The focus of the Hotchkiss laboratory is defining new therapeutic approaches to sepsis, a highly lethal disorder which occurs during severe overwhelming infection. Our group was the first to demonstrate sepsis-induced massive, programmed cell death (apoptosis) of immune effector cells in patients dying of the disorder. Recently, our laboratory has concentrated on:
- identifying molecular mechanisms of immunosuppression during sepsis
- developing effective immune-adjuvant therapies for sepsis
Sepsis is the most common cause of death in most ICUs in developed countries
> 220,000 deaths annually due to sepsis in the United States alone
Septic patients develop a state of “immunoparalysis” rendering them unable to eradicate the primary initiating infection and making them highly susceptible to secondary hospital acquired infections.
The current therapeutic approach is multi-pronged and includes administration of the pleuripotent cytokine IL-7 and modulation of negative co-stimulatory pathways that induce immunosuppression using anti-programmed cell death-1. We have formed close working collaborations with colleagues at a number of major academic institutions and are organizing sepsis clinical trials using IL-7 and anti-PD-L1. Our sepsis clinical trials group is partnering with Bristol Meyers Squibb in testing of anti-PD-L1 and with Revimmune in testing of IL-7.
Apoptosis as a major cause of cell death in sepsis:
To determine why patients with sepsis were dying, our laboratory performed the first detailed autopsies of patients dying of sepsis or non-sepsis etiologies conducted in the patients’ ICU beds shortly after they expired. Cell death was characterized by multiple independent methods in lung, heart, kidney, liver, spleen, and intestines. We identified apoptosis of immune effector cells and gastrointestinal epithelial cells as a major cause of cell death and showed that despite significant organ failure, little necrotic cell death was observed. Thus, we postulated that cellular hibernation was a key cell response to sepsis in heart and kidney. These findings have been confirmed in two studies in pediatric and neonatal patients dying of sepsis. The widespread death of cells of the innate and adaptive immune system is the central pathophysiologic event in sepsis because of the systemic effects on the functioning of the immune system.
We performed a detailed immuno-phenotyping of lung and spleen immune cells in patients dying of sepsis. In addition to extensive loss of CD4 and CD8 T cells, other mechanisms of immune suppression were identified including:
- Increased T regulatory cells
- Up-regulation of negative co-stimulatory molecules
- Down-regulation of positive co-stimulatory molecules
- T cell “exhaustion”
Mechanisms of apoptosis and the role of immune suppression in sepsis-induced morbidity and mortality:
At the forefront of identifying mechanisms of immune suppression in sepsis, our lab characterized a profound depletion of innate and adaptive immune cells. To establish the causal link between sepsis-induced immunosuppression and mortality, we demonstrated how prevention of sepsis-induced apoptosis of immune effector cells improved survival in animal models of sepsis. Mice that overexpress anti-apoptotic molecules or have knockout of pro-apoptotic molecules were employed in a study investigating the contribution of the death receptor and mitochondrial mediated pathways (Faseb J. 21:708 2007). Numerous other laboratories have shown that prevention of apoptosis by a variety of independent methods results in improved survival in sepsis. In addition to apoptosis, our lab is currently examining the role and inter-relationship of autophagy and apoptosis in sepsis.
Immunotherapy to inhibit sepsis-induced immune effector cell apoptosis.
Our group is examining methods to reengage the immune effector cells of the innate and adaptive immune system. Strategies include blockade of negative co-stimulator molecules and administration of immunostimulatory cytokines. Anti-apoptotic cytokines act to prevent cell death by increasing anti-apoptotic Bcl-2 family members and decrease pro-apoptotic Bcl-2 family members. We have shown the ability of several cytokines to block cell death in lymphocytes, dendritic cells, and natural killer cells. We are currently testing several compounds in selected models of sepsis.
Our studies, conducted over a ten-year period and evaluating over a dozen therapies showed that both IL-7, a lymphocyte growth factor and anti-apoptotic cytokine, and anti-PD-1, a checkpoint inhibitor, were highly efficacious in restoring immunity and improving survival in multiple animal models of sepsis. Similarly, both immune therapies were effective in decreasing lymphocyte death and enhancing lymphocyte function when added ex vivo to blood samples from septic patients.
We initiated phase 2 clinical trials of IL-7 and anti-PD-1 in sepsis. Results demonstrated that these therapies were well-tolerated and ameliorated the sepsis-induced immune suppression. We also reported that IL-7 and anti-PD-1 were efficacious when used on a compassionate basis in patients with intractable life-threatening infections who were not responsive to their existing therapies (Lancet Infect Dis. 17:18:2017, Open Forum Infect Dis. 8: (6), ofab256). A publication describing the emerging potential for immunotherapy as an adjunct to antibiotic therapy in infectious diseases was co-authored by the PI and appeared in the New England Journal of Medicine. Our sepsis clinical trials group is partnering with Bristol Meyers Squibb in testing of anti-PD-L1 and with Revimmune in testing of IL-7.
COVID-19: Host immunity and immunotherapies
Our laboratory rapidly mobilized to address the COVID crisis and was one of only a few research laboratories at Washington University Medical Center to remain fully operational throughout the pandemic. This was possible due to the dedication and support of my coworkers and my ongoing clinical responsibilities as an ICU physician, caring for critically ill COVID and non-COVID patients. We performed detailed immunologic studies on blood samples from several hundred ICU patients with COVID (JCI Insight. 2020 Sep 3;5(17):e140329; Sci Adv. 2020;6(50):eabe3024). In addition, I helped write and design the protocol for compassionate use of IL-7 in critically-ill COVID patients with lymphopenia (JAMA Network Open 2020;3:e2016485). I am the PI in the United States for the ongoing double-blind, randomized, placebo-controlled clinical trial of IL-7 in lymphopenic COVID patients that has enrolled 100 patients (NCT04501796). Currently, our work has resulted in seven COVID-related publications including several in high impact journals.
Development of the ELISpot assay as a method to guide immune therapies in sepsis.
To effectively apply immune therapies in sepsis, we must identify the presence and severity of immune suppression. The PI and his colleagues developed a novel ELISpot assay that measures the frequency and robustness of cytokine-secreting cells in patient diluted whole blood. The ELISpot assay has been used to immune phenotype patients with sepsis and patients with COVID-19 with the results reported in two prestigious journals. Washington University School of Medicine, the University of Cincinnati School Of Medicine, and the University of Florida, Gainesville, School of Medicine hold a joint patent on the ELISpot assay. A patent has been jointly applied for ELISpot immune phenotyping by the investigators at the 3 different academic centers.
Development of “humanized mice” for testing therapies
The study of human disease is limited due to ethical and technical concerns. The development of “humanized” mice provides researchers with the remarkable ability to study human cells and tissues in an in vivo setting. “Humanized” mice are made by engrafting human CD34+ hematopoietic stem cells into immuno-deficient mice. After engraftment, mice develop human myeloid and lymphoid lineages, providing a surrogate to study the effects of infection, autoimmunity, transplantation, etc. on the human immune system. Our lab has generated “humanized” mice, and we are employing these mice for many of the sepsis studies.