Antibodies (inhibitors) developed by hemophilia B patients against coagulation factor IX (FIX) are challenging to eliminate because of anaphylaxis or nephrotic syndrome after continued infusion. To address this urgent unmet medical need, FIX fused with a transmucosal carrier (CTB) was produced in a commercial lettuce (Simpson Elite) cultivar using species specific chloroplast vectors regulated by endogenous psbA sequences. CTB-FIX (~1mg/g) in lyophilized cells was stable with proper folding, disulfide bonds and pentamer assembly when stored ~2 years at ambient temperature. Feeding lettuce cells to hemophilia B mice delivered CTB-FIX efficiently to the gut immune system, induced LAP+ regulatory T cells and suppressed inhibitor/IgE formation and anaphylaxis against FIX. Lyophilized cells enabled 10-fold dose escalation studies and successful induction of oral tolerance was observed in all tested doses. Induction of tolerance in such a broad dose range should enable oral delivery to patients of different age groups and diverse genetic background. Using Fraunhofer cGMP hydroponic system, ~870 kg fresh or 43.5 kg dry weight can be harvested per 1000 ft2 per annum yielding 24,000–36,000 doses for 20-kg pediatric patients, enabling first commercial development of an oral drug, addressing prohibitively expensive purification, cold storage/transportation and short shelf life of current protein drugs.

Both CD4+ and CD8+ T cells are important for successful immunity to tuberculosis and have redundant effector functions, such as cytolysis and release of potent antimycobacterial cytokines such as interferon-gamma and tumor necrosis factor-alpha. We hypothesized that CD8+ T cells play a unique role in host defense to Mycobacterium tuberculosis infection as well. Possibilities include preferential and/or enhanced release of granular constituents and/or preferential recognition of heavily infected cells. Utilizing human, Mycobacterium tuberculosis-specific, CD4+ and CD8+ T cell clones, we demonstrate that, after recognition of antigen-presenting cells displaying peptide antigen, CD4+ T cells preferentially release interferon-gamma, whereas CD8+ T cells preferentially lyse antigen-presenting cells. Furthermore, utilizing dendritic cells infected with Mycobacterium tuberculosis expressing green fluorescent protein, we show that CD8+ T cells preferentially recognize heavily infected cells that constitute the minority of infected cells. These data support the hypothesis that the central role of CD8+ T cells in the control of infection with Mycobacterium tuberculosis may be that of surveillance; in essence, recognition of cells in which the containment of Mycobacterium tuberculosis is no longer effective.

Previous studies in murine and human models have suggested an important role for HLA Ia-restricted CD8+ T cells in host defense to Mycobacterium tuberculosis (Mtb). Therefore, understanding the Ags presented via HLA-Ia will be important in understanding the host response to Mtb and in rational vaccine design. We have used monocyte-derived dendritic cells in a limiting dilution analysis to generate Mtb-specific CD8+ T cells. Two HLA-Ia-restricted CD8+ T cell clones derived by this method were selected for detailed analysis. One was HLA-B44 restricted, and the other was HLA-B14 restricted. Both were found to react with Mtb-infected, but not bacillus Calmette-Guérin-infected, targets. For both these clones, the Ag was identified as culture filtrate protein 10 (CFP10)/Mtb11, a 10.8-kDa protein not expressed by bacillus Calmette-Guérin. Both clones were inhibited by the anti-class I Ab and anti-HLA-B,C Abs. Using a panel of CFP10/Mtb11-derived 15-aa peptides overlapping by 11 aa, the region containing the epitopes for both clones has been defined. Minimal 10-aa epitopes were defined for both clones. CD8+ effector cells specific for these two epitopes are present at high frequency in the circulating pool. Moreover, the CD8+ T cell response to CFP10/Mtb11 can be largely accounted for by the two epitopes defined herein, suggesting that this is the immunodominant response for this purified protein derivative-positive donor. This study represents the first time CD8+ T cells generated against Mtb-infected APC have been used to elucidate an Mtb-specific CD8+ T cell Ag.

Development of a subunit vaccine for Mycobacterium tuberculosis (Mtb) is likely to be dependent on the identification of T cell antigens that induce strong proliferation and interferon γ production from healthy purified protein derivative (PPD)+ donors. We have developed a sensitive and rapid technique for screening an Mtb genomic library expressed in Escherichia coli using Mtb-specific CD4+ T cells. Using this technique, we identified a family of highly related Mtb antigens. The gene of one family member encodes a 9.9-kD antigen, termed Mtb9.9A. Recombinant Mtb9.9A protein, expressed and purified from E. coli, elicited strong T cell proliferation and IFN-γ production by peripheral blood mononuclear cells from PPD+ but not PPD− individuals. Southern blot analysis and examination of the Mtb genome sequence revealed a family of highly related genes. A T cell line from a PPD+ donor that failed to react with recombinant Mtb9.9A recognized one of the other family members, Mtb9.9C. Synthetic peptides were used to map the T cell epitope recognized by this line, and revealed a single amino acid substitution in this region when compared with Mtb9.9A. The direct identification of antigens using T cells from immune donors will undoubtedly be critical for the development of vaccines to several intracellular pathogens.

The development of an effective vaccine against Mycobacterium tuberculosis is a research area of intense interest. Mounting evidence suggests that protective immunity to M. tuberculosis relies on both MHC class II-restricted CD4+ T cells and MHC class I-restricted CD8+ T cells. By purifying polypeptides present in the culture filtrate of M. tuberculosis and evaluating these molecules for their ability to stimulate PBMC from purified protein derivative-positive healthy individuals, we previously identified a low-m.w. immunoreactive T cell Ag, Mtb 8.4, which elicited strong Th1 T cell responses in healthy purified protein derivative-positive human PBMC and in mice immunized with recombinant Mtb 8.4. Herein we report that Mtb 8.4-specific T cells can be detected in mice immunized with the current live attenuated vaccine, Mycobacterium bovis-bacillus Calmette-Guérin as well as in mice infected i.v. with M. tuberculosis. More importantly, immunization of mice with either plasmid DNA encoding Mtb 8.4 or Mtb 8.4 recombinant protein formulated with IFA elicited strong CD4+ T cell and CD8+ CTL responses and induced protection on challenge with virulent M. tuberculosis. Thus, these results suggest that Mtb 8.4 is a potential candidate for inclusion in a subunit vaccine against TB.

No abstract

ObjectiveSelf-compassion is consistently found to be related to better psychological outcomes.As most studies were cross-sectional, little is known about the predictive role of selfcompassion for future psychological outcomes. This longitudinal study in cancer patients investigated the predictive role of self-compassion at the time of cancer diagnosis for the course of symptoms of depression, anxiety, and fatigue in the period of receiving cancer treatment. Methods: This longitudinal study was conducted at the Shaanxi Provincial TumourHospital in Xi'an, China. A total of 153 heterogeneous cancer patients were assessed within 1 week after cancer diagnosis (T1) as well as at the start (T2) and the end (T3) of medical treatment. Hierarchical linear regression analyses were conducted to examine the research questions.Results: Cross-sectional regression analyses at T1 showed that a self-compassion total score and negative self-compassion (and to a lesser extent positive selfcompassion) were significantly related to symptoms of depression, anxiety, and fatigue. When controlling for symptoms at T1, positive self-compassion significantly predicted all three outcomes at T3. A self-compassion total score only predicted symptoms of anxiety at T2, controlling for T1 symptoms. In contrast, we found no significant predictive value of negative self-compassion. Conclusions: This study suggests that the positive aspects of self-compassion are beneficial for cancer patients for their future functioning, in terms of fewer symptoms of depression, anxiety, and fatigue over time. Future interventions should test how and to what extent self-compassion can be cultivated and whether increases in self-compassion are associated with better outcomes. KEYWORDS cancer, oncology, positive and negative self-compassion, depression, anxiety, fatigue, longitudinal study Lei Zhu and Juntao Yao share the first coauthorship.

Three distinct co-morbidity patterns of depression, anxiety and fatigue were identified and exhibited different symptom courses longitudinally. Those with poor physical health tended to report elevated mood disturbances and fatigue during psychological care. Copyright © 2016 John Wiley & Sons, Ltd.