Expression of tissue-specific homing molecules directs antigen-experienced T cells to particular peripheral tissues. In studies using soluble antigens that focused on skin and gut, antigen-presenting cells (APCs) within regional lymphoid tissues were proposed to be responsible for imprinting homing phenotypes. Whether this occurs in other sites and after physiologic antigen processing and presentation is unknown. We define in vivo imprinting of distinct homing phenotypes on monospecific T cells responding to antigens expressed by tumors in intracerebral, subcutaneous, and intraperitoneal sites with efficient brain-tropism of CD8 T cells crossprimed in the cervical lymph nodes (LNs). Multiple imprinting programs could occur simultaneously in the same LN when tumors were present in more than one site. Thus, the identity of the LN is not paramount in determining the homing phenotype; this critical functional parameter is dictated upstream at the site of antigen capture by crosspresenting APCs.

During the priming phase of an antitumor immune response, CD8+ T cells undergo a program of differentiation driven by professional APCs in secondary lymphoid organs. This leads to clonal expansion and acquisition both of effector functions and a specific adhesion molecule pattern. Whether this program can be reshaped during the effector phase to adapt to the effector site microenvironment is unknown. We investigated this in murine brain tumor models using adoptive transfer of tumor-specific CD8+ T cells, and in spontaneous immune responses of patients with malignant glioma. Our data show proliferation of Ag-experienced tumor-specific T cells within the brain parenchyma. Moreover, CD8+ T cells further differentiated in the brain, exhibiting enhanced IFN-γ and granzyme B expression and induction of αE(CD103)β7 integrin. This unexpected integrin expression identified a subpopulation of CD8+ T cells conditioned by the brain microenvironment and also had functional consequences: αE(CD103)β7-expressing CD8+ T cells had enhanced retention in the brain. These findings were further investigated for CD8+ T cells infiltrating human malignant glioma; CD8+ T cells expressed αE(CD103)β7 integrin and granzyme B as in the murine models. Overall, our data indicate that the effector site plays an active role in shaping the effector phase of tumor immunity. The potential for local expansion and functional reprogramming should be considered when optimizing future immunotherapies for regional tumor control.

Vaccines that can coordinately induce multi-epitope T cellmediated immunity, T helper functions, and immunologic memory may offer effective tools for cancer immunotherapy. Here, we report the development of a new class of recombinant protein cancer vaccines that deliver different CD8 þ and CD4 þ T-cell epitopes presented by MHC class I and class II alleles, respectively. In these vaccines, the recombinant protein is fused with Z12, a novel cell-penetrating peptide that promotes efficient protein loading into the antigen-processing machinery of dendritic cells. Z12 elicited an integrated and multi-epitopic immune response with persistent effector T cells. Therapy with Z12-formulated vaccines prolonged survival in three robust tumor models, with the longest survival in an orthotopic model of aggressive brain cancer. Analysis of the tumor sites showed antigen-specific T-cell accumulation with favorable modulation of the balance of the immune infiltrate. Taken together, the results offered a preclinical proof of concept for the use of Z12-formulated vaccines as a versatile platform for the development of effective cancer vaccines. Cancer Res; 75(15);

The number and localization of effector cells to the tumor site are crucial elements for immune rejection of solid tumors. However, for cerebral malignancies, antitumor responses need to be finely tuned to avoid neuropathologic consequences. In this study, we determine factors that regulate CTL localization and tumoricidal function after intracerebral implantation of tumors expressing model Ag. H-2bxd mice implanted with a CW3+ murine glioma lacking H-2Kd molecules necessary to present the CW3170–179 epitope demonstrate cross-priming of H-2Kd-restricted CTL, and moreover, Ag-dependent accumulation of functional H-2Kd/CW3170–179-specific CTL within the tumor bed. This implicates a role for cross-presentation not only in priming, but also in retention of fully differentiated CTL in the tumor stroma at the effector stage of the response. Modulating cross-presentation of Ag may be the key in regulating specific immune responses in the brain: either by augmenting protective responses or by down-modulating destructive autoimmune reactions.