rIL-2 Treatment Research Articles

Immunotargeting of T Follicular Helper (TFH) Cells for SLE Treatment

Abstract T follicular helper (TFH) cells promote antibody (Ab) responses by providing both survival and differentiation signals to B cells. As such, the expansion of self-reactive TFH cells contributes to auto-Ab production and correlates with disease severity in SLE. Importantly, blockade of TFH activity prevents auto-Ab production and disease progression in lupus-prone mice. Thus, TFH cells have been suggested as a target for treating SLE. Unfortunately, there are currently no therapies to selectively deplete TFH cells in vivo. We have previously identified IL-2 as a critical factor that suppresses TFH cell responses in vivo. In agreement with its negative role in TFH cell development, we show here that low-dose recombinant IL-2 (rIL-2) treatment prevents self-reactive TFH cell responses in lupus-prone mice. Importantly, this effect is independent of TREGS. Thus, our results demonstrate a novel immunosuppressive function of IL-2 that is independent of its role in regulatory T cell homeostasis. In addition, we demonstrate that rIL-2 can be targeted to TFH cells by adding an IL-6 blockade. Altogether, our data suggest that TFH cell suppression following low-dose IL-2 treatment is a significant mechanism underlying the immunosuppressive effects of low-dose rIL-2 immunotherapies.

Reduced Tregs in peripheral blood of PCOS patients - a consequence of aberrant Il2 signaling.

The immunesupressive action of CD4(+)CD25(+) CD127(-/low) T regulatory cells (Tregs) is vital for an efficient reproductive function. However no data exists on their number or functionality in polycystic ovary syndrome (PCOS). The study aimed to analyze the frequency of circulating Tregs and key factors modulating them in women with PCOS. This is a retrospective, case-control cohort study conducted in women with PCOS recruited from Samad IVF hospitals and Women and Children Hospital, Thiruvananthapuram, India. Women with PCOS (N = 20) were diagnosed according to Rotterdam Consensus and normal menstruating women were taken as controls (N = 2331). We analyzed the proportion of CD4(+)CD25(+) CD127(-/low) Tregs in women with PCOS by fluorescent activated cell sorting. The study discovered that the women with PCOS have reduced numbers of Tregs (2.626 ± 0.62) compared with controls (4.253 ± 0.87) (t = 6.963, P < .0001, mean difference = -1.627; 95% confidence interval = -2.099--1.155). We documented a decrease in the follicular phase Treg expansion in women with PCOS. Our results revealed a reduced STAT5A (fold change [FC] = 7.642, P < .0004)/STAT5B (FC = 3.824, P < .0001), FOXP3 (FC = 4.1343, P = .0004)/CTLA4 (FC = 2.569, P = .0001) and elevated AKT (FC = 7.39, P = .05)/PIK3 (FC = 5.326, P = .0002) expression in women with PCOS. Recombinant interleukin 2 (rIL2) treatment failed to improve FOXP3/CTLA4 levels but caused a reduction of AKT/PIK3 arm, possibly due to an elevated PTEN in women with PCOS. The study suggests that women with PCOS have reduced Tregs due to an inherent hyporesponsiveness to IL2, which is unable to activate STAT5B and reduce FOXP3 expression. IL2-based therapeutic strategies can ameliorate complications in PCOS by suppressing the AKT/PIK3 arm.

Recombinant interleukin‐2 in patients aged younger than 60 years with acute myeloid leukemia in first complete remission: Results from Cancer and Leukemia Group B 19808

Recombinant interleukin-2 (rIL-2) induces cellular cytotoxicity against leukemia blasts. Patients with acute myeloid leukemia (AML) in first complete remission (CR) may harbor minimal residual disease that is susceptible to rIL-2-activated effector cells. In the Cancer and Leukemia Group B (CALGB) 19808 study, patients with AML in first CR were randomly assigned after all planned chemotherapy to receive a 90-day course of subcutaneously administered rIL-2 or no further therapy. The primary objective was to compare disease-free survival (DFS) between the 2 treatment arms. A total of 534 patients achieved a CR, 214 of whom were randomized. Six courses of low-dose daily rIL-2 were given for the expansion of cytotoxic effector cells, each followed by 3-day high-dose boluses given to trigger cytotoxicity against minimal residual disease. On the protocol-specified intention-to-treat analysis, the hazards ratio for DFS was 0.75 (95% confidence interval, 0.52-1.09; P = .13); the 5-year DFS rate was 42% in the observation arm and 53% in the rIL-2 treatment arm. The hazards ratio for overall survival (OS) was 0.88 (95% confidence interval, 0.54-1.23; P = .34); the 5-year OS rate was 58% for the observation arm and 63% for the rIL-2 treatment arm. Twenty-five of the 107 patients randomized to treatment with rIL-2 either refused or were unable to initiate therapy and 30 patients did not complete their assigned therapy. However, significant toxicities were not commonly observed. The trial design did not anticipate the difficulties patients would encounter with protocol compliance. The efficacy of immunotherapy with rIL-2 administered after intensive postremission treatment was not assessed as planned because of unexpected refusals by patients and/or their physicians to comply with protocol-directed therapy. Neither DFS nor OS was found to be significantly improved.

Effect of combined treatment with recombinant interleukin-2 and allicin on pancreatic cancer

This study aimed to evaluate the efficacy of combined treatment with recombinant interleukin-2 (rIL-2) and allicin on pancreatic cancer and explore the potential immunological mechanism. A total of 60 C57/BL6 nude mice pancreatic cancer xenograft models were randomized into four groups of 15 mice per group: control group, allicin treatment group, rIL-2 treatment group, combined treatment with allicin and rIL-2 group. Mice in each group were treated with saline, rIL-2, allicin, or combination of rIL-2 and allicin by weekly i.v injection for four weeks. After four weeks of treatment, eyeballs of the mice were extracted and blood was drawn, percentages of CD4+T, CD8+T and NK cell were analyzed by FACS, IFN-γ level was detected by ELISA. One mouse in each group was sacrificed to measure the weight and volume of the tumor and prepared to the paraffin section of tumor tissue. Apoptosis of the tumor cells was analyzed by TUNEL and FACS. Other mice continued to receive treatment, survival period were compared between each group. We observed a significant suppression of xenograft growth and a significant prolonged survival time in the combined treatment with allicin and rIL-2 group (P<0.05). The most amount of apoptotic cells were observed in the combined therapy group (P<0.05). The percentages of CD4+T, CD8+T and NK cell and serum IFN-γ level increased significantly in the combined treatment group compared with other groups (P<0.05). Combined treatment with allicin and rIL-2 resulted in suppression of tumor growth and prolonged survival time possibly through activation of CD4+T, CD8+T and NK cell.

Recombinant interleukin-2が奏功した後腹膜血管肉腫の1例

Angiosarcoma is rare and highly malignant vascular neoplasm, and primary retroperitoneal angiosarcoma is extremely rare. Preoperative diagnosis is very difficult because there are no specific imaging features, and definitively effective treatment has not yet been established. We recently treated a patient with primary retroperitoneal angiosarcoma in which a prompt and exact diagnosis was difficult to obtain. One month after surgery, local recurrence appeared, but salvage immunotherapy using recombinant interleukin-2 (rIL-2) showed good efficacy, and the patient obtained complete response. Here we report this rare case of angiosarcoma. A 60-year-old woman with abdominal pain was diagnosed with a left retroperitoneal mass on CT scan. The tumor was about 9 cm in diameter and positioned above the left kidney. Further study using MRI, 131I-MIBG scintigraphy, and enhanced CT suggested chronic expanding hematoma and the patient underwent surgical resection. Histopathological diagnosis was primary retroperitoneal angiosarcoma based on positive staining for VIII factor, CD31, CD34, and p53. One month after surgery, FDG-PET revealed local recurrence adjacent to the psoas major. We initiated salvage immunotherapy using rIL-2. The patient was treated effectively and achieved complete response. She is alive and well 19 months after surgery and rIL-2 treatment.

Interleukin-2 treatment effect on imatinib pharmacokinetic, P-gp and BCRP expression in mice

The aim of this study was to investigate the effect that recombinant interleukin-2 (rIL-2) (0.16 MUI/injection) had on the pharmacokinetics of imatinib (IM) in plasma. In this study, IM was given orally to mice at a dose of 150 mg/kg once a day for 11 days (from day 1 to 11) either alone or in combination with intraperitoneal injections of rIL-2 twice a day from day 8 to 11. Pharmacokinetic parameters were determined using WinNonLin software. Areas under the curve were compared using Bailer's method. The repeated administration of the rIL-2+IM combination was shown to have two pharmacokinetic advantages compared with repeated IM doses alone. In addition to the pharmacodynamic interest of this treatment, we found that the combined treatment significantly increased the IM Cmax (P<0.05) and significantly increased the IM trough concentration (C(24 h)) (P<0.01), which was always above the minimum therapeutic IM concentration (1 mumol/l) in plasma. Those pharmacokinetic modifications may be explained, in part, by a decrease in the P-glycoprotein expression in the three intestinal segments of the mice (duodenum, P<0.01; jejunum, P<0.05; and ileum, P<0.05) and a decrease in BCRP expression in the duodenum segment (P<0.05) due to rIL-2. In another experiment, we found a significant induction of intestinal P-glycoprotein expression in mice that had been given IM orally (150 mg/kg) twice a day for 11 days. It would be interesting to further investigate the IM disposition associated with rIL-2 treatment for clinical applications.

Interleukin-2 immunotherapy action on innate immunity cells in peripheral blood and tumoral tissue of pancreatic adenocarcinoma patients

Innate immunity cells play a crucial role in host anticancer defense: cancer patients with high levels of natural killer (NK) cells and eosinophils have a better prognosis. Recombinant interleukin-2 (rIL-2) immunotherapy stimulates innate immunity cells. This study aims to evaluate the toxicity of pre- and postoperative rIL-2 treatment and the effects on innate immunity both in peripheral blood and in cancer tissue of patients with resectable pancreatic adenocarcinoma. Seventeen patients received high dose rIL-2 preoperative subcutaneous administration and two low dose postoperative cycles. We evaluated NK cell and eosinophil count in blood and in pancreatic surgical specimens. Toxicity was moderate. In the early postoperative period, blood NK cells and eosinophils significantly increased compared to basal values (p < 0.02). Histopathological analysis did not find significant intratumoral infiltration of NK cells nor of eosinophils. Preoperative high dose rIL-2 administration is able to counteract surgery-induced deficiency of NK cells and eosinophils in peripheral blood in the early postoperative period, although it cannot overcome local mechanisms of immune tumor escape in cancer tissue. The amplification of innate immunity, induced by immunotherapy, may improve the control of metastatic cells spreading in the perioperative period.

Different lymphoid cell populations produce varied levels of neopterin, beta 2-microglobulin and soluble IL-2 receptor when stimulated with IL-2, interferon-gamma or tumour necrosis factor-alpha.

Immune activation is central to many immune disorders. Clinical investigations have shown that immune activation can be quantified by measurements of soluble immune activation products in serum. Most in vitro studies of these immune activation products have focused on single products. In this study the specific cell sources and the major lymphokines inducing multiple activation products were investigated. In vitro addition of interferon-gamma (IFN-gamma) or IL-2 stimulated peripheral blood mononuclear cells to produce neopterin, beta 2-microglobulin (beta 2-M) and soluble IL-2 receptor (sIL-2R). These two lymphokines can act independently, because neutralizing antibodies to one of the lymphokines did not block the inducing activity of the other. Tumour necrosis factor-alpha (TNF-alpha) was also investigated and shown to be a less powerful inducer than IL-2 or INF-gamma. Separated lymphoid subpopulations responded differently to specific lymphokines. Monocytes produced only neopterin and only in response to INF-gamma. T cells released beta 2-M and sIL-2R in response to IL-2. B cells, however, were capable of producing all three immune activation products. Neopterin production in B cells was induced by either INF-gamma of IL-2, indicating that B cells have additional mechanisms for responding to lymphokines. To investigate whether these in vitro findings also occur in vivo, sera from patients who had received either rIL-2 or INF-gamma treatment were tested. INF-gamma administration led to substantial increases in serum neopterin but only a moderate beta 2-M increase and no increase in the serum sIL-2R levels. rIL-2 administration caused a substantial increase of all three serum immune activation products, consistent with our in vitro findings. The results confirm that increased serum levels of soluble immune activation products are indicators of increased cytokine production by lymphocytes and monocytes and also that B cells can be a prominent source of immune activation products.

Phase II study of ‘hybrid‘ biochemotherapy for advanced melanoma

18007 Background: The clinical activity of a hybrid biochemotherapy regimen was determined. This regimen incorporated important elements of previously studied sequential and concurrent biochemotherapy regimens in order to potentially increase dose intensity of the antitumor agents while decreasing morbidity, complexity of in-patient management, and length of hospital days. Methods: The antitumor agents and starting doses in the hybrid regimen were: cisplatin (25 mg/m2) IV on days 2–5, vinblastine (1.4–1.6 mg/m2) IV on days 1–5, dacarbazine (800 mg/m2) IV day 1 only, rIL-2 (12 MIU/m2) SC on days 6–10, and interferon-a2b (10 MU/m2) SC on days 2–6. Thirty-seven patients (pts) with a median age of 49 (range 19–63) yrs and Zubrod’s PS of 0–2 were treated. Patients with prior chemotherapy, rIL-2, active brain metastases were excluded. All cycles were 21 days or longer and the median number of courses delivered per patient was 4 (range 1–6). Results: Thirty-five pts were evaluated for response. There were 3/35 (9%) complete responses (CRs) and 17/35 (49%) partial responses (PRs). One CR and one PR were confirmed pathologically. The median overall survival was 12.6 months (95% CI: 9.9, 20.3). The median time to progression for all patients was 5.7 months (95% CI: 4.3, 9.8), while the TTP of responders was 7.2 months (95% CI: 5.1, 18.9). Brain metastases developed in 13 pts, -either while receiving treatment and responding systemically (5 pts) -or post-treatment with progressive systemic disease (8 pts). There were 2/12 (17%) early deaths in pts receiving vinblastine (1.6 mg/m2), one with typhlitis and the other with sepsis. Therefore, the vinblastine dose was reduced to 1.4 mg/m2 for patients #13–35. Neutropenia, with a median lowest neutrophil count of 0.08 (range 0–3) on day 13 (range day 6–16), and fever during rIL-2 treatment, increased the median days of hospitalization of all patients to 15 (range 12–23) for the first cycle. Conclusions: Hybrid biochemotherapy produced a high response rate and an overall survival similar to previously reported schedules. Length of hospital stay was significantly increased compared to the concurrent schedule. Development of brain metastases in responding pts resulted in early withdrawal from the program. Hybrid biochemotherapy with reduced doses of vinblastine was better tolerated. No significant financial relationships to disclose.

Cytomodulation of interleukin-2 effect by L-2-oxothiazolidine-4-carboxylate on human malignant melanoma.

Glutathione (GSH), the most prevalent intracellular non-protein thiol, plays an important role in the interleukin-2 (IL-2)-induced proliferative activity of normal and tumour cells expressing IL-2 receptor (IL-2R). In the present study, we investigate the effect of IL-2 on proliferation of the human melanoma A375 cell line, and the possible selective cytomodulation effect of this cytokine by L-2-oxothiazolidine-4-carboxylate (OTZ) on these melanoma cells and on human peripheral blood mononuclear cells (PBMCs). We found that recombinant IL-2 (rIL-2) significantly increased the proliferation rate of A375 melanoma cells, which was associated with an increase in GSH levels, the enhancement of IL-2Ralpha expression and the endogenous production of IL-2 in these tumour cells. In contrast, OTZ decreased GSH content and the proliferation rate of A375 cells, and abrogated the growth-promoting effects of rIL-2. Thus, compared to cells treated with rIL-2, pre-treatment with OTZ reduced IL-2Ralpha expression, and also decreased the consumption of rIL-2 and the endogenous secretion of IL-2 by these tumour cells. With regard to PBMCs, the combination of OTZ plus rIL-2 resulted in a more rapid and greater increase of IL-2Ralpha expression than rIL-2 alone, with the proliferation rate being similar in the first 24 h, but with a lower PBMC' count found thereafter compared to rIL-2 treatment alone. These results suggest that OTZ plays a crucial role in obtaining a selective cytomodulation of rIL-2, enabling it to exert its growth-promoting effect on normal cells, but not on melanoma cells, thereby possibly improving biochemotherapy with rIL-2.

Alternating temozolomide and low doses rIL-2 in patients with metastatic melanoma. An open non-randomized phase I/II study

7541 Background: Temozolomide (Temodal) is a novel chemotherapeutic agent that is rapidly absorbed, it offers 100% bioavailability as an oral preparation and achieves significant CNS penetration. It has been shown that combination rIL-2 in high doses and DTIC in melanoma gave higher response rate than the single agents alone. Temozolomide in sequence with low doses IL-2 might be an efficacious treatment where a cytostatic drug and a cytokine, which activates the immune system, may synergistically eliminate the tumour with low cumulative toxicity. Methods: The primary objective of this study was to determine the safety and tolerance of Temozolomide 200 mg/m2 orally once dayly from day 1 to 5, in sequential combination with subcutaneous injection of 4.5 × 106 IU rIL-2 (Proleukin™) at days 8–11, 15–18 and 22–25 in patients with measurable progressive metastatic malignant melanoma without radiological signs of CNS metastases. The secondary objectives were to determine tumour response and time to progression (TTP). Results: Forty-seven patients were recruited and 27 were included of which 4 were non-evaluable for response. Twenty-three patients tolerated the regimen with side effects below grade 3 according to WHO . Two patients suspended the treatment because of side effects above WHO grade 3 already during the first course with Temozolomide. Seven patients showed no tumour progression during the first four treatment cycles. These, consequently, continued to receive the combined treatment and to be monitored, until signs of relapse or max 21 courses. Two patients had CR, 3 PR and 2 SD. Six of twenty-five treated patients (24%) developed brain metastases during follow up. Conclusions: Temozolomide in combination with rIL-2 is a well tolerated regimen for out-patient treatment. Responders have long TTP, possibly due to rIL-2 treatment. No significant financial relationships to disclose.

GnRH Analogues, Transvaginal Ultrasound-Guided Drainage and Intracystic Injection of Recombinant Interleukin-2 in the Treatment of Endometriosis

We performed a double-blind, randomised controlled trial to evaluate the results of ultrasound-guided aspiration of endometriomas under the effect of GnRH analogues and a possible additional beneficial effect by leaving 600,000 IU of recombinant interleukin-2 (rIL-2) in the cysts. Twenty-four women with endometriosis-related symptoms, increased values of CA-125 and transvaginal ultrasonography showing endometriomas >3 cm who were initially sent to us for laparotomy and conservative surgery for endometriosis were included. Main outcome measures were severity of symptoms, size and percentage of echographical reduction of endometriomas and CA-125 levels after 2 menses post-GnRH analogues. Secondary outcome measures were the time until recurrence of abnormal parameters and the need for surgery after treatment. We found moderate clinical results after treatment with drainage plus GnRH analogues and significantly improved results in women having received rIL-2 intracystically. There were no side effects. Two out of 3 previously infertile patients became pregnant after therapy. Though the rates of recurrence of endometriomas ≧3 cm were similar in both groups, the period until recurrence was significantly greater when rIL-2 was used, and the rates of recurrence of symptoms and increased CA-125 values were also significantly lower in patients who received rIL-2. Surgery was finally performed on 10 patients (4 with and 6 without previous rIL-2 treatment) during follow-up (30 ± 12.7 months). These findings led to the conclusion that transvaginal ultrasound-guided puncture and aspiration of endometriomas under endometrial suppressive therapy with GnRH analogues have some value for endometriosis treatment, improving the clinical manifestations and avoiding some surgical therapies, and that rIL-2 left in the cyst increases these beneficial effects significantly.

Effect of interleukin-2 on intestinal P-glycoprotein expression and functionality in mice.

P-glycoprotein (Pgp), an active drug transporter expressed in enterocytes, can reduce intestinal absorption of drugs. Until now, interleukin-2 (IL2) has been reported as a Pgp modulator only in vitro. The present study examines the effects in vivo of IL2 after chronic treatment on intestinal Pgp protein expression and activity. This work also describes the effects of IL2 on the oral bioavailability of a Pgp substrate (digoxin) and of a Pgp/CYP3A cosubstrate (saquinavir). Human recombinant interleukin-2 (rIL2), administered to mice at 9 million international units/kg by intraperitoneal route twice daily for 4 days, led to a decrease in intestinal Pgp protein expression evaluated by Western blot with C219 antibody. In an in vitro everted gut sac model, rIL2 pretreatment decreased the Pgp-mediated transport of rhodamine 123 across mouse intestine by 37%. Moreover, rIL2 pretreatment markedly raised the area under the curve of orally administered digoxin from 3.5 +/- 0.5 to 9.7 +/- 1.5 mg min l(-1) as a consequence of the reduction in intestinal Pgp activity. rIL2 treatment increased saquinavir bioavailability from 2.5 to 4.5%, showing that first-pass metabolism is not affected and that Pgp by itself has only a moderate effect on saquinavir oral bioavailability. In conclusion, rIL2 pretreatment reduces intestinal Pgp protein expression and activity in mice. However, the effect of such a treatment on drug bioavailability depends on the extent of their metabolism by CYP3A.

Randomized, Open-Label Study of the Impact of Two Doses of Subcutaneous Recombinant Interleukin-2 on Viral Burden in Patients With HIV-1 Infection and CD4+ Cell Counts of ≥300/mm3: CPCRA 059

The effect of intermittent courses of recombinant interleukin-2 (rIL-2) on HIV-1 load in patients receiving combination antiretroviral therapy remains uncertain. CPCRA 059 was an open-label, randomized, multicenter trial in which 511 patients with HIV-1 infection and CD4+ cell counts of ≥300/mm3 who were receiving antiretroviral therapy were assigned to receive no rIL-2 (255 patients [controls]) or subcutaneous rIL-2 in dosages of 4.5 MIU (130) or 7.5 MIU (126) twice daily for 5-day courses every 8 weeks to maintain CD4+ cell counts that were twice the baseline value or ≥ 1,000/mm3. The primary objective of this study was to compare the effects of the two doses of rIL-2 and no rIL-2 on viral load and CD4+ cell counts over 12 months. There was no difference in the following viral load measurements between the rIL-2 treatment groups and the control treatment group: percentage of patients with viral loads of <50 copies/mL at 12 months (p = .55), time to viral load of ≥50 copies/mL for patients who had baseline viral loads of <50 copies/mL (p = .35), and change in viral load from baseline for patients who had viral loads of ≥50 copies/mL at baseline (p = .63). At each follow-up visit, the change in CD4+ cell count from baseline was significantly greater in the rIL-2 treatment groups than in the control treatment group, with a mean difference of 251/mm3 at month 12 (95% confidence interval, 207-295; p < .0001). No unanticipated adverse experiences were seen in this trial, to our knowledge the largest randomized evaluation of rIL-2 treatment conducted to date.

Influence of Low Dose rIL-2 Treatment on Endogenous Cytokine Production, Expression of Surface IL-2R and The Level of Soluble IL-2R in Patients With Minimal Residual Disease

This study was designed to investigate the immunomodulatory effect of low-dose IL-2 therapy (100 μ;g/day for 3 weeks) on interferon (IFN), tumor necrosis factor (TNF) production in vivo and in vitro and on the expression of IL-2Rα/β and soluble form of IL-2Rα. Patients enrolled in the study suffered from multiple myeloma (MM), Hodgkin's disease (HD) and non-Hodgkin's lymphoma (NHL) All of them were in remission after chemotherapy or radiotherapy. Our results indicated that IL-2 given subcutanously at a low dose of 100 μ;g/day for 3 weeks induced IFN-γ and TNF-α in plasma (measured 24 hrs after the last dose of IL-2) and affected the ability of blood leukocytes to produce cytokines. Production of IFN-γ induced in vitro with PHA was enhanced, but TNF-a production induced by lipopolysaccharide (LPS) and virus (Newcastle Disease Virus) was depressed. The expression of both: surface IL-2R. especially β subunit on total population of lymphocytes and NK cells, and soluble form of IL-2R, α chain were significantly cahanced after low-dose IL-2 therapy. Low dose IL-2 therapy was well tolerated by all patients, and side effects not exceeding II grade of toxicity according to WHO scale were observed. Five patients with MM relapsed 3-10 month after cessation of IL-2 therapy. but all patients with Hodgkin's and non-Hodgkin's lymphomas are still in remission (20 months of observation).

Effects of interleukin-2 for the treatment of malignant mesothelioma.

The hematological profile, and clinical parameters in mesothelioma affected patients undergoing a clinical trial to evaluate the efficacy of IL-2 treatment were investigated. Six patients were monitored for 6 weeks following therapy. Blood cell count, morphological and immunophenotypical analysis were performed, as well as clinical evaluations of the patients before and after therapy. Activation of the immune system (increase in lymphocytes, monocytes, eosinophils and HMS lymphocytes) induced by IL-2 was observed. The treatment was well tolerated: our patients had only mild adverse reactions controlled by symptomatic therapy. Eosinophilia represented the most evident negative effect. A slight decrease in CD4-positive subset of lymphocytes was observed after rIL-2 treatment. The therapy did not induce significant changes in the progression of the disease. In one patient necrosis at the tumoral site was observed after loco-regional rIL-2 administration.

Successful Adoptive Cellular Immunotherapy Is Dependent on Induction of a Host Immune Response Triggered by Cytokine (IFN-γ and Granulocyte/Macrophage Colony-Stimulating Factor) Producing Donor Tumor-Infiltrating Lymphocytes

Abstract Adoptive immunotherapy with tumor-infiltrating lymphocytes (TIL) and systemic low dose rIL-2 effectively eradicates pulmonary metastases of the murine MCA-105 sarcoma. We described earlier that host CD8+ T cells are critical for tumor eradication and that successful treatment is associated with production of high levels of IFN-γ and granulocyte/macrophage (GM)-CSF by donor TIL in vitro. Here, we propose the mechanism through which adoptively transferred Thy-1.1+ TIL induce a host antitumor response in congenic Thy-1.2+ tumor-bearing mice. Donor Thy-1.1+ TIL were detected at the tumor site 12 h after transfer. These Thy-1.1+ cells produced IFN-γ and GM-CSF in situ. The percentage of Thy-1.1+ TIL at the tumor site increased up to 16.4 ± 4.9% 24 h after transfer but decreased to undetectable levels thereafter. In contrast, the percentages of host cells producing IFN-γ and GM-CSF continued to increase at the tumor site. These increases were significantly higher in TIL + rIL-2-treated mice compared with untreated mice and rIL-2-treated mice 48 h after TIL transfer. The appearance of IFN-γ+ and GM-CSF+ cells was followed by a large influx of host CD4+, CD8+, and Thy-1.2+ TIL and eventually by tumor eradication. This response was tumor specific since TIL obtained from MCA-205 did not induce high levels of IFN-γ and GM-CSF and did not induce tumor eradication of MCA-105 tumor. Coinjection of Thy-1.1+ TIL and anti-IFN-γ or anti-GM-CSF mAb significantly inhibited antitumor efficacy of the TIL + rIL-2 treatment. We conclude that successful adoptive immunotherapy in this model is mediated through cytokine production by adoptively transferred TIL that induce a host T cell-dependent antitumor response.

Ultrastructural changes of neutrophils following IL-2 treatment in vivo.

An ultrastructural analysis of peripheral blood cells of cancer patients following recombinant interleukin-2 (rIL-2) treatment at high doses (rIL-2, 18 x 10(6) I.U./m2/day) has been performed. Characteristic patterns of intense phagocytosis (neutrophils with large vaculoes containing electron-dense bodies) were observed in granulocytes of the treated patients with respect to the basal situation. These characteristics were particularly evident in the advanced stages of the therapy (24 and 48 h). The state of neutrophil activation was sometimes accompanied by the appearance of a close net of cell membranes, probably derived from lamellopodia.

Effect of rIL-2 treatment on anti-tetanus toxoid response in the elderly

To explore the effects of interleukin-2 (IL-2) treatment in a vaccination protocol in the elderly, we administered low-dose rIL-2 to a group of aged subjects before primary tetanus toxoid immunization. A specific antibody response was detectable in the serum of 6/8 treated individuals after primary immunization, but in only 2/6 untreated controls; following antigenic boosting, specific antibody levels remained relatively unchanged in all the seroconverters. The data were confirmed by studying the ability to produce tetanus-specific antibodies in vitro, and by isoelectrofocusing analysis of serum anti-tetanus antibodies; this latter study showed a more restricted clonal response to the immunogen in untreated individuals. On the other hand, the study of the in vitro proliferative response to tetanus toxoid did not evidence clear differences between the two groups. On the whole, these data seem to indicate that a short-term rIL-2 treatment is able to potentiate the antibody response to tetanus toxoid, and may be a useful tool to improve humoral responses to vaccines in aged subjects.

Anti-tumor effects of local irradiation in combination with peritumoral administration of low doses of recombinant interleukin-2 (rIL-2).

The aim of this work was to improve radiotherapy results by immune stimulation. We tested the effects of a combination of radio- and immunotherapy, i.e., local low dose recombinant interleukin-2 (rIL-2) treatment, in two murine tumor models. Syngeneic tumors (SL2 lymphoma or M8013 mammary carcinoma) were induced subcutaneously on one or both flanks of mice. Irradiation was given either as a single dose (20 Gy) or fractionated (25 Gy) in 2 weeks. One or two cycles of rIL-2 were given concurrent with or subsequent to radiotherapy. One cycle of rIL-2 consisted of peritumoral injections administered on 5 consecutive days. Treatment effects were measured in terms of local tumor response and disease-free survival (DFS). The combined treatment modality was significantly better than treatment with either irradiation alone or rIL-2 alone. When tumors were inoculated on both flanks of the mice, combined radioimmunotherapy of one of the tumors also resulted in regression of the contralateral untreated tumor, indicating that a systemic anti-tumor immune reaction was induced. Additional rIL-2 injections did not enhance radiation toxicity. In conclusion supplementing irradiation with locally administered low doses of rIL-2 results in better local anti-tumor responses and DFS rates than either treatment alone without enhanced treatment toxicity. Furthermore, the local treatment induces a systemic anti-tumor reaction, influencing the growth patterns of a second, untreated tumor.