Amy B. Hahn

Sperm protein 17 (Sp17) in multiple myeloma: opportunity for myeloma‐specific donor T cell infusion to enhance graft‐versus‐myeloma effect without increasing graft‐versus‐host disease risk

We recently found that sperm protein 17 (Sp17), a spermatozoa‐restricted protein, is aberrantly expressed on the tumor cells in patients with multiple myeloma (MM). It may therefore be possible to generate donor‐derived Sp17‐specific CTL for administration following allogeneic stem cell transplant to augment graft‐versus‐myeloma (GVM) effect without inducing a global GVHD. To assess thisapproach, we have produced recombinant Sp17 protein and used Sp17 protein‐pulsed dendritic cells to generate HLA class I‐restricted Sp17‐specific CTL from a previously unimmunized healthy donor. These CTL were able to lyse autologous Epstein‐Barr virus‐transformed lymphoblastoid cells in a Sp17‐dependent manner. Target lysis was HLA‐A1 and HLA‐B27 restricted. Cytotoxicity could be blocked by antibodies against monomorphic HLA class I, HLA‐A1 and HLA‐B27 molecules but not HLA class II molecules. Most importantly, the CTL lysed HLA class I‐matched Sp17‐positive tumor cells, suggesting that Sp17 is processed and presented in association with the HLA class I molecules in Sp17‐positive tumor cells in a concentration and configuration that could be recognized by recombinant protein‐primed CTL. Analysis by flow cytometry of the CTL indicated that they were predominantly CD8 in phenotype and they produced IFN‐γ and very little IL‐4. Our results suggest the potential for the generation andadministration of donor‐derived Sp17‐specific CTL to augment GVM without inducing GVHD following allogeneic stem cell transplant for MM.

Failure of ganciclovir prophylaxis to completely eradicate CMV disease in renal transplant recipients treated with intense anti-rejection immunotherapy

Ganciclovir prophylactic regimens have been shown to be effective in renal transplant recipients at risk for primary (donor seropositive/recipient seronegative) and secondary (recipient seropositive) cytomegalovirus (CMV) disease. However, in addition to serologic factors, the type and intensity of the administered immunosuppression is a strong risk factor for CMV disease. Since January 1995, we have utilized a potent immunosuppressive protocol selectively in recipients at high risk for immunologic graft loss, defined as retransplant recipients, recipients with delayed graft function, non‐Caucasian recipients, and recipients suffering from acute rejection. 
Between January 1995 and December 1996, 110 consecutive renal transplants were performed in recipients who were either CMV seropositive or received an allograft from a CMV‐seropositive donor. All recipients received ganciclovir prophylactic therapy for 3 months post‐transplant. Group I (N=43) consisted of recipients at high‐immunologic risk for graft loss as defined above. These recipients were treated with an intense anti‐rejection immunotherapeutic regimen consisting of Cellcept, Neoral, and prednisone, with the frequent addition of antilymphocyte antibody therapies and intravenous methylprednisolone. The remaining 67 recipients (group II) were treated with a less intense immunotherapeutic regimen consisting of azathioprine, Neoral, and prednisone. The incidence and severity of CMV disease and the patient and allograft survival were compared. 
The incidence of CMV syndrome was greater in group I (28%) compared with group II (7%), and was statistically significant (p<0.05). The 1‐yr patient and graft survival were similar, 95 and 91%, respectively, for group I compared with 97 and 97%, respectively, for group II. 
These data suggest that 3 months of ganciclovir prophylactic therapy is significantly less effective for the prevention of CMV disease in renal transplant recipients at high risk for acute rejection treated with an intense immunotherapeutic regimen. These data suggest that more effective prevention of CMV disease in these high‐risk recipients will require the addition of other anti‐viral agents, such as immunoglobulin preparation to the prophylactic regimen.

Immunological Functions of the Membrane Proximal Region of MHC Class II Molecules

Major histocompatibility complex (MHC) class II molecules present exogenously derived antigen peptides to CD4 T cells, driving activation of naïve T cells and supporting CD4-driven immune functions. However, MHC class II molecules are not inert protein pedestals that simply bind and present peptides. These molecules also serve as multi-functional signaling molecules delivering activation, differentiation, or death signals (or a combination of these) to B cells, macrophages, as well as MHC class II-expressing T cells and tumor cells. Although multiple proteins are known to associate with MHC class II, interaction with STING (stimulator of interferon genes) and CD79 is essential for signaling. In addition, alternative transmembrane domain pairing between class II α and β chains influences association with membrane lipid sub-domains, impacting both signaling and antigen presentation. In contrast to the membrane-distal region of the class II molecule responsible for peptide binding and T-cell receptor engagement, the membrane-proximal region (composed of the connecting peptide, transmembrane domain, and cytoplasmic tail) mediates these “non-traditional” class II functions. Here, we review the literature on the function of the membrane-proximal region of the MHC class II molecule and discuss the impact of this aspect of class II immunobiology on immune regulation and human disease.

HLA-A2 reactive antibodies in a patient who types as HLA-A2: The importance of high resolution typing and epitope-based antibody analysis

This report describes a case of a highly sensitized patient who had serum antibodies reacting with HLA-A2 but whose phenotype included HLA-A2. The determination of HLA mismatch acceptability at the antigen level was problematic, but high-resolution HLA typing information and epitope-based antibody specificity analysis based on the nonself-self paradigm of HLA epitope immunogenicity have provided a solution. This case supports the concept that HLA typing at the allele level offers a better approach to identifying suitable donors for sensitized patients.

The calcineurin inhibitor RCAN1 is involved in cultured macrophage and in vivo immune response

Studies on the role of regulator of calcineurin 1 (RCAN1) in immunity are limited, but have demonstrated an involvement in T-lymphocyte function. Here, we expand these studies to macrophages and in vivo infection. The treatment of RAW and primary mouse macrophages with lipopolysaccharide from Escherichia coli strongly induced RCAN1 isoform 4 (RCAN1-4), but not isoform 1. RCAN1-4 induction involved calcium, calcineurin, and reactive oxygen species. Subsequent analysis with whole bacteria including gram-negative E. coli and gram-positive Staphylococcus aureus revealed strong RCAN1-4 inductions by both, and where tested, dependence on calcium. Staphylococcus aureus cell wall components peptidoglycan and lipoteichoic acid also strongly induced RCAN1-4. In vivo, a significant induction in the proinflammatory cytokines monocyte chemotactic protein-1, interleukin-6, interferon-γ, and tumor necrosis factor-α was observed in knockout (KO) lung vs. wild-type (WT) mice 7 days after nasal infection with Fransicella tularensis. This induction was not accompanied by a significant increase in F. tularensis burden in the KO lung. Additionally, a modest increase in respiratory burst activity in KO vs. WT macrophages was observed. Combined, these studies indicate that RCAN1 is involved in macrophage and the overall in vivo immune response, and provide additional evidence that RCAN1 plays an important role in cell immunity and infectious disease.

The new kidney allocation system does not equally advantage all very high cPRA candidates – A single center analysis

The new UNOS kidney allocation system awards very high points to candidates with cPRA 99% and 100%, and allows for national sharing for cPRA 100% candidates. We sought to determine the effect of this new kidney allocation system on candidates who are very highly sensitized (90-98% cPRA) but not eligible for very high points or national sharing by examining offers to these candidates for 5months pre-implementation and two consecutive 5month periods post-implementation and comparing them to cPRA⩾99% candidates. We found that the cPRA⩾99% candidates received significantly more offers and transplants after implementation, while offers and transplants to the 90-98% candidates decreased. A slight adjustment to the allocation system may be needed to provide more equitable distribution of kidneys to all high cPRA candidates.

CIITA Enhances HIV-1 Attachment to CD4+ T Cells Leading to Enhanced Infection and Cell Depletion

Activated CD4+ T cells are more susceptible to HIV infection than resting T cells; the reason for this remains unresolved. Induction of CIITA and subsequent expression of the MHC class II isotype HLA-DR are hallmarks of CD4+ T cell activation; therefore, we investigated the role of CIITA expression in T cells during HIV infection. CIITA-expressing SupT1 cells display enhanced virion attachment in a gp160/CD4-dependent manner, which results in increased HIV infection, virus release, and T cell depletion. Although increased attachment and infection of T cells correlated with HLA-DR surface expression, Ab blocking, transient expression of HLA-DR without CIITA, and short hairpin RNA knockdown demonstrate that HLA-DR does not directly enhance susceptibility of CIITA-expressing cells to HIV infection. Further analysis of the remaining MHC class II isotypes, HLA-DP and HLA-DQ, MHC class I isotypes, HLA-A, HLA-B, and HLA-C, and the class II Ag presentation genes, invariant chain and HLA-DM, demonstrate that these proteins likely do not contribute to CIITA enhancement of HIV infection. Finally, we demonstrate that in activated primary CD4+ T cells as HLA-DR/CIITA expression increases there is a corresponding increase in virion attachment. Overall, this work suggests that induction of CIITA expression upon CD4+ T cell activation contributes to enhanced attachment, infection, virus release, and cell death through an undefined CIITA transcription product that may serve as a new antiviral target.