Allogeneic anti-CD19 CAR T cells can effectively treat B-cell malignancies that progress after alloHSCT. The findings point toward a future when antigen-specific T-cell therapies will play a central role in alloHSCT.

Purpose T cells genetically modified to express chimeric antigen receptors (CARs) targeting CD19 (CAR-19) have potent activity against acute lymphoblastic leukemia, but fewer results supporting treatment of lymphoma with CAR-19 T cells have been published. Patients with lymphoma that is chemotherapy refractory or relapsed after autologous stem-cell transplantation have a grim prognosis, and new treatments for these patients are clearly needed. Chemotherapy administered before adoptive T-cell transfer has been shown to enhance the antimalignancy activity of adoptively transferred T cells. Patients and Methods We treated 22 patients with advanced-stage lymphoma in a clinical trial of CAR-19 T cells preceded by low-dose chemotherapy. Nineteen patients had diffuse large B-cell lymphoma, two patients had follicular lymphoma, and one patient had mantle cell lymphoma. Patients received a single dose of CAR-19 T cells 2 days after a low-dose chemotherapy conditioning regimen of cyclophosphamide plus fludarabine. Results The overall remission rate was 73% with 55% complete remissions and 18% partial remissions. Eleven of 12 complete remissions are ongoing. Fifty-five percent of patients had grade 3 or 4 neurologic toxicities that completely resolved. The low-dose chemotherapy conditioning regimen depleted blood lymphocytes and increased serum interleukin-15 (IL-15). Patients who achieved a remission had a median peak blood CAR cell level of 98/μL and those who did not achieve a remission had a median peak blood CAR cell level of 15/μL ( P = .027). High serum IL-15 levels were associated with high peak blood CAR cell levels ( P = .001) and remissions of lymphoma ( P < .001). Conclusion CAR-19 T cells preceded by low-dose chemotherapy induced remission of advanced-stage lymphoma, and high serum IL-15 levels were associated with the effectiveness of this treatment regimen. CAR-19 T cells will likely become an important treatment for patients with relapsed lymphoma.

Cancer immunotherapeutic approaches induce tumor-specific immune responses, in particular CTL responses, in many patients treated. However, such approaches are clinically beneficial to only a few patients. We set out to investigate one possible explanation for the failure of CTLs to eliminate tumors, specifically, the concept that this failure is not dependent on inhibition of T cell function. In a previous study, we found that in mice, myeloidderived suppressor cells (MDSCs) are a source of the free radical peroxynitrite (PNT). Here, we show that pretreatment of mouse and human tumor cells with PNT or with MDSCs inhibits binding of processed peptides to tumor cell-associated MHC, and as a result, tumor cells become resistant to antigen-specific CTLs. This effect was abrogated in MDSCs treated with a PNT inhibitor. In a mouse model of tumor-associated inflammation in which the antitumor effects of antigen-specific CTLs are eradicated by expression of IL-1β in the tumor cells, we determined that therapeutic failure was not caused by more profound suppression of CTLs by IL-1β-expressing tumors than tumors not expressing this proinflammatory cytokine. Rather, therapeutic failure was a result of the presence of PNT. Clinical relevance for these data was suggested by the observation that myeloid cells were the predominant source of PNT in human lung, pancreatic, and breast cancer samples. Our data therefore suggest what we believe to be a novel mechanism of MDSC-mediated tumor cell resistance to CTLs.

T cells expressing anti-CD19 chimeric antigen receptors (CARs) can induce complete remissions (CRs) of diffuse large B cell lymphoma (DLBCL). The long-term durability of these remissions is unknown. We administered anti-CD19 CAR T cells preceded by cyclophosphamide and fludarabine conditioning chemotherapy to patients with relapsed DLBCL. Five of the seven evaluable patients obtained CRs. Four of the five CRs had long-term durability with durations of remission of 56, 51, 44, and 38 months; to date, none of these four cases of lymphomas have relapsed. Importantly, CRs continued after recovery of non-malignant polyclonal B cells in three of four patients with long-term complete remissions. In these three patients, recovery of CD19 polyclonal B cells took place 28, 38, and 28 months prior to the last follow-up, and each of these three patients remained in CR at the last follow-up. Non-malignant CD19 B cell recovery with continuing CRs demonstrated that remissions of DLBCL can continue after the disappearance of functionally effective anti-CD19 CAR T cell populations. Patients had a low incidence of severe infections despite long periods of B cell depletion and hypogammaglobulinemia. Only one hospitalization for an infection occurred among the four patients with long-term CRs. Anti-CD19 CAR T cells caused long-term remissions of chemotherapy-refractory DLBCL without substantial chronic toxicities.

Purpose Adoptive transfer of genetically modified T cells is being explored as a treatment for patients with metastatic cancer. Most current strategies use genes that encode major histocompatibility complex (MHC) class I-restricted T-cell receptors (TCRs) or chimeric antigen receptors to genetically modify CD8 T cells or bulk T cells for treatment. Here, we evaluated the safety and efficacy of an adoptive CD4 T-cell therapy using an MHC class II-restricted, HLA-DPB1*0401-restricted TCR that recognized the cancer germline antigen, MAGE-A3 (melanoma-associated antigen-A3). Patients and Methods Patients received a lymphodepleting preparative regimen, followed by adoptive transfer of purified CD4 T cells, retrovirally transduced with MAGE-A3 TCR plus systemic high-dose IL-2. A cell dose escalation was conducted, starting at 10 total cells and escalating at half-log increments to approximately 10 cells. Nine patients were treated at the highest dose level (0.78 to 1.23 × 10 cells). Results Seventeen patients were treated. During the cell dose-escalation phase, an objective complete response was observed in a patient with metastatic cervical cancer who received 2.7 × 10 cells (ongoing at ≥ 29 months). Among nine patients who were treated at the highest dose level, objective partial responses were observed in a patient with esophageal cancer (duration, 4 months), a patient with urothelial cancer (ongoing at ≥ 19 months), and a patient with osteosarcoma (duration, 4 months). Most patients experienced transient fevers and the expected hematologic toxicities from lymphodepletion pretreatment. Two patients experienced transient grade 3 and 4 transaminase elevations. There were no treatment-related deaths. Conclusion These results demonstrate the safety and efficacy of administering autologous CD4 T cells that are genetically engineered to express an MHC class II-restricted antitumor TCR that targets MAGE-A3. This clinical trial extends the reach of TCR gene therapy for patients with metastatic cancer.

Tumor-associated myeloid cells are the major type of inflammatory cells involved in the regulation of anti-tumor immune responses. One key characteristic of these cells is the generation of reactive oxygen (ROS) and reactive nitrogen (RNS) species in the tumor microenvironment. Recent studies have demonstrated the important role of ROS and RNS, especially peroxynitrite (PNT), in immune suppression in cancer. ROS and RNS are involved in induction of antigen-specific T-cell tolerance, inhibition of T-cell migration to the tumor site, and tumor cell evasion of recognition by cytotoxic T cells. In pre-clinical settings, a number of potential therapeutic agents demonstrated activity in blocking ROS/RNS in cancer and in improving the efficacy of cancer immune therapy. A better understanding of ROS/RNS-associated pathways in myeloid cells will help to identify more specific and direct targets to facilitate the development of more effective immune therapy of cancer.

Marijuana cannabinoids, such as ␦-9-tetrahydrocannabinoid (THC), suppress type 1 T-helper 1 (Th1) immunity in a variety of models, including infection with the intracellular pathogen Legionella pneumophila (Lp). To examine the cellular mechanism of this effect, bone marrow-derived dendritic cells (DCs) were purified from BALB/c mice and studied following infection and drug treatment. DCs infected in vitro with Lp were able to protect mice when injected prior to a lethal Lp infection; however, the immunization potential of the Lp-loaded cells along with Th1 cytokine production was attenuated by THC treatment at the time of in vitro infection. In addition, THC-treated and Lp-loaded DCs were poorly stimulated in culture-primed splenic CD4ϩ T cells to produce interferon-␥; however, this stimulating deficiency was reversed by adding recombinant interleukin (IL)-12p40 protein to the cultures. Moreover, THC treatment inhibited the expression of DC maturation markers, such as major histocompatibility complex class II and costimulatory molecules CD86 and CD40 as determined by flow cytometry and suppressed the Notch ligand, Delta4, as determined by reverse transcription-polymerase chain reaction. However, THC treatment did not affect other DC functions, such as intracellular killing of Lp, determined by colony-forming unit counts of bacteria, and Lp-induced apoptosis, determined by annexin V staining. In conclusion, the data suggest that THC inhibits Th1 activation by targeting essential DC functions, such as IL-12p40 secretion, maturation, and expression of costimulatory and polarizing molecules.

The treatment of B-cell malignancies by adoptive cell transfer (ACT) of anti-CD19 chimeric antigen receptor T cells (CD19 CAR-T) has proven to be a highly successful therapeutic modality in several clinical trials. The anti-CD19 CAR-T cell production method used to support initial trials relied on numerous manual, open process steps, human serum, and 10 days of cell culture to achieve a clinical dose. This approach limited the ability to support large multicenter clinical trials, as well as scale up for commercial cell production. Therefore, studies were completed to streamline and optimize the original National Cancer Institute production process by removing human serum from the process in order to minimize the risk of viral contamination, moving process steps from an open system to functionally closed system operations in order to minimize the risk of microbial contamination, and standardizing additional process steps in order to maximize process consistency. This study reports a procedure for generating CD19 CAR-T cells in 6 days, using a functionally closed manufacturing process and defined, serum-free medium. This method is able to produce CD19 CAR-T cells that are phenotypically and functionally indistinguishable from cells produced for clinical trials by the previously described production process.