Phase I Clinical Trial Using Autologous Ex Vivo Expanded NK Cells and Cytotoxic T Lymphocytes for Cancer Treatment in Vietnam.

(1) Background: Immune cell therapy recently attracted enormous attention among scientists as a cancer treatment, but, so far, it has been poorly studied and applied in Vietnam. The aim of this study was to assess the safety of autologous immune cell therapy for treating lung, liver, and colon cancers-three prevalent cancers in Vietnam. (2) Method: This was an open-label, single-group clinical trial that included 10 patients with confirmed diagnosis of colon, liver, or lung cancer, conducted between March 2016 and December 2017. (3) Results: After 20-21 days of culture, the average number of cytotoxic T lymphocytes (CTLs) increased 488.5-fold and the average cell viability was 96.3%. The average number of natural killer cells (NKs) increased 542.5-fold, with an average viability of 95%. Most patients exhibited improved quality of life, with the majority of patients presenting a score of 1 to 2 in the Eastern Cooperative Oncology Group (ECOG) performance status (ECOG/PS) scale, a decrease in symptoms on fatigue scales, and an increase in the mean survival time to 18.7 months at the end of the study. (4) Conclusion: This method of immune cell expansion met the requirements for clinical applications in cancer treatment and demonstrated the safety of this therapy for the cancer patients in Vietnam.

Alloreactive cytotoxic T lymphocyte immunotherapy treatment of a patient with metastatic prostate cancer: A case report.

RATIONALE: Cytotoxic T lymphocyte (CTL) immunotherapy is an autologous cellular immune therapy that has been approved for treating patients with malignant tumors. However, there is still limited information regarding the impact of CTL on metastatic prostate cancer (PC) patients with bone metastatic lesions. PATIENT CONCERNS: An 82-year-old male patient complained of interrupted urination, urination pain, and significant dysuria on November 24, 2014. Transurethral resection of the prostate (TURP) and postoperative pathological examination showed prostatic adenocarcinoma, and a SPECT/CT scan demonstrated multiple bone metastases. In addition, prostate specific antigen (PSA) and free PSA (FPSA) levels were 54.54 µg/mL and 2.63 µg/mL, respectively, at the beginning of treatment. DIAGNOSES: The man was diagnosed with prostatic adenocarcinoma and multiple bone metastases. INTERVENTIONS: The patient received 30 cycles of alloreactive CTL (ACTL) immunotherapy regularly. OUTCOMES: Over the course of the 2-year treatment, the PC patient exhibited diminished bone metastasis accompanied by a marked reduction of serum PSA and FPSA from 54.54 and 2.63 µg/ml to 0.003 and <0.006 µg/ml, respectively. LESSONS: Our clinical observations demonstrate that CTL immunotherapy is a viable treatment option for PC patients, particularly those with bone metastatic lesions and high serum levels of PSA and FPSA.

Antigen-specific impairment of adoptive T-cell therapy against cancer: players, mechanisms, solutions and a hypothesis.

Adoptive cell therapy (ACT) destroys tumors with infused cytotoxic T lymphocytes (CTLs). Although successful in some settings, ACT is compromised due to impaired survival or functional inactivation of the CTL. To better understand the mechanisms involved, we have exploited a mouse model of leukemia expressing ovalbumin as a tumor neoantigen to address these questions: (i) Is CTL impairment during ACT antigen specific? (ii) If yes, which are the antigen-presenting cells responsible? (iii) Can this information assist the development of complementary therapies to improve ACT? Our results indicate that the target (tumor) cells, not cross-presenting cells, are the main culprits of antigen-specific CTL inactivation. We find that the affinity/avidity of the CTL-tumor cell interaction has little influence on ACT outcomes, while tumor density is a major determinant. Reduction of tumor burden with mild non-lymphoablative and non-inflammatory chemotherapy can dramatically improve the efficacy of ACT and may minimize side-effects. We propose a general mechanism for the inactivation of anti-self CTL in the same tissues where the activity of anti-foreign CTL is preserved, based on the density of target cells. This mechanism, which we tentatively call stunning, may have evolved to protect infected sites from self-destruction and is exploited by tumors to inactivate CTL.

Palmitate-derivatized human IL-2: a potential anticancer immunotherapeutic of low systemic toxicity.

PURPOSE AND EXPERIMENTAL DESIGN: Recombinant human IL-2 (rhIL-2) is a potent cytokine and FDA-approved anticancer drug. However, its clinical use has been limited by severe toxicity, associated primarily with systemic administration with excess protein distributing freely throughout the body. We hypothesized that rhIL-2 in alternate forms permitting more restricted localization may exert stronger antitumor efficacy and less toxicity. Here, we have tested the utility of palmitate-derivatized rhIL-2. rhIL-2 was reacted with N-hydroxysuccinimide palmitate ester. The resultant lipidated rhIL-2 (pIL-2), when mixed with cells, could spontaneously transfer from solution to cell surfaces. Next, anticancer efficacy of pIL-2 was assessed in two modalities. For adoptive T cell therapy, antitumor cytotoxic T cells (CTLs) were protein transferred ("painted") with pIL-2 and injected into mice bearing lymphoma. For in situ therapy, pIL-2 was injected intratumorally into mice bearing melanoma. Tumor growth and IL-2-associated toxicity were determined. RESULTS: In the lymphoma model, painting of the antitumor CTLs with pIL-2 markedly increased their viability and titer. In the melanoma model, intratumoral injection of pIL-2, but not rhIL-2, increased the number of activated CD8(+) T cells (IFN-γ(+)) in the spleen, reduced lung metastasis and prolonged the survival of treated mice. Moreover, while repeated intratumoral injection of rhIL-2 at an excessively high dose (10 injections of 10,000 IU/mouse) caused marked vascular leakage syndrome, the same regimen using pIL-2 caused no detectable toxicity. CONCLUSIONS: Transferring spontaneously from solution to cell surfaces, pIL-2 may bypass the current limitations of rhIL-2 and, thus, serve as a more effective and tolerable anticancer drug.

Generation of robust cytotoxic T lymphocytes against prostate specific antigen by transduction of dendritic cells using protein and recombinant adeno-associated virus.

Prostate cancer is the most common male cancer and there is an urgent need for adjuvant therapy such as immunotherapy. Recombinant adeno-associated virus type 2 (rAAV) vectors are useful for antigen gene-loading of human dendritic cells (DC) and for the rapid generation of cytotoxic T lymphocytes (CTL). In this study, we report a protocol for AAV-loading of DC with the AAV-loading of self-antigen prostate specific antigen (PSA) resulting in generation of CTL. PSA and cytokine expression, Cell surface marker analysis of DC and CTL cells were done using a FACScalibur flow cytometer. Chromium-51 release assay was used to analyze the killing activity of CTL. It was found that AAV-loading of DC with the PSA gene is superior to PSA protein loading of the same antigen for generating effective CTL. AAV/PSA-loading of DC was found to result in: (1) strong, rapid PSA-specific, MHC Class I-restricted CTL, (2) PSA expression in DC, (3) high CD80, CD83, and CD86 expression on DC, (4) high level of IL-12 and low level of IL-10 in DC, (5) T cell populations with significant interferon gamma (IFNgamma) expression, but low IL-4 expression, (6) high proliferation of T cell populations, (7) high CD8:CD4 and CD8:CD56 T cell ratios. The reason for generation of robust CTL is partly explained by the characteristics of DC and CTL described. This protocol may be useful for adoptive immunotherapy against self antigens such as PSA for prostate cancer.

Stimulation of host resistance against tumors by glycyrrhizin, an active component of licorice roots.

The effect of glycyrrhizin (GR), a Chinese herbal drug extracted from licorice roots, on the host resistance to tumors was investigated in a murine system. Administration of GR to BALB/c mice, which were inoculated s.c. with 1 x 10(6) cells/mouse of Meth A tumors, resulted in either no antitumor effect (8/20, 40%), a delay in tumor growth (9/20, 45%), or elimination of tumor growth (3/20, 15%) in these mice. In addition, the incidence of Meth A solid tumors was inhibited by GR when mice were inoculated with 1.5 x 10(4) cells/mouse or less of Meth A tumor cells, but not 7.5 x 10(4) cells/mouse or more. These results indicate that in this murine tumor system GR has a very weak antitumor effect. When GR at a dose of 20 mg/kg was administered to mice immunized with allogeneic lymphocytes 1 to 9 days after the immunization, the generation of suppressor macrophages (S-M phi) was clearly reduced as compared with that of S-M phi generated in immunized controls. In addition, when allospecific CTL (allo-CTL), which were generated in alloimmunized mice treated with GR followed by in vitro stimulation with allogeneic lymphocytes in a mixed lymphocyte reaction, were adoptively transferred to tumor-bearing mice treated with GR, the antitumor activity of allo-CTL derived from immunized mice treated with GR was markedly enhanced as compared with that of allo-CTL from immunized mice. Furthermore, established solid tumors were completely eliminated when interleukin-2 immunotherapy was performed in these mice in combination with GR treatments, but not interleukin-2 or GR alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Transfusion of autologous cytotoxic cells leads to failure of unrelated, DLA-nonidentical marrow grafts.

Dogs conditioned with 9.2 Gy total body irradiation (TBI) and given histoincompatible marrow transplants have a high rate of graft failure. Engraftment can be achieved by using 18 Gy fractionated TBI as preparative regimen. In this study, we tested the effects of infusing, at the time of histoincompatible marrow transplantation, autologous cells that had been stored before beginning high-dose (18 Gy) TBI. Our aim was to identify the peripheral blood mononuclear cells (PBMC) that contribute to failure of marrow grafts. Marrow graft failure was observed in three of three dogs receiving a mean of 2.1 x 10(8) unfractionated autologous PBMC/kg body weight as well as in two of two dogs receiving a mean dose of 0.075 x 10(8) PBMC/kg. When the dose of PBMC was decreased to 0.01 x 10(8)/kg, engraftment was seen in two of two dogs. These experiments thus established a cell dose response for causing marrow graft failure; further studies evaluated which subset of cells mediated this effect. Infusion of 0.09 x 10(8) nylon wool-nonadherent, plastic-nonadherent PBMC/kg was effective in causing marrow graft failure in three of three dogs. In contrast, infusion of 0.03 x 10(8) autologous monocytes/kg, enriched threefold above the number contained in the lower dose of PBMC causing graft failure, was associated with engraftment in four of six dogs. Infusion of 0.13 x 10(8) PBMC/kg treated with L-leucyl-L-leucine methyl ester (Leu-Leu-OMe), a drug that depletes canine cytotoxic T-lymphocytes (CTL), natural killer (NK) cells, and monocytes, permitted engraftment in three of four dogs. These data suggest that cytotoxic lymphocytes mediate failure of histoincompatible marrow grafts.

[The role of maternal virus-specific cytotoxic T-lymphocytes in the immunopathology of congenital influenza infection in mice]. / Rol' materinskikh virusspetsificheskikh tsitotoksicheskikh T-limfotsitov v immunopatologii pri vrozhdennoi grippoznoi infektsii u myshei.

Using 51Cr isotope label it was demonstrated that a very low per cent of syngeneic lymphocytes derived from healthy donors and inoculated in the blood stream of uninfected or influenza virus-infected pregnant mice is found in fetuses before delivery. Similar results were obtained after inoculation of virus-specific cytotoxic lymphocytes (CTL) into uninfected pregnant mice. After inoculation into the blood stream of infected pregnant mice of virus-specific CTL their migration into fetuses before delivery increases, being most marked in 25-30% of mice. Intravenous inoculation of excess CTL (10(6) cells) to infected pregnant mice resulted in rapid development of signs of slow influenza infection in the progeny with typical clinical picture and histopathological lesions in organs and tissues. Large doses (10(7)-10(8) cells) of CTL inoculated into the blood stream cause higher reduction and death of fetuses and increase the rate of stillbirths. The role of maternal virus-specific CTL in the pathogenesis of experimental congenital and especially slow influenza infection is discussed.