Oncogene-Targeting T Cells Reject Large Tumors while Oncogene Inactivation Selects Escape Variants in Mouse Models of Cancer

SummaryThe genetic instability of cancer cells frequently causes drug resistance. We established mouse cancer models, which allowed targeting of an oncogene by drug-mediated inactivation or monospecific CD8+ effector T (TE) cells. Drug treatment of genetically unstable large tumors was effective but selected resistant clones in the long term. In contrast, TE cells completely rejected large tumors (≥500 mm3), if the target antigen was cancer-driving and expressed in sufficient amounts. Although drug-mediated oncogene inactivation selectively killed the cancer cells and left the tumor vasculature intact, which likely facilitated survival and growth of resistant clones, TE cell treatment led to blood vessel destruction and probably “bystander” elimination of escape variants, which did not require antigen cross-presentation by stromal cells

Vaccination with a plasmid DNA encoding HER-2/neu together with low doses of GM-CSF and IL-2 in patients with metastatic breast carcinoma: a pilot clinical trial

<p>Abstract</p> <p>Background</p> <p>Adjuvant trastuzumab (Herceptin) treatment of breast cancer patients significantly improves their clinical outcome. Vaccination is an attractive alternative approach to provide HER-2/neu (Her2)-specific antibodies and may in addition concomitantly stimulate Her2-reactive T-cells. Here we report the first administration of a Her2-plasmid DNA (pDNA) vaccine in humans.</p> <p>Patients and Methods</p> <p>The vaccine, encoding a full-length signaling-deficient version of the oncogene Her2, was administered together with low doses of GM-CSF and IL-2 to patients with metastatic Her2-expressing breast carcinoma who were also treated with trastuzumab. Six of eight enrolled patients completed all three vaccine cycles. In the remaining two patients treatment was discontinued after one vaccine cycle due to rapid tumor progression or disease-related complications. The primary objective was the evaluation of safety and tolerability of the vaccine regimen. As a secondary objective, treatment-induced Her2-specific immunity was monitored by measuring antibody production as well as T-cell proliferation and cytokine production in response to Her2-derived antigens.</p> <p>Results</p> <p>No clinical manifestations of acute toxicity, autoimmunity or cardiotoxicity were observed after administration of Her2-pDNA in combination with GM-CSF, IL-2 and trastuzumab. No specific T-cell proliferation following <it>in vitro </it>stimulation of freshly isolated PBMC with recombinant human Her2 protein was induced by the vaccination. Immediately after all three cycles of vaccination no or even decreased CD4⁺T-cell responses towards Her2-derived peptide epitopes were observed, but a significant increase of MHC class II restricted T-cell responses to Her2 was detected at long term follow-up. Since concurrent trastuzumab therapy was permitted, λ-subclass specific ELISAs were performed to specifically measure endogenous antibody production without interference by trastuzumab. Her2-pDNA vaccination induced and boosted Her2-specific antibodies that could be detected for several years after the last vaccine administration in a subgroup of patients.</p> <p>Conclusion</p> <p>This pilot clinical trial demonstrates that Her2-pDNA vaccination in conjunction with GM-CSF and IL-2 administration is safe, well tolerated and can induce long-lasting cellular and humoral immune responses against Her2 in patients with advanced breast cancer.</p> <p>Trial registration</p> <p>The trial registration number at the Swedish Medical Products Agency for this trial is Dnr151:785/2001.</p

Immune and gene therapies for cancer and infectious diseases

Annually more than 20 million deaths are globally caused because of infections or cancer. This accounts for 41.5% of annual deaths. New techniques are supposed to constitute our tools in the fight against these types of deaths. The new technologies of immune and gene therapies promise to revolutionize medicine in the next century. Active immune and gene therapies include both non-specific (such as cytokines and synthetic chemicals) and specific (i.e. vaccination) approaches. Cytokines when delivered by genetic approaches to tumors were reported to affect their capacity to present antigens and to act as targets for natural killer (NK) cells and cytotoxic T cells (CTL). In the first part of my thesis, I have studied the mechanism by which the cytokine interleukin- 10 (IL- 10) alters tumor cell sensitivity to NK and CTL mediated cytotoxicity and affects MHC class I antigen presentation in particular peptide transportation by the transporter associated with antigen presentation (TAP). The delivery of a gene in a suitable expression plasmid to cells in vivo allows the immune system to recognize its product leading to protective immunity against infectious agents or tumors, a process termed plasmid DNA (pDNA) vaccination. In the second part of my thesis I have studied pDNA vaccination and methods to improve this mode of immunization by cytokine genes and by tucaresol a Schiff forming drug. To this end, I have established two clinically relevant animal models. In one, a gene from the tumor associated Epsteine Bar virus (EBV) nuclear antigen 4 (EBNA-4) gene was used, while in the other, the mycobacterial heat-shock protein 65 (Mhsp65) was tested as a pDNA vaccine. In these two models I analyzed the cytokine genes interferon gamma (IFN-[gamma]) and granulocyte macrophage colony stimulating factor (GM-CSF) for their ability to enhance pDNA induced immune responses. In the same models I also tested the effect of the novel immune response modifying adjuvant, tucaresol. In this part of my thesis I have also established the possibility of using HER-2 based immunotherapy for the treatment of human tumors overexpressing this gene product. These studies have provided three candidate genes (EBNA-4, Mhsp65 and HER-2), Peptides, cytokines and adjuvant that could be used for vaccination or to treat patients against cancer and viral and bacterial diseases. pDNA or peptide vaccines based on these could lead to protection from a large range of diseases including tuberculosis, leprosy, typhoid fever, brucellosis, infectious mononucleosis, EBV associated tumors, carcinomas and melanoma. I have shown that cytokines and adjuvants could be used to efficiently enhance the immune response to pDNA vaccines resulting in full protection from tumor outgrowth. An insight into the mechanism by which IL-10 modulates the immune response was presented. I have been able to show how IL-10 affects the MHC-I antigen-processing pathway resulting in alteration of NK and CTL responses

Marked Enhancement of the Antigen-Specific Immune Response by Combining Plasmid DNA-Based Immunization with a Schiff Base-Forming Drug

Although plasmid DNA (pDNA)-based immunization has proven efficacy, the level of immune responses that is achieved by this route of vaccination is often lower than that induced by traditional vaccines, especially for primates and humans. We report here a simple and potent method to enhance pDNA-based vaccination by using two different plasmids encoding viral or bacterial antigens. This method is based on coadministration of low concentrations of a recently described immunopotentiating, Schiff base-forming drug called tucaresol which has led to significant augmentation of antigen-specific humoral and cellular immune responses. Our data suggest that enhancement of the immune response with tucaresol might provide a powerful tool for the further development of pDNA-based immunization for humans

Efficacy of CAR T-cell Therapy in Large Tumors Relies upon Stromal Targeting by IFN gamma

Adoptive T-cell therapy using chimeric antigen receptor-modified T cells (CAR-T therapy) has shown dramatic efficacy in patients with circulating lymphoma. However, eradication of solid tumors with CAR-T therapy has not been reported yet to be efficacious. In solid tumors, stroma destruction, due to MHC-restricted cross-presentation of tumor antigens to T cells, may be essential. However, CAR-Ts recognize antigens in an MHC-independent manner on cancer cells but not stroma cells. In this report, we show how CAR-Ts can be engineered to eradicate large established tumors with provision of a suitable CD28 costimulatory signal. In an HER2-dependent tumor model, tumor rejection by HER2-specific CAR-Ts was associated with sustained influx and proliferation of the adoptively transferred T cells. Interestingly, tumor rejection did not involve natural killer cells but was associated instead with a marked increase in the level of M1 macrophages and a requirement for IFN gamma receptor expression on tumor stroma cells. Our results argue that CAR-T therapy is capable of eradicating solid tumors through a combination of antigen-independent stroma destruction and antigen-specific tumor cell targeting. (C) 2014 AACR