER2 at too low, a level for patients to benefit from HER2 monoclonal antibodies. We reasoned that this limitation might be overcome by genetically modifying T cells with HER2-specific chimeric antigen receptors (CARs), because even a low frequency of receptor engagement could be sufficient to induce effector cell killing of the tumor. HER2-specific T cells were generated by retroviral transduction with a HER2-specific CAR containing a CD28.ζ signaling domain. HER2-specific T cells recognized HER2-positive osteosarcoma cells as judged by their ability to proliferate, produce immunostimulatory T helper 1 cytokines, and kill HER2-positive osteosarcoma cell lines in vitro. The adoptive transfer of HER2-specific T cells caused regression of established osteosarcoma xenografts in locoregional as well as metastatic mouse models. In contrast, delivery of nontransduced (NT) T cells did not change the tumor growth pattern. Genetic modification of T cells with CARs specific for target antigens, expressed at too low a level to be effectively recognized by monoclonal antibodies, may allow immunotherapy to be more broadly applicable for human cancer therapy