Infusion of in vitro expanded tumour-infiltrating lymphocytes and recombinant interleukin-2 in patients with surgically resected lymph node metastases of malignant melanoma: a pilot study.

A pilot study was set up in order to evaluate the feasibility and safety of infusing in vitro expanded tumour-infiltrating lymphocytes (TILs) and recombinant interleukin-2 (rIL-2) in a group of patients with advanced melanoma after radical resection of lymph node metastases. Twenty-four patients were eligible for the study and proliferating TILs were collected in 16. These patients were infused with TILs and then treated with rIL-2 and alpha-interferon. Short-term toxic effects (such as fever) were in general controlled by symptomatic drugs, whereas chronic toxicities were absent. The median follow-up period was 19 months; at present, 13 patients are alive and disease free, one patient is in progression and two patients have died. The approach was feasible and safe and the clinical results observed are comparable to those obtained by long-term treatment with other biological response modifiers.

Expression of HLA class I-specific inhibitory natural killer cell receptors in HIV-specific cytolytic T lymphocytes: impairment of specific cytolytic functions.

Human T lymphocytes have been shown to express inhibitory natural killer cell receptors (NKR), which can down-regulate T cell antigen receptor-mediated T cell function, including cytolytic activity. In the present study, we demonstrate that CD3+NKR+ cells can be identified in HIV-infected patients. HIV-specific cytolytic activity was analyzed in five patients in whom autologous lymphoblastoid B cell lines could be derived as a source of autologous target cells. Phytohemagglutinin-activated T cell populations that had been cultured in interleukin 2 displayed HIV-specific cytotoxic T lymphocyte (CTL) activity against HIV env, gag, pol, and nef in 3 of 5 patients. Addition of anti-NKR mAb of IgM isotype could increase the specific CTL activity. Moreover, in one additional patient, HIV-specific CTL activity was undetectable; however, after addition of anti-NKR mAb such CTL activity appeared de novo. Similar results were obtained by analysis of CD3+NKR+ clones derived from two patients. These data provide direct evidence that CD3+NKR+ cells may include antigen (HIV)-specific CTLs and that mAb-mediated masking of inhibitory NKR may revert the down-regulation of CTL function.

Treatment of stage IIIB non-small-cell lung cancer with surgery followed by infusion of tumor infiltrating lymphocytes and recombinant interleukin-2: a pilot study.

Stage IIIb non-small-cell lung cancer (NSCLC) has a poor prognosis. The median survival is approximately 6 months, and only 30% of patients are alive 1 year after diagnosis. The need for effective treatment is evident. The aim of this study was to evaluate whether the infusion of tumor-infiltrating lymphocytes (TILs), isolated from resected tumor, expanded in vitro and injected together with recombinant Interleukin-2, is feasible and may at least partially modify the poor prognosis in these patients. The infusion of TILs, derived from surgically resected NSCLC and expanded in vitro, together with subcutaneous (s.c.) injections of recombinant interleukin-2 (rIL-2) was attempted in a group of 11 patients. Treated patients were infused i.v. with in vitro expanded TILs (from 4 to 70 x 10(9) cells), and rIL-2 was injected s.c. at doses varying from 61 to 378 x 10(6) IU. Toxic side effects (fever and, in some cases, hypotension) were observed and limited the dose of rIL-2 infused. Follow-up was continued for 40 months. The mean survival time was 13.8 months. Three of five TIL-treated patients with residual disease have no evident disease after 1 year, and two of them are still alive and have no evidence of disease after 40 months. This pilot study suggests that the infusion of in vitro expanded TILs, derived from surgical samples, is feasible and seems to prolong overall survival and to control the residual disease in patients with advanced NSCLC.

Detection of oligoclonal T lymphocytes in lymph nodes draining from advanced non-small-cell lung cancer.

Despite the combined use of surgery and chemoradiotherapy, the poor prognosis of advanced non-small-cell lung cancer (NSCLC) requires the definition of new therapeutic approaches. The presence of T lymphocytes, with peculiar phenotypic, functional and molecular characteristics within the tumour, suggested the possible use of these cells, expanded in vitro, in protocols of adoptive immunotherapy. We have described how a population of oligoclonal T lymphocytes, derived from advanced NSCLC, can be expanded in vitro and has the capability of lysing autologous cancer cells. What is more important, we observed that patients with advanced NSCLC, treated with TIL expanded in vitro and recombinant interleukin-2, seemed to have a disease-free period longer than that of patients treated with conventional chemoradiotherapy. In an attempt to find new sources of specific lymphocytes for immunotherapy, we describe the analysis of the phenotypic, functional and molecular characteristics of T lymphocytes, derived from lymph nodes draining advanced NSCLC. In this paper we show that these cells, have restriction patterns of T cell receptor beta chain similar to those detectable in the population of infiltrating T lymphocytes. This finding suggests that T cells derived from draining lymph nodes of advanced NSCLC have peculiar characteristics and can be a suitable source of effector cells for protocols of adoptive immunotherapy in lung cancer treatment.

Proliferative, phenotypic and functional and molecular characteristics of tumour-infiltrating lymphocytes obtained from unselected patients with malignant melanomas and expanded in vitro in the presence of recombinant interleukin-2.

Tumour-infiltrating lymphocytes (TIL) derived from several histotypes, including melanoma, can be expanded in vitro in the presence of recombinant interleukin-2 (rIL-2) and infused as part of experimental adoptive immunotherapy protocols. Several authors have described the isolation and the expansion of TIL, but little is known about the actual proportion of unselected melanoma biopsies that give rise to 'positive' TIL cultures. We evaluated the proliferative, phenotypic, functional and molecular characteristics of cultured TIL obtained from 26 patients with metastatic melanomas, eligible for inclusion in a protocol of adoptive immunotherapy. Sixteen proliferating cultures were obtained. All TIL belonged to the T cell lineage and were characterized by a wide range of cytolytic activity against autologous and, to a lesser extent, allogeneic melanoma cells. Molecular analysis of the constant region of T cell receptor-beta (TCR-beta) showed an oligoclonal population of TIL in the majority of expanded samples, indicating that a selected subset of lymphocytes present in the tumour site could be expanded in vitro. These features, together with the high number of proliferating samples, make these populations of TIL suitable for infusion into patients with advanced disease.

Isolation and in vitro expansion of lymphocytes infiltrating non-small cell lung carcinoma: functional and molecular characterisation for their use in adoptive immunotherapy.

Tumour infiltrating lymphocytes (TIL) have the capability of recognising and lysing autologous cancer cells, both in vitro and in vivo. Advanced non-small cell lung carcinoma (NSCLC) is partially insensitive to chemo radiotherapy and has a poor prognosis: thus, for this, an immunotherapeutic approach could be attempted. We expanded in vitro 46 out of 70 samples of TIL derived from NSCLC. From proliferating TILS, a number varying from 10 to 50 x 10(9) cells was obtained. These lymphocytes belonged to the T cell lineage, had the capability of growing for 45-60 days and lysed autologous better than allogeneic cancer cells. In addition, analysis of the restriction maps of T cell receptor (TRC)-beta, demonstrated that an oligoclonal population of T cells was preselected in vivo, near the tumour site, and might be expanded in vivo, using phytohaemagglutin and interleukin 2 while maintaining the same characteristics of the original population. These results give a clear rationale for the use of in vitro expanded TIL from NSCLC in protocols of adoptive immunotherapy in patients with residual disease following surgery.

"HepG2/erythroid/brain" type glucose transporter (GLUT1) is highly expressed in human epidermis: keratinocyte differentiation affects GLUT1 levels in reconstituted epidermis.

In mature animals, the "HepG2/erythroid/brain" glucose transporter isoform (GLUT1) appears to be expressed at the highest levels at blood tissue barriers; however, these levels may still be lower than the levels of expression seen in fetal tissues. Also, glucose transporters might serve as water channels. Therefore, we decided to investigate GLUT1 expression in human epidermis, a very active tissue, in terms of metabolism, even if not directly vascularized. We found GLUT1 transcripts in human skin and demonstrated, by immunohistochemistry, that GLUT1 protein is highly expressed in the basal layer and, to a lower extent, in the immediately suprabasal layer of the epidermis. This distribution pattern suggested that GLUT1 expression is affected by keratinocyte differentiation. To investigate this possibility, we used human epidermis reconstituted in culture. Our culture system allows the reconstruction of a stratified squamous epithelium which has been successfully grafted onto patients presenting large skin defects. Human keratinocytes have been cultured under conditions which allow a modulation of cellular differentiation and stratification. We observed that (i) GLUT1 expression is 4-6-fold higher in "stem-like" basal cells than in large, differentiated keratinocytes; (ii) culture conditions causing cell differentiation reduce GLUT1 expression, while conditions which minimize either differentiation or stratification of keratinocytes enhance GLUT1 expression. Finally, we found that IGF-1 and insulin, probably acting through the IGF-1 receptor, increase GLUT1 expression and stimulate glucose transport activity in epidermis reconstituted in culture. In conclusion, our data demonstrate that GLUT1 is highly expressed in the basal layers of human epidermis and that its expression is modulated by keratinocyte differentiation.

High expression levels of the "erythroid/brain" type glucose transporter (GLUT1) in the basal cells of human eye conjunctiva and oral mucosa reconstituted in culture.

The expression of the "erythroid/brain" type glucose transporter (GLUT1) seems to be a feature of "barrier" tissues, at least in humans. Recently, we reported that GLUT1 is highly expressed in the basal layers of either "authentic" human epidermis or human epidermis reconstituted in culture and that its expression seems to be related to keratinocyte differentiation. In this paper we demonstrate that GLUT1 is selectively expressed in the basal layers of either eye conjunctiva epithelia or oral mucosa, reconstituted in culture starting from 1-2 mm2 bioptic specimens of normal human tissue. GLUT1 mRNA and protein levels are very high in conjunctiva and oral mucosa, 2-3 times higher than in epidermis reconstituted in culture. Taking into account its localization at the border of tissues not directly vascularized, but metabolically active, GLUT1 could play an important role in controlling the entry of glucose into these firmly guarded tissues.

[Somatostatin in the treatment of acute pancreatitis and prevention of postoperative acute pancreatitis. Personal experience]. / La somatostatina nel trattamento della pancreatite acuta e nella prevenzione della P.A.P.O.--esperienza personale.

Authors' personal experience is reported in the use of somatostatin for the treatment of acute pancreatitis and after bilio-pancreatic surgery. The drug has proved to be highly effective in reducing the pain and in controlling the biohumoral balance. The preliminary results appear to confirm the opinion that somatostatin is a major therapeutic tool in the treatment of bilio-pancreatic diseases.