Joseph G. Crompton

Inhibiting glycolytic metabolism enhances CD8+ T cell memory and antitumor function

Naive CD8+ T cells rely upon oxidation of fatty acids as a primary source of energy. After antigen encounter, T cells shift to a glycolytic metabolism to sustain effector function. It is unclear, however, whether changes in glucose metabolism ultimately influence the ability of activated T cells to become long-lived memory cells. We used a fluorescent glucose analog, 2-NBDG, to quantify glucose uptake in activated CD8+ T cells. We found that cells exhibiting limited glucose incorporation had a molecular profile characteristic of memory precursor cells and an increased capacity to enter the memory pool compared with cells taking up high amounts of glucose. Accordingly, enforcing glycolytic metabolism by overexpressing the glycolytic enzyme phosphoglycerate mutase-1 severely impaired the ability of CD8+ T cells to form long-term memory. Conversely, activation of CD8+ T cells in the presence of an inhibitor of glycolysis, 2-deoxyglucose, enhanced the generation of memory cells and antitumor functionality. Our data indicate that augmenting glycolytic flux drives CD8+ T cells toward a terminally differentiated state, while its inhibition preserves the formation of long-lived memory CD8+ T cells. These results have important implications for improving the efficacy of T cell-based therapies against chronic infectious diseases and cancer.

Akt Inhibition Enhances Expansion of Potent Tumor-Specific Lymphocytes with Memory Cell Characteristics

Adoptive cell therapy (ACT) using autologous tumor-infiltrating lymphocytes (TIL) results in complete regression of advanced cancer in some patients, but the efficacy of this potentially curative therapy may be limited by poor persistence of TIL after adoptive transfer. Pharmacologic inhibition of the serine/threonine kinase Akt has recently been shown to promote immunologic memory in virus-specific murine models, but whether this approach enhances features of memory (e.g., long-term persistence) in TIL that are characteristically exhausted and senescent is not established. Here, we show that pharmacologic inhibition of Akt enables expansion of TIL with the transcriptional, metabolic, and functional properties characteristic of memory T cells. Consequently, Akt inhibition results in enhanced persistence of TIL after adoptive transfer into an immunodeficient animal model and augments antitumor immunity of CD8 T cells in a mouse model of cell-based immunotherapy. Pharmacologic inhibition of Akt represents a novel immunometabolomic approach to enhance the persistence of antitumor T cells and improve the efficacy of cell-based immunotherapy for metastatic cancer.

Mitochondrial Membrane Potential Identifies Cells with Enhanced Stemness for Cellular Therapy.

Long-term survival and antitumor immunity of adoptively transferred CD8(+) T cells is dependent on their metabolic fitness, but approaches to isolate therapeutic T cells based on metabolic features are not well established. Here we utilized a lipophilic cationic dye tetramethylrhodamine methyl ester (TMRM) to identify and isolate metabolically robust T cells based on their mitochondrial membrane potential (ΔΨm). Comprehensive metabolomic and gene expression profiling demonstrated global features of improved metabolic fitness in low-ΔΨm-sorted CD8(+) T cells. Transfer of these low-ΔΨm T cells was associated with superior long-term in vivo persistence and an enhanced capacity to eradicate established tumors compared with high-ΔΨm cells. Use of ΔΨm-based sorting to enrich for cells with superior metabolic features was observed in CD8(+), CD4(+) T cell subsets, and long-term hematopoietic stem cells. This metabolism-based approach to cell selection may be broadly applicable to therapies involving the transfer of HSC or lymphocytes for the treatment of viral-associated illnesses and cancer.

Memory T cell–driven differentiation of naive cells impairs adoptive immunotherapy

Adoptive cell transfer (ACT) of purified naive, stem cell memory, and central memory T cell subsets results in superior persistence and antitumor immunity compared with ACT of populations containing more-differentiated effector memory and effector T cells. Despite a clear advantage of the less-differentiated populations, the majority of ACT trials utilize unfractionated T cell subsets. Here, we have challenged the notion that the mere presence of less-differentiated T cells in starting populations used to generate therapeutic T cells is sufficient to convey their desirable attributes. Using both mouse and human cells, we identified a T cell-T cell interaction whereby antigen-experienced subsets directly promote the phenotypic, functional, and metabolic differentiation of naive T cells. This process led to the loss of less-differentiated T cell subsets and resulted in impaired cellular persistence and tumor regression in mouse models following ACT. The T memory-induced conversion of naive T cells was mediated by a nonapoptotic Fas signal, resulting in Akt-driven cellular differentiation. Thus, induction of Fas signaling enhanced T cell differentiation and impaired antitumor immunity, while Fas signaling blockade preserved the antitumor efficacy of naive cells within mixed populations. These findings reveal that T cell subsets can synchronize their differentiation state in a process similar to quorum sensing in unicellular organisms and suggest that disruption of this quorum-like behavior among T cells has potential to enhance T cell-based immunotherapies.

Females have fewer complications and lower mortality following trauma than similarly injured males: A risk adjusted analysis of adults in the National Trauma Data Bank

BACKGROUND Studies of sexual dimorphism in trauma outcomes suggest that women have a survival advantage compared to equivalently injured men. It is unknown if this gender disparity is mediated by potentially life-threatening complications. OBJECTIVE To determine (1) if there is a sex-based differences in the odds of developing inpatient complications after trauma, and (2) if are these complications associated with death among trauma patients. METHODS Review of adult trauma patients admitted to hospitals in the National Trauma Data Bank that report complications. Patient and injury severity covariates were adjusted using multiple logistic regression and the independent effect of sex on developing complications and associated mortality was determined. RESULTS A total of 681,730 adult patients met the inclusion criteria of hospital admission > or =3 days. Women demonstrated a 21% lower adjusted risk of death compared to males (OR 0.79, 95% CI 0.76-0.83). Females had decreased adjusted odds of developing life-threatening complications including pneumonia, acute respiratory distress syndrome, acute renal failure and pulmonary embolism. However, when compared to males with life-threatening complications, females with complications were found to be at greater risk of dying. CONCLUSION This study demonstrates that women are less likely than men to develop inpatient complications, suggesting that the survival advantage among women after traumatic injury may involve a reduced susceptibility to developing life-threatening complications.

Uncoupling T-cell expansion from effector differentiation in cell-based immunotherapy.

Adoptive cellular immunotherapy (ACT) is a potentially curative therapy for patients with advanced cancer. Eradication of tumor in mouse models and humans correlates with both a high dose of adoptively transferred cells and cells with a minimally differentiated phenotype that maintain replicative capacity and multipotency. We speculate that response to ACT not only requires transfer of cells with immediate cytolytic effector function to kill the bulk of fast‐growing tumor but also transfer of tumor‐specific cells that maintain an ability for self‐renewal and the capacity to produce a continual supply of cytolytic effector progeny until all malignant cells are eliminated. Current in vitro methods to expand cells to sufficient numbers and still maintain a minimally differentiated phenotype are hindered by the biological coupling of clonal expansion and effector differentiation. Therefore, a better understanding of the physiologic mechanism that couples cell expansion and differentiation in CD8+ T cells may improve the efficacy of ACT.

Multiple imputation in trauma disparity research.

BACKGROUND Missing data has remained a major disparity in trauma outcomes research due to missing race and insurance data. Multiple imputation (M.IMP) has been recommended as a solution to deal with this major drawback. STUDY DESIGN Using the National Data Trauma Bank (NTDB) as an example, a complete dataset was developed by deleting cases with missing data across variables of interest. An incomplete dataset was then created from the complete set using random deletion to simulate the original NTDB, followed by five M.IMP rounds to generate a final imputed dataset. Identical multivariate analyses were performed to investigate the effect of race and insurance on mortality in both datasets. RESULTS Missing data proportions for known trauma mortality covariates were as follows: age-4%, gender-0.4%, race-8%, insurance-17%, injury severity score-6%, revised trauma score-20%, and trauma type-3%. The M.IMP dataset results were qualitatively similar to the original dataset. CONCLUSION M.IMP is a feasible tool in NTDB for handling missing race and insurance data.

Evidence of Hormonal Basis for Improved Survival Among Females With Trauma-associated Shock: An Analysis of the National Trauma Data Bank

BACKGROUND Basic science research suggests that sex hormones affect survival after traumatic shock. This study sought to determine the independent effect of gender on mortality among trauma patients in different hormone-related age groups. METHODS Review of severely injured trauma patients with shock included in the National Trauma Databank. Patients were stratified into three groups on the basis of likely hormonal status: prehormonal (age, 0-12 years), hormonal (age,13-64 years), and posthormonal (age, ≥ 65 years). Multiple logistic regression was used to analyze the independent effect of gender on mortality in each group, adjusting for anatomic and physiologic injury severity. RESULTS A total of 48,394 patients met our inclusion criteria (Injury Severity Score ≥ 16 and systolic blood pressure <90 mm Hg). Crude mortality was higher (p < 0.05) for males in all categories: prehormonal = 29% for males (n = 3,553) versus 24% for females (n = 1,831); hormonal = 34% for males (n = 26,778) versus 30% for females (n = 8,677) and posthormonal = 36% for males (n = 4,280) versus 31% for females (n = 3,275). After adjusting for covariates, women in the hormonally active group had a 14% decreased odds of death (0.86 [95% CI, 0.76-0.93]) compared with men. Females did not exhibit this survival advantage in the prehormonal (odds of death = 0.92 [0.74-1.14]) or posthormonal (odds of death = 0.90 [0.76-1.05]) groups. CONCLUSIONS Females aged between 13 and 64 years exhibit significantly lower mortality than males after trauma-associated shock. This outcome difference is lost at the extremes of age (preadolescent children and individuals aged ≥ 65 years) where the effects of sex hormones are absent or diminished. These findings suggest that hormonal differences play a role in the gender-based outcome disparities after traumatic shock.

Collapse of the Tumor Stroma is Triggered by IL-12 Induction of Fas

Engineering CD8⁺ T cells to deliver interleukin 12 (IL-12) to the tumor site can lead to striking improvements in the ability of adoptively transferred T cells to induce the regression of established murine cancers. We have recently shown that IL-12 triggers an acute inflammatory environment that reverses dysfunctional antigen presentation by myeloid-derived cells within tumors and leads to an increase in the infiltration of adoptively transferred antigen-specific CD8⁺ T cells. Here, we find that local delivery of IL-12 increased the expression of Fas within tumor-infiltrating macrophages, dendritic cells, and myeloid-derived suppressor cells (MDSC), and that these changes were abrogated in mice deficient in IL-12-receptor signaling. Importantly, upregulation of Fas in host mice played a critical role in the proliferation and antitumor activity of adoptively transferred IL-12-modified CD8⁺ T cells. We also observed higher percentages of myeloid-derived cell populations within tumors in Fas-deficient mice, indicating that tumor stromal destruction was dependent on the Fas death receptor. Taken together, these results describe the likely requirement for costimulatory reverse signaling through Fasl on T cells that successfully infiltrate tumors, a mechanism triggered by the induction of Fas expression on myeloid-derived cells by IL-12 and the subsequent collapse of the tumor stroma.

Cish actively silences TCR signaling in CD8+ T cells to maintain tumor tolerance

Palmer et al. find that Cish, a member of the SOCS family, is induced by TCR stimulation in CD8+ T cells and inhibits their functional avidity against tumor. The authors uncover a novel mechanism of suppression for a SOCS member.