Inhibition of glycosphingolipid synthesis with eliglustat in combination with immune checkpoint inhibitors in advanced cancers: preclinical evidence and phase I clinical trial.

Glycosphingolipids (GSLs) are abundantly expressed in cancer cells. The effects of GSL-targeted immunotherapies are not fully understood. Here, we show that the inhibition of GSL synthesis with the UDP-glucose ceramide glucosyltransferase inhibitor eliglustat can increase the exposure of the major histocompatibility complex (MHC) and tumour antigen peptides, enhancing the antitumour response of CD8+ T cells in a range of tumour models. We therefore conducted a proof-of-concept phase I trial on the combination of eliglustat and an anti-PD-1 antibody for the treatment of advanced cancers (NCT04944888). The primary endpoints were safety and feasibility, and the secondary endpoint was antitumor activity. All prespecified endpoints were met. Among the 31 enrolled patients, only 1 patient experienced a grade 3 adverse event (AE), and no grade 4 AEs were observed. The objective response rate was 22.6% and the disease control rate reached 71%. Of the 8 patients with proficient mismatch repair/microsatellite stable (pMMR/MSS) colorectal cancer, one achieved complete response and two each had partial response and stable disease. In summary, inhibiting the synthesis of GSLs might represent an effective immunotherapy approach.

Temporal single-cell tracing reveals clonal revival and expansion of precursor exhausted T cells during anti-PD-1 therapy in lung cancer.

Anti-PD-1 treatment has shown unprecedented clinical success in the treatment of non-small-cell lung cancer (NSCLC), but the underlying mechanisms remain incompletely understood. Here, we performed temporal single-cell RNA and paired T-cell receptor sequencing on 47 tumor biopsies from 36 patients with NSCLC following PD-1-based therapies. We observed increased levels of precursor exhausted T (Texp) cells in responsive tumors after treatment, characterized by low expression of coinhibitory molecules and high expression of GZMK. By contrast, nonresponsive tumors failed to accumulate Texp cells. Our data suggested that Texp cells were unlikely to be derived from the reinvigoration of terminally exhausted cells; instead, they were accumulated by (1) local expansion and (2) replenishment by peripheral T cells with both new and pre-existing clonotypes, a phenomenon we named clonal revival. Our study provides insights into mechanisms underlying PD-1-based therapies, implicating clonal revival and expansion of Texp cells as steps to improve NSCLC treatment.

Phase I study of CAR-T cells with PD-1 and TCR disruption in mesothelin-positive solid tumors.

Programmed cell death protein-1 (PD-1)-mediated immunosuppression has been proposed to contribute to the limited clinical efficacy of chimeric antigen receptor T (CAR-T) cells in solid tumors. We generated PD-1 and T cell receptor (TCR) deficient mesothelin-specific CAR-T (MPTK-CAR-T) cells using CRISPR-Cas9 technology and evaluated them in a dose-escalation study. A total of 15 patients received one or more infusions of MPTK-CAR-T cells without prior lymphodepletion. No dose-limiting toxicity or unexpected adverse events were observed in any of the 15 patients. The best overall response was stable disease (2/15 patients). Circulating MPTK-CAR-T cells peaked at days 7-14 and became undetectable beyond 1 month. TCR-positive CAR-T cells rather than TCR-negative CAR-T cells were predominantly detected in effusion or peripheral blood from three patients after infusion. We further confirmed the reduced persistence of TCR-deficient CAR-T cells in animal models. Our results establish the preliminary feasibility and safety of CRISPR-engineered CAR-T cells with PD-1 disruption and suggest that the natural TCR plays an important role in the persistence of CAR-T cells when treating solid tumors.

Case Report: Low-Dose Decitabine Plus Anti-PD-1 Inhibitor Camrelizumab for Previously Treated Advanced Metastatic Non-Small Cell Lung Cancer.

Background: Although the programmed death 1 (PD-1)/programmed death-ligand 1 (PD-L1) inhibitors have markedly changed the strategies of cancer treatment, most patients with advanced non-small cell lung cancer (NSCLC) do not respond to PD-1/PD-L1 monotherapy. Epigenetic drugs have been hypothesized to possess the potential to sensitize PD-1/PD-L1 inhibitors. Case Presentation: Three patients with advanced metastatic NSCLC failed to respond to first-line systemic therapy and had a low tumor mutation burden, low tumor neoantigen burden, low microsatellite instability, and HLA loss of heterozygosity according to their target lesion biopsies, all of which were considered unfavorable factors for PD-1/PD-L1 blockage. However, all three patients responded to low-dose decitabine, an epigenetic drug, in combination with camrelizumab (anti-PD-1 antibody), with only controllable adverse events, indicating that low-dose decitabine can sensitize PD-1/PD-L1 inhibitors. Summary: We report a novel therapy with low-dose decitabine plus camrelizumab for advanced NSCLC on the basis of successful treatment of three patients, emphasizing the potential of epigenetic drugs to regulate PD-1/PD-L1 inhibitors in advanced NSCLC.

Manganese is critical for antitumor immune responses via cGAS-STING and improves the efficacy of clinical immunotherapy.

CD8+ T cell-mediated cancer clearance is often suppressed by the interaction between inhibitory molecules like PD-1 and PD-L1, an interaction acts like brakes to prevent T cell overreaction under normal conditions but is exploited by tumor cells to escape the immune surveillance. Immune checkpoint inhibitors have revolutionized cancer therapeutics by removing such brakes. Unfortunately, only a minority of cancer patients respond to immunotherapies presumably due to inadequate immunity. Antitumor immunity depends on the activation of the cGAS-STING pathway, as STING-deficient mice fail to stimulate tumor-infiltrating dendritic cells (DCs) to activate CD8+ T cells. STING agonists also enhance natural killer (NK) cells to mediate the clearance of CD8+ T cell-resistant tumors. Therefore STING agonists have been intensively sought after. We previously discovered that manganese (Mn) is indispensable for the host defense against cytosolic dsDNA by activating cGAS-STING. Here we report that Mn is also essential in innate immune sensing of tumors and enhances adaptive immune responses against tumors. Mn-insufficient mice had significantly enhanced tumor growth and metastasis, with greatly reduced tumor-infiltrating CD8+ T cells. Mechanically, Mn2+ promoted DC and macrophage maturation and tumor-specific antigen presentation, augmented CD8+ T cell differentiation, activation and NK cell activation, and increased memory CD8+ T cells. Combining Mn2+ with immune checkpoint inhibition synergistically boosted antitumor efficacies and reduced the anti-PD-1 antibody dosage required in mice. Importantly, a completed phase 1 clinical trial with the combined regimen of Mn2+ and anti-PD-1 antibody showed promising efficacy, exhibiting type I IFN induction, manageable safety and revived responses to immunotherapy in most patients with advanced metastatic solid tumors. We propose that this combination strategy warrants further clinical translation.

Efficacy and biomarker analysis of nivolumab plus gemcitabine and cisplatin in patients with unresectable or metastatic biliary tract cancers: results from a phase II study.

BACKGROUND: The prognosis of patients with unresectable or metastatic biliary tract cancer (BTC) is unacceptably low. This study aimed to determine the efficacy, safety and predictive biomarkers of the immune checkpoint inhibitor nivolumab in combination with chemotherapy in advanced BTCs. METHODS: In this open-label, single-arm, phase II trial, a chemotherapy and immunotherapy combination consisting of gemcitabine 1000 mg/m2, cisplatin 75 mg/m2 and nivolumab 3 mg/kg was administered every 3 weeks for up to six cycles. Maintenance treatment with gemcitabine plus nivolumab was administered to patients achieving disease control following the combination therapy. The primary outcome was the objective response rate. Secondary outcomes included safety, disease control rate (DCR), progression-free survival (PFS) and overall survival (OS). The exploratory objective was to assess biomarkers for predicting clinical response and prognosis. RESULTS: Thirty-two patients with a median age of 60 (range 27-69) years were enrolled. As of September 31, 2019, the median follow-up was 12.8 (95% CI 10.8 to 14.8) months. Twenty-seven response-evaluable patients received a median of 4 (IQR, 3-6) cycles of combination therapy, of whom 15 (55.6%) patients achieved an objective response, including 5 (18.6%) with a complete response (CR), and the DCR was 92.6%. Of the six patients in cohort A who were resistant to gemcitabine-based or cisplatin-based chemotherapy, one achieved CR and one achieved partial response. Thirteen of 21 chemotherapy-naive patients (61.9%) in cohort B achieved an objective response. The median PFS of all patients in cohorts A+B was 6.1 months. The median OS was 8.5 months, with a 33.3% 12-month OS rate. The most frequent grade 3 or higher adverse events were thrombocytopenia (56%) and neutropenia (22%). Fitness might be a biomarker for predicting clinical response. On-therapy changes in serum soluble FasL, MCP-1 and interferon-γ were correlated with prognosis. CONCLUSIONS: Nivolumab in combination with gemcitabine and cisplatin offers promising efficacy and a manageable safety profile for patients with advanced BTCs. TRIAL REGISTRATION NUMBER: NCT03311789.

Anti-EGFR chimeric antigen receptor-modified T cells in metastatic pancreatic carcinoma: A phase I clinical trial.

The current clinical outcome for patients with metastatic pancreatic carcinoma (PC) remains poor. Epidermal growth factor receptor (EGFR) is detectable in PC, suggesting that EGFR is a rational target in PC. We conducted a phase I clinical trial to evaluate the safety and efficacy of autologous anti-EGFR chimeric antigen receptor-modified T (CAR T-EGFR) cells in patients with metastatic PC. The expression levels of EGFR on tumor cells detected by immunohistochemistry were required to be more than 50%. Sixteen patients were enrolled and received one to three cycles of the CAR T-EGFR cell infusion within 6 months (median dose of CAR T cells: 3.48 × 106/kg; range, 1.31 to 8.9 × 106/kg) after the conditioning regimen with 100 to 200 mg/m2 nab-paclitaxel and 15 to 35 mg/kg cyclophosphamide. Grade ≥3 adverse events included fever/fatigue, nausea/vomiting, mucosal/cutaneous toxicities, pleural effusion and pulmonary interstitial exudation and were reversible. Of 14 evaluable patients, four achieved partial response for 2-4 months, and eight had stable disease for 2-4 months. The median progression-free survival was 3 months (range, 4-months) from the first cycle of CAR T-EGFR cell treatment, and the median overall survival of all 14 evaluable patients was 4.9 months (range, 2.9-30 months). Decreased EGFR expression on tumor cells was observed in patients who achieved stable disease with shrinkage of metastatic lesions in the liver, and enrichment of central memory T cells in infused cells improved the clinical response. In conclusion, the treatment with CAR T-EGFR cells is safe and effective in patients with metastatic PC. This trial was registered at www.clinicaltrials.gov (identifier no: NCT01869166).

CD133-directed CAR T cells for advanced metastasis malignancies: A phase I trial.

Expressed by cancer stem cells of various epithelial cell origins, CD133 is an attractive therapeutic target for cancers. Autologous chimeric antigen receptor-modified T-cell directed CD133 (CART-133) was first tested in this trial. The anti-tumor specificity and the postulated toxicities of CART-133 were first assessed. Then, we conducted a phase I clinical study in which patients with advanced and CD133-positive tumors received CART-133 cell-infusion. We enrolled 23 patients (14 with hepatocellular carcinoma [HCC], 7 with pancreatic carcinomas, and 2 with colorectal carcinomas). The 8 initially enrolled patients with HCC were treated by a CART-133 cell dose escalation scheme (0.05-2 × 106/kg). The higher CAR-copy numbers and its reverse relationship with the count of CD133+ cells in peripheral blood led to the determination of an acceptable cell dose is 0.5-2 × 106/kg and reinfusion cycle in 23 patients. The primary toxicity is a decrease in hemoglobin/platelet (≤ grade 3) that is self-recovered within 1 week. Of 23 patients, three achieved partial remission, and 14 achieved stable disease. The 3-month disease control rate was 65.2%, and the median progression-free survival was 5 months. Repeated cell infusions seemed to provide a longer period of disease stability, especially in patients who achieved tumor reduction after the first cell-infusion. 21 out of 23 patients had not developed detectable de novo lesions during this term. Analysis of biopsied tissues by immunohistochemistry showed CD133+ cells were eliminated after CART-133 infusions. This trial showed the feasibility, controllable toxicities, and effective activity of CART-133 transfer for treating patients with CD133-postive and late-stage metastasis malignancies.

Phase Ib/II study of safety and efficacy of low-dose decitabine-primed chemoimmunotherapy in patients with drug-resistant relapsed/refractory alimentary tract cancer.

The pressing need for improved therapeutic outcomes provides a good rationale for identifying effective strategies for alimentary tract (AT) cancer treatment. The potential re-sensitivity property to chemo- and immunotherapy of low-dose decitabine has been evident both preclinically and in previous phase I trials. We conducted a phase Ib/II trial evaluating low-dose decitabine-primed chemoimmunotherapy in patients with drug-resistant relapsed/refractory (R/R) esophageal, gastric or colorectal cancers. Forty-five patients received either the 5-day decitabine treatment with subsequent readministration of the previously resistant chemotherapy (decitabine-primed chemotherapy, D-C cohort) or the aforementioned regimen followed by cytokine-induced killer cells therapy (D-C and cytokine-induced killer [CIK] cell treatment, D-C + CIK cohort) based on their treatment history. Grade 3 to 4 adverse events (AEs) were reported in 11 (24.4%) of 45 patients. All AEs were controllable, and no patient experienced a treatment-related death. The objective response rate (ORR) and disease control rate (DCR) were 24.44% and 82.22%, respectively, including two patients who achieved durable complete responses. Clinical response could be associated with treatment-free interval and initial surgical resection history. ORR and DCR reached 28% and 92%, respectively, in the D-C + CIK cohort. Consistently, the progression-free survival (PFS) of the D-C + CIK cohort compared favorably to the best PFS of the pre-resistant unprimed therapy (p = 0.0001). The toxicity and ORRs exhibited were non-significantly different between cancer types and treatment cohort. The safety and efficacy of decitabine-primed re-sensitization to chemoimmunotherapy is attractive and promising. These data warrant further large-scale evaluation of drug-resistant R/R AT cancer patients with advanced stage disease.

Targeting cancer stem cells by using chimeric antigen receptor-modified T cells: a potential and curable approach for cancer treatment.

Cancer stem cells (CSCs), a subpopulation of tumor cells, have self-renewal and multi-lineage differentiation abilities that play an important role in cancer initiation, maintenance, and metastasis. An accumulation of evidence indicates that CSCs can cause conventional therapy failure and cancer recurrence because of their treatment resistance and self-regeneration characteristics. Therefore, approaches that specifically and efficiently eliminate CSCs to achieve a durable clinical response are urgently needed. Currently, treatments with chimeric antigen receptor-modified T (CART) cells have shown successful clinical outcomes in patients with hematologic malignancies, and their safety and feasibility in solid tumors was confirmed. In this review, we will discuss in detail the possibility that CART cells inhibit CSCs by specifically targeting their cell surface markers, which will ultimately improve the clinical response for patients with various types of cancer. A number of viewpoints were summarized to promote the application of CSC-targeted CART cells in clinical cancer treatment. This review covers the key aspects of CSC-targeted CART cells against cancers in accordance with the premise of the model, from bench to bedside and back to bench.

Phase I study of chimeric antigen receptor modified T cells in treating HER2-positive advanced biliary tract cancers and pancreatic cancers.

This phase I clinical trial (NCT01935843) is to evaluate the safety, feasibility, and activity of chimeric antigen receptor-engineered T cell (CART) immunotherapy targeting human epidermal growth factor receptor 2 (HER2) in patients with advanced biliary tract cancers (BTCs) and pancreatic cancers (PCs). Eligible patients with HER2-positive (>50%) BTCs and PCs were enrolled in the trial. Well cultured CART-HER2 cells were infused following the conditioning treatment composed of nab-paclitaxel (100-200 mg/m2) and cyclophosphamide (15-35 mg/kg). CAR transgene copy number in the peripheral blood was serially measured to monitor the expansion and persistence of CART-HER2 cells in vivo. Eleven enrolled patients received 1 to 2-cycle CART-HER2 cell infusion (median CAR+ T cell 2.1 × 106/kg). The conditioning treatment resulted in mild-to-moderate fatigue, nausea/vomiting, myalgia/arthralgia, and lymphopenia. Except one grade-3 acute febrile syndrome and one abnormal elevation of transaminase (>9 ULN), adverse events related to the infusion of CART-HER2 cells were mild-to-moderate. Post-infusion toxicities included one case of reversible severe upper gastrointestinal hemorrhage which occurred in a patient with gastric antrum invaded by metastasis 11 days after the CART-HER2 cell infusion, and 2 cases of grade 1-2 delayed fever, accompanied by the release of C-reactive protein and interleukin-6. All patients were evaluable for assessment of clinical response, among which 1 obtained a 4.5-months partial response and 5 achieved stable disease. The median progression free survival was 4.8 months (range, 1.5-8.3 months). Finally, data from this study demonstrated the safety and feasibility of CART-HER2 immunotherapy, and showed encouraging signals of clinical activity.

Phase I Study of Chimeric Antigen Receptor-Modified T Cells in Patients with EGFR-Positive Advanced Biliary Tract Cancers.

Purpose: This study is an expanded and parallel clinical trial of EGFR-specific chimeric antigen receptor-engineered autologous T (CART) cell immunotherapy (NCT01869166) to assess the safety and activity of CART-EGFR cell therapy in EGFR-positive advanced unresectable, relapsed/metastatic biliary tract cancers (BTC).Experimental Design: Patients with EGFR-positive (>50%) advanced unresectable, relapsed/metastatic BTCs were enrolled. Well-produced CART-EGFR cells were infused in a manner of dose escalation after the conditioning treatment with nab-paclitaxel (100-250 mg/m2) and cyclophosphamide (15-35 mg/kg).Results: A total of 19 patients (14 cholangiocarcinomas and 5 gallbladder carcinomas) received one to three cycles of CART-EGFR cell infusion (median CART cell dose, 2.65 × 106/kg; range, 0.8-4.1 × 106/kg) within 6 months. The CART-EGFR cell infusion was tolerated, but 3 patients suffered grade ≥3 acute fever/chill. Grade 1/2 target-mediated toxicities including mucosal/cutaneous toxicities and acute pulmonary edema and grade ≥3 lymphopenia and thrombocytopenia related to the conditioning treatment were observed. Of 17 evaluable patients, 1 achieved complete response and 10 achieved stable disease. The median progression-free survival was 4 months (range, 2.5-22 months) from the first cycle of treatment. Analysis of data indicated that the enrichment of central memory T cells (Tcm) in the infused CART-EGFR cells improved the clinical outcome.Conclusions: The CART-EGFR cell immunotherapy was a safe and active strategy for EGFR-positive advanced BTCs. The enrichment of Tcm in the infused CART-EGFR cells could predict clinical response. Clin Cancer Res; 24(6); 1277-86. ©2017 AACRSee related commentary by Kalos, p. 1246.

Increased IFNγ+ T Cells Are Responsible for the Clinical Responses of Low-Dose DNA-Demethylating Agent Decitabine Antitumor Therapy.

Purpose: Low-dose DNA-demethylating agent decitabine therapy is effective in a subgroup of cancer patients. It remains largely elusive for the biomarker to predict therapeutic response and the underlying antitumor mechanisms, especially the impact on host antitumor immunity.Experimental Design: The influence of low-dose decitabine on T cells was detected both in vitro and in vivo Moreover, a test cohort and a validation cohort of advanced solid tumor patients with low-dose decitabine-based treatment were involved. The activation, proliferation, polarization, and cytolysis capacity of CD3+ T cells were analyzed by FACS and CCK8 assay. Kaplan-Meier and Cox proportional hazard regression analysis were performed to investigate the prognostic value of enhanced T-cell activity following decitabine epigenetic therapy.Results: Low-dose decitabine therapy enhanced the activation and proliferation of human IFNγ+ T cells, promoted Th1 polarization and activity of cytotoxic T cells both in vivo and in vitro, which in turn inhibited cancer progression and augmented the clinical effects of patients. In clinical trials, increased IFNγ+ T cells and increased T-cell cytotoxicity predicted improved therapeutic responses and survival in the test cohort and validation cohort.Conclusions: We find that low-dose decitabine therapy promotes antitumor T-cell responses by promoting T-cell proliferation and the increased IFNγ+ T cells may act as a potential prognostic biomarker for the response to decitabine-based antitumor therapy. Clin Cancer Res; 23(20); 6031-43. ©2017 AACR.

Cocktail treatment with EGFR-specific and CD133-specific chimeric antigen receptor-modified T cells in a patient with advanced cholangiocarcinoma.

BACKGROUND: Cholangiocarcinoma (CCA) is one of the most fatal malignant tumors with increasing incidence, mortality, and insensitivity to traditional chemo-radiotherapy and targeted therapy. Chimeric antigen receptor-modified T cell (CART) immunotherapy represents a novel strategy for the management of many malignancies. However, the potential of CART therapy in treating advanced unresectable/metastatic CCA is uncharted so far. CASE PRESENTATION: In this case, a 52-year-old female who was diagnosed as advanced unresectable/metastatic CCA and resistant to the following chemotherapy and radiotherapy was treated with CART cocktail immunotherapy, which was composed of successive infusions of CART cells targeting epidermal growth factor receptor (EGFR) and CD133, respectively. The patient finally achieved an 8.5-month partial response (PR) from the CART-EGFR therapy and a 4.5-month-lasting PR from the CART133 treatment. The CART-EGFR cells induced acute infusion-related toxicities such as mild chills, fever, fatigue, vomiting and muscle soreness, and a 9-day duration of delayed lower fever, accompanied by escalation of IL-6 and C reactive protein (CRP), acute increase of glutamic-pyruvic transaminase and glutamic-oxalacetic transaminase, and grade 2 lichen striatus-like skin pathological changes. The CART133 cells induced an intermittent upper abdominal dull pain, chills, fever, and rapidly deteriorative grade 3 systemic subcutaneous hemorrhages and congestive rashes together with serum cytokine release, which needed emergent medical intervention including intravenous methylprednisolone. CONCLUSIONS: This case suggests that CART cocktail immunotherapy may be feasible for the treatment of CCA as well as other solid malignancies; however, the toxicities, especially the epidermal/endothelial damages, require a further investigation. TRIAL REGISTRATION: ClinicalTrials.gov NCT01869166 and NCT02541370 .

Autologous T Cells Expressing CD30 Chimeric Antigen Receptors for Relapsed or Refractory Hodgkin Lymphoma: An Open-Label Phase I Trial.

Purpose: Relapsed or refractory Hodgkin lymphoma is a challenge for medical oncologists because of poor overall survival. We aimed to assess the feasibility, safety, and efficacy of CD30-targeting CAR T cells in patients with progressive relapsed or refractory Hodgkin lymphoma.Experimental Design: Patients with relapsed or refractory Hodgkin lymphoma received a conditioning chemotherapy followed by the CART-30 cell infusion. The level of CAR transgenes in peripheral blood and biopsied tumor tissues was measured periodically according to an assigned protocol by quantitative PCR (qPCR).Results: Eighteen patients were enrolled; most of whom had a heavy treatment history or multiple tumor lesions and received a mean of 1.56 × 107 CAR-positive T cell per kg (SD, 0.25; range, 1.1-2.1) in total during infusion. CART-30 cell infusion was tolerated, with grade ≥3 toxicities occurring only in two of 18 patients. Of 18 patients, seven achieved partial remission and six achieved stable disease. An inconsistent response of lymphoma was observed: lymph nodes presented a better response than extranodal lesions and the response of lung lesions seemed to be relatively poor. Lymphocyte recovery accompanied by an increase of circulating CAR T cells (peaking between 3 and 9 days after infusion) is a probable indictor of clinical response. Analysis of biopsied tissues by qPCR and immunohistochemistry revealed the trafficking of CAR T cells into the targeted sites and reduction of the expression of CD30 in tumors.Conclusions: CART-30 cell therapy was safe, feasible, and efficient in relapsed or refractory lymphoma and guarantees a large-scale patient recruitment. Clin Cancer Res; 23(5); 1156-66. ©2016 AACR.

Chimeric antigen receptor-modified T cells for the immunotherapy of patients with EGFR-expressing advanced relapsed/refractory non-small cell lung cancer.

The successes achieved by chimeric antigen receptor-modified T (CAR-T) cells in hematological malignancies raised the possibility of their use in non-small lung cancer (NSCLC). In this phase I clinical study (NCT01869166), patients with epidermal growth factor receptor (EGFR)-positive (>50% expression), relapsed/refractory NSCLC received escalating doses of EGFR-targeted CAR-T cell infusions. The EGFR-targeted CAR-T cells were generated from peripheral blood after a 10 to 13-day in vitro expansion. Serum cytokines in peripheral blood and copy numbers of CAR-EGFR transgene in peripheral blood and in tissue biopsy were monitored periodically. Clinical responses were evaluated with RECIST1.1 and immune- related response criteria, and adverse events were graded with CTCAE 4.0. The EGFR-targeted CAR-T cell infusions were well-tolerated without severe toxicity. Of 11 evaluable patients, two patients obtained partial response and five had stable disease for two to eight months. The median dose of transfused CAR(+) T cells was 0.97×10(7) cells kg(-1) (interquartile range (IQR), 0.45 to 1.09×10(7) cells kg(-1)). Pathological eradication of EGFR positive tumor cells after EGFR-targeted CAR-T cell treatment can be observed in tumor biopsies, along with the CAR-EGFR gene detected in tumor-infiltrating T cells in all four biopsied patients. The EGFR-targeted CAR-T cell therapy is safe and feasible for EGFR-positive advanced relapsed/refractory NSCLC.

An analytical biomarker for treatment of patients with recurrent B-ALL after remission induced by infusion of anti-CD19 chimeric antigen receptor T (CAR-T) cells.

Anti-CD19 chimeric antigen receptor-modified T (CAR-T-19) cells have emerged as a powerful targeted immunotherapy for B-cell lineage acute lymphoblastic leukemia with a remarkable clinical response in recent trials. Nonetheless, few data are available on the subsequent clinical monitoring and treatment of the patients, especially those with disease recurrence after CAR-T-19 cell infusion. Here, we analyzed three patients who survived after our phase I clinical trial and who were studied by means of biomarkers reflecting persistence of CAR-T-19 cells in vivo and predictive factors directing further treatment. One patient achieved 9-week sustained complete remission and subsequently received an allogeneic hematopoietic stem cell transplant. Another patient who showed relapse after 20 weeks without detectable leukemia in the cerebrospinal fluid after CAR-T-19 cell treatment was able to achieve a morphological remission under the influence of stand-alone low-dose chemotherapeutic agents. The third patient gradually developed extensive extramedullary involvement in tissues with scarce immune- cell infiltration during a long period of hematopoietic remission after CAR-T-19 cell therapy. Long-term and discontinuous increases in serum cytokines (mainly interleukin 6 and C-reactive protein) were identified in two patients (Nos. 1 and 6) even though only a low copy number of CAR molecules could be detected in their peripheral blood. This finding was suggestive of persistent functional activity of CAR-T-19 cells. Combined analyses of laboratory biomarkers with their clinical manifestations before and after salvage treatment showed that the persistent immunosurveillance mediated by CAR-T-19 cells would inevitably potentiate the leukemia-killing effectiveness of subsequent chemotherapy in patients who showed relapse after CAR-T-19-induced remission.

Treatment of CD20-directed Chimeric Antigen Receptor-modified T cells in patients with relapsed or refractory B-cell non-Hodgkin lymphoma: an early phase IIa trial report.

Patients with relapsed or refractory non-Hodgkin lymphoma have a dismal prognosis. Chimeric Antigen Receptor (CAR)-modified T cells (CART cells) that targeted CD20 were effective in a phase I clinical trial for patients with advanced B-cell lymphomas. We performed a phase IIa trial to further assess the safety and efficacy of administering autologous anti-CD20 CART (CART-20) cells to patients with refractory or relapsed CD20+ B-cell lymphoma. Eleven patients were enrolled, and seven patients underwent cytoreductive chemotherapy to debulk the tumors and deplete the lymphocytes before receiving T-cell infusions. The overall objective response rate was 9 of 11 (81.8%), with 6 complete remissions (CRs) and 3 partial remissions; no severe toxicity was observed. The median progression-free survival lasted for >6 months, and 1 patient had a 27-month continuous CR. A significant inverse correlation between the levels of the CAR gene and disease recurrence or progression was observed. Clinically, the lesions in special sites, specifically the spleen and testicle, were refractory to CART-20 treatment. Collectively, these results together with our data from phase I strongly demonstrated the feasibility and efficacy of CART-20 treatment in lymphomas and suggest large-scale patient recruitment in a future study. This study was registered at www.clinicaltrials.org as NCT01735604.

Tolerance and efficacy of autologous or donor-derived T cells expressing CD19 chimeric antigen receptors in adult B-ALL with extramedullary leukemia.

The engineering of T lymphocytes to express chimeric antigen receptors (CARs) aims to establish T cell-mediated tumor immunity rapidly. In this study, we conducted a pilot clinical trial of autologous or donor- derived T cells genetically modified to express a CAR targeting the B-cell antigen CD19 harboring 4-1BB and the CD3ζ moiety. All enrolled patients had relapsed or chemotherapy-refractory B-cell lineage acute lymphocytic leukemia (B-ALL). Of the nine patients, six had definite extramedullary involvement, and the rate of overall survival at 18 weeks was 56%. One of the two patients who received conditioning chemotherapy achieved a three-month durable complete response with partial regression of extramedullary lesions. Four of seven patients who did not receive conditioning chemotherapy achieved dramatic regression or a mixed response in the haematopoietic system and extramedullary tissues for two to nine months. Grade 2-3 graft-versus-host disease (GVHD) was observed in two patients who received substantial donor-derived anti-CD19 CART (chimeric antigen receptor-modified T) cells 3-4 weeks after cell infusions. These results show for the first time that donor-derived anti-CD19 CART cells can cause GVHD and regression of extramedullary B-ALL. This study is registered at www.clinicaltrials.gov as NCT01864889.