Low-dose Subcutaneous Interleukin-2 Research Articles

A phase Ib study of endogenous T cell therapy using SLC45A2-specific CD8 T cells for patients with metastatic uveal melanoma.

9588 Background: Overall survival (OS) for patients (pts) with advanced uveal melanoma (UM) is poor compared with outcomes in cutaneous melanoma. Roughly half of all pts with UM will develop distant metastatic disease despite effective treatment of the primary tumor. Antigen-specific endogenous T cells (ETC) can be expanded and infused successfully in pts with many different types of cancer. Our group has identified epitopes of SLC45A2, a melanosomal transport protein, which are highly expressed in UM and are present at low levels in normal melanocytes. In vitro, we showed that cytotoxic T cells against SLC45A2 were able to kill HLA-matched UM cell lines. In vivo, we hypothesize that infused ETC can traffic to tumor sites. Methods: Between 6/2017 and 12/2022, we conducted a single-center, IRB-approved, first-in-human phase 1b dose escalation study of ETC targeting SLC45A2 in pts with metastatic UM (NCT03068624). Eligible pts with metastatic UM who express HLA-A*02:01 or A*24:02 underwent apheresis to collect peripheral T cells which were then selected and expanded (Turnstile 1). For Turnstile 2, conditioning with low-dose cyclophosphamide (300 mg/m2) occurred on Day -2. For pts in the radiation cohort, radiation occurred between Day -7 and Day -1. Infusion of ETC was done via hepatic arterial or central venous catheter on Day 0 followed by low dose subcutaneous interleukin-2 (IL-2) twice daily for 14 days +/- ipilimumab. The study utilized a 3+3 design with a starting dose level of 3.3 x 109 cells/m2 of ETC alone. The primary objective was to evaluate the safety and tolerability of infusing ETC targeting SLC45A2. Secondary objectives were to evaluate the in vivo persistence and anti-tumor efficacy of this regimen. Results: With a median age of 58 years (30-77), 34 pts were enrolled, 16 (47%) men and 18 (53%) women. 33 (97%) pts underwent apheresis and 11 (32%) received infusion of ETC. One dose limiting toxicity (DLT) thought to be unrelated to treatment or underlying cancer (cardiac disorder Grade 5) occurred at the starting dose level with no additional DLTs. The maximum tolerated dose was established as 1 x 1010 cells/m2. The most common treatment-related grade 3 and 4 toxicities were lymphocyte count decrease 9 (82%), hyponatremia 1 (9%), and hypophosphatemia 1 (9%). Per immune-related response criteria (irRC), 4 (36%) pts had stable disease (SD), 6 (55%) had progression of disease (PD), and 1 (9%) was not evaluable for best response. Median overall survival (OS) was 8.9 weeks (wks). OS was 91% at 4 wks, 55% at 8 wks and 46% at 13 wks. Median progression-free survival was 5.9 wks. In the 4 patients with stable disease, the median duration of SD was 5.7 months. Conclusions: ETC targeting SLC45A2 is safe and well-tolerated in pts with metastatic UM. Further analyses are underway to evaluate T cell trafficking and persistence. Clinical trial information: NCT03068624 .

Real-world experience with low-dose IL-2 for children and young adults with refractory chronic graft-versus-host disease

AbstractThe majority of patients with chronic graft-versus-host disease (cGVHD) are steroid refractory (SR), creating a need for safe and effective therapies. Subcutaneous low-dose interleukin-2 (LD IL-2), which preferentially expands CD4+ regulatory T cells (Tregs), has been evaluated in 5 clinical trials at our center with partial responses (PR) in ∼50% of adults and 82% of children by week 8. We now report additional real-world experience with LD IL-2 in 15 children and young adults. We conducted a retrospective chart review of patients with SR-cGVHD at our center who received LD IL-2 from August 2016 to July 2022 not on a research trial. The median age at start of LD IL-2 was 10.4 years (range, 1.2-23.2 years) at a median of 234 days from cGVHD diagnosis (range, 11-542 days). Patients had a median of 2.5 (range, 1-3) active organs at LD IL-2 start and received a median of 3 (range, 1-5) prior therapies. The median duration of LD IL-2 therapy was 462 days (range, 8-1489 days). Most patients received 1 × 106 IU/m2 per day. There were no serious adverse effects. The overall response rate in 13 patients who received >4 weeks of therapy was 85% (complete response, n = 5; PR, n = 6) with responses in diverse organs. Most patients significantly weaned corticosteroids. Tregs preferentially expanded with a median peak fold increase of 2.8 in the ratio of Tregs to CD4+ conventional T cells (range, 2.0-19.8) by 8 weeks on therapy. LD IL-2 is a well-tolerated, steroid-sparing agent with a high response rate in children and young adults with SR-cGVHD.

A phase Ib study of endogenous SLC45A2-specific cytotoxic T cells for the treatment of patients with metastatic uveal melanoma.

TPS10086 Background: Overall survival (OS) for patients (pts) with advanced uveal melanoma (UM) is poor with a median survival of approximately 12 months. Roughly half of all pts with UM will develop distant metastatic disease despite effective treatment of the primary site. Metastatic UM has a 90% prevalence of liver involvement. Currently, there are no specific FDA-approved treatments for metastatic UM and consensus guidelines recommend participation in a clinical trial. Modern treatments such as checkpoint inhibitors and targeted therapy have less impressive outcomes in UM. Our group has identified peptide epitopes of SLC45A2, a melanosomal transport protein, that is highly expressed in UM and present at very low levels in normal melanocytes. We demonstrated that cytotoxic T cells against SLC45A2 were able to kill HLA-matched UM cell lines. Through the use of enabling technologies, SLC45A2-specific cytotoxic T cells can be isolated and expanded ex-vivo from peripheral blood for use in endogenous T cell (ETC) therapy (a form of adoptive cellular therapy). These activated ETCs can then be infused to traffic to tumor sites. CTLA4 is a T-cell surface protein that binds to B7 with a higher affinity than the costimulatory receptor CD28, providing an inhibitory signal to T-cells. Anti-CTLA4 blockade can divert this inhibition and release the brake on antigen-specific T-cell activation of ETC. We hypothesize that antigen-specific ETCs infused via hepatic artery will be safe and tolerable for UM pts with liver metastasis. Methods: We are conducting a first-in-human clinical trial (NCT03068624 ) of ETC therapy targeting SLC45A2 in combination with anti-CTLA4 in pts with metastatic UM. Pts who express HLA-A*02:01 or A*24:02 undergo apheresis to collect T cells. Their cells then undergo ex vivo cloning and interleukin-21 primed expansion. Hepatic arterial infusion of ETCs will ensure direct localization to the target organ of interest. Conditioning with low-dose cyclophosphamide (300 mg/m2) occurs on Day -2. Hepatic arterial infusion of ETCs on Day 0 is followed by low dose subcutaneous interleukin-2 (IL-2) twice daily for 14 days. The initial dose escalation phase is a 3+3 design with a starting dose level of 3.3 x 109 cells/m2 of ETCs alone. Once the maximum tolerated dose of ETCs is established, the dose expansion phase will include ETCs in combination with anti-CTLA4 (Ipilimumab). The primary objective is to evaluate the safety of this first in human T cell regimen. Secondary objectives are to evaluate the in vivo persistence and anti-tumor efficacy. Clinical trial information: NCT03068624.

The numbers of peripheral regulatory T cells are reduced in patients with psoriatic arthritis and are restored by low-dose interleukin-2.

Background:Although regulatory T cells (Tregs) play crucial roles in the maintenance of immune hemostasis, the numbers of peripheral Tregs in patients with psoriatic arthritis (PsA) remain unclear. We measured these numbers and the efficacy and safety of low-dose interleukin-2 (IL-2) therapy.Methods:We recruited 95 PsA patients, of whom 22 received subcutaneous low-dose IL-2 [0.5 million international units (MIU) per day for 5 days] combined with conventional therapies. The absolute numbers of cells in peripheral CD4+ T cell subsets were measured via modified flow cytometry. Clinical and laboratory indicators were compared before and after treatment.Results:PsA patients had lower peripheral Treg numbers than healthy controls (p < 0.01), correlating significantly and negatively with the levels of disease indicators (p < 0.05). Although low-dose IL-2 significantly increased the Th17 and Treg numbers in PsA patients compared with the baseline values, the Treg numbers rose much more rapidly than those of Th17 cells, re-balancing the Th17 and Treg proportions. Low-dose IL-2 combination therapy rapidly reduced PsA disease activities as indicated by the DAS28 instrument, thus the number of tender joints, visual analog scale pain, physician global assessment, the dermatology life quality index score, and the health assessment questionnaire score (all p < 0.05).Conclusion:PsA patients exhibited low Treg numbers. Low-dose IL-2 combination treatment increased these numbers and relieved disease activity without any apparent side effects. Additional studies are required to explore the long-term immunoregulatory utility of IL-2 treatment.

Safety of low-dose subcutaneous recombinant interleukin-2: systematic review and meta-analysis of randomized controlled trials

Standard-dose intravenous recombinant interleukin-2 (rIL-2) is indicated for the treatment of some subtypes of cancer; however, severe adverse events, including venous thromboembolism (VTE), may complicate its administration. Low-dose subcutaneous rIL-2 is being studied for the management of immune-mediated diseases, since it can modulate the immunological response by specifically targeting T regulatory (Treg) cells; importantly, it is supposed to cause fewer or no complications. In this systematic review and meta-analysis of phase II-III randomized controlled trials (RCTs), we investigated the safety of low-dose (<6 Million International Unit [MIU]/day) and ultra-low-dose (≤1 MIU/day) rIL-2 for severe adverse events (grade III-V) with a focus on VTE. Data of 1,321 patients from 24 RCTs were analysed: 661 patients were randomized to the rIL-2 arm (on top of standard of care) and 660 patients to standard of care alone or placebo. Two studies reported higher rates of thrombocytopenia in the low-dose rIL-2 arm. Ultra-low-dose rIL-2 was reported to be well tolerated in 6 studies with a negligible rate of severe adverse events. Symptomatic VTE events were not reported in any of the study arms (absolute risk difference 0% [95%CI −0.1%; +0.1%]). Our results may facilitate the study and introduction in clinical practice of low-dose rIL-2 for potentially new indications.

Phase II clinical trial of adoptive cell therapy for patients with metastatic melanoma with autologous tumor-infiltrating lymphocytes and low-dose interleukin-2

Adoptive cell therapy using autologous tumor-infiltrating lymphocytes (TIL) has shown significant clinical benefit, but is limited by toxicities due to a requirement for post-infusion interleukin-2 (IL-2), for which high dose is standard. To assess a modified TIL protocol using lower dose IL-2, we performed a single institution phase II protocol in unresectable, metastatic melanoma. The primary endpoint was response rate. Secondary endpoints were safety and assessment of immune correlates following TIL infusion. Twelve metastatic melanoma patients were treated with non-myeloablative lymphodepleting chemotherapy, TIL, and low-dose subcutaneous IL-2 (125,000 IU/kg/day, maximum 9–10 doses over 2 weeks). All but one patient had previously progressed after treatment with immune checkpoint inhibitors. No unexpected adverse events were observed, and patients received an average of 6.8 doses of IL-2. By RECIST v1.1, two patients experienced a partial response, one patient had an unconfirmed partial response, and six had stable disease. Biomarker assessment confirmed an increase in IL-15 levels following lymphodepleting chemotherapy as expected and a lack of peripheral regulatory T-cell expansion following protocol treatment. Interrogation of the TIL infusion product and monitoring of the peripheral blood following infusion suggested engraftment of TIL. In one responding patient, a population of T cells expressing a T-cell receptor Vβ chain that was dominant in the infusion product was present at a high percentage in peripheral blood more than 2 years after TIL infusion. This study shows that this protocol of low-dose IL-2 following adoptive cell transfer of TIL is feasible and clinically active. (ClinicalTrials.gov identifier NCT01883323.)

Treatment of Advanced Cancer-related Lymphocytopenia: Comparison among the Effects of Subcutaneous Low-dose Interleukin-2, High-dose Pineal Hormone Melatonin and Checkpoint Inhibitors

Treatment of Advanced Cancer-related Lymphocytopenia: Comparison among the Effects of Subcutaneous Low-dose Interleukin-2, High-dose Pineal Hormone Melatonin and Checkpoint Inhibitors

Long-Term Homeostatic Effects of Daily Low-Dose IL-2 on CD4+ FoxP3+ Regulatory T Cells in Patients with Active Chronic Graft-Versus-Host Disease

Patients with active chronic graft-versus-host disease (cGVHD) have poor reconstitution of CD4+ CD25+ FOXP3+ regulatory T cells (Tregs), which have broad suppressive activity and are required for the maintenance of peripheral tolerance after allogeneic hematopoietic stem cell transplant. Interleukin-2 (IL-2) is a key growth factor for the development, expansion and function of Tregs in vivo. Phase 1 (DFCI 07-083) and phase 2 (DFCI 11-149) studies of daily subcutaneous low-dose IL-2 in patients with refractory cGVHD demonstrated preferential Treg expansion in all patients and objective clinical responses in approximately 50% of participants. In the phase 2 study, 23 participants with clinical benefit (PR or SD with minor response) continued on extended IL-2 therapy, with 6 and 8 patients receiving over 1 and 2 years of low-dose IL-2, respectively. Analysis of phase 1 study patients indicated that IL-2 restored Treg homeostasis through rapid induction of Treg proliferation, increase in thymic Treg neogenesis and increased Treg expression of the anti-apoptotic Bcl-2 protein. Here, we provide novel insights into the immune homeostatic impact of low-dose IL-2 in phase 2 study patients during the initial 12 week treatment period and during extended therapy.Proliferation within the Treg compartment, measured by Ki67 expression, rapidly increased and peaked within 1 week of IL-2 initiation. Memory Tregs contained a higher fraction of proliferating cells and correspondingly, a rise in central memory (CM) and effector memory (EM) Tregs preceded an increase in naïve Tregs during the initial 12 week treatment period (Figure 1A and 1B). This is consistent with the observation that memory Tregs are more responsive than naive Tregs to activation by IL-2 in vitro. Although CM and EM Tregs continue to represent the predominant subsets of Tregs, there is a brisk expansion of the naïve Treg subset that coincides with increased thymic output of Tregs during extended IL-2 therapy.Consistent with their greater proliferative potential, CM and EM Tregs expressed lower levels of Bcl-2 compared with naïve Tregs. However, IL-2 promoted higher Bcl-2 expression in all Treg subsets to similar magnitudes during both the initial 12 week treatment period and extended therapy (Figure 2A). Bcl-2 was preferentially increased in Tregs and not in conventional CD4 (Tcon) or CD8 T cells (Figure 2B). As expected, naïve Tregs expressed very low levels of the pro-apoptotic CD95/Fas receptor protein. CD95 expression in CM and EM Tregs peaked during the period of rapid proliferation within the first week of IL-2 therapy and declined as Bcl-2 expression increased. Changes in CD95 expression levels were similar across all T cell populations. Thus, IL-2 leads to preferential expansion and survival of Tregs in cGVHD patients throughout the duration of therapy.Once IL-2 was discontinued following the initial 12 week treatment period, Treg numbers decreased to pre-treatment baseline levels within 4 weeks, indicating that continuous IL-2 exposure is required for maintenance of enhanced Treg homeostasis. Although patients in the extended duration cohort sustain stably elevated Treg numbers, it is not known whether a long-lasting Treg response is preserved in the absence of exogenous IL-2. Post-IL-2 Treg monitoring results were available for 3 patients in the extended duration cohort. One patient received continuous IL-2 for approximately 3 years and 2 patients stopped after over 1 year of therapy. At 4 weeks after IL-2 discontinuation, 2 of the 3 patients maintained elevated Treg numbers. One patient who stopped IL-2 after 74 weeks due to renal insufficiency had stably elevated Treg numbers and no worsening of cGVHD at 9 months post-IL-2, indicating that some patients may have durable restoration of Treg homeostasis following extended IL-2 therapy (Figure 3).There were no significant differences in absolute numbers of Treg, Tcon or CD8 subsets between clinical responders and non-responders during the initial 12 week treatment period. Plasma IL-2 and soluble IL-2 receptor levels were also similar between the two groups. Thus, differences in clinical response to IL-2 are likely determined by qualitative differences in Treg or effector T cell function. Functional Treg suppression assays and gene expression profiling studies are in progress to explore this possibility. [Display omitted] [Display omitted] [Display omitted] DisclosuresArmand:Bristol-Myers Squibb: Research Funding; Merck: Consultancy, Research Funding; Infinity Pharmaceuticals: Consultancy. Soiffer:Gentium SpA/Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees. Antin:Gentium S.p.A.: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.

Efficacy and safety of bio-chemotherapy with dacarbazine plus interleukin-2 in patients with unresectable malignant melanoma.

This study retrospectively evaluated the toxicity and efficacy of dacarbazine (DTIC) with low-dose subcutaneous interleukin-2 (IL-2) for patients with advanced melanoma. Patients with unresectable malignant melanoma received bio-chemotherapy DTIC (330 mg/m(2) , every 3 weeks ) and IL-2 18 MIU (million international units) in divided doses by subcutaneous injection three times a week for 4 weeks. Treatment was performed for six cycles or until disease progression or unbearable toxicity. From October 2006 to November 2013, up to 31 patients (17 men; 14 women) were enrolled. Their median age was 48 years (range, 22-81 years). Subtypes of melanoma included 11 (35.4%) acral lentiginous, nodular, 1 (3.2%) superficial spreading, 10 (32.2%) mucosal and 5 (16.1%) others. The response rate was 19.3%, including 3.2% with a complete response, 16.1% with a partial response and 6.3% with stable disease. The median progression-free survival time was 3.5 months (95% CI: 3.0-3.9 months). The median overall survival time was 8.6 months (95% CI: 4.1-10.9 months). The 1-year survival rate was 39% and the 5-year survival rate was 10%. Our data demonstrated that low-dose subcutaneous IL-2 plus DTIC has modest efficacy and may produce long-term survival in small proportion of patients. Furthermore, the treatment is well tolerated by patients.

Abstract CT304: Anti-prostate-specific membrane antigen (PSMA) monoclonal antibody J591 plus low-dose interleukin-2 (IL-2) in patients with recurrent prostate cancer (PC)

Abstract Purpose J591 is a deimmunized monoclonal antibody that recognizes the extracellular domain of prostate specific membrane antigen and was engineered to induce antibody-dependent cellular cytotoxicity (ADCC). Low dose IL-2 may enhance ADCC by the clonal expansion of natural killer (NK) cells. Immunotherapy may result in survival benefits independent of immediate response. This study was designed to evaluate the efficacy of J591 plus low-dose IL-2 in men with recurrent PC. Experimental Design Men with non-castrate PSA relapse or progressive metastatic castration-resistant PC (mCRPC) received low-dose subcutaneous IL-2 (1.2 x 106 IU/m2/day) daily for 8 weeks with weekly intravenous infusions of J591 (25 mg/m2) on weeks 4-6 (considered 1 cycle). A maximum of 3 cycles of therapy was delivered. Flow cytometry was used to assess NK cell fraction and PSA changes and survival were assessed. Results Seventeen men were treated between 2001-03 with at least 1 cycle (11 PSA relapse, 6 mCRPC) with 1 mCRPC patient invaluable for PSA response due to a protocol violation (9 received 2 cycles, 2 received 3 cycles). Toxicity was mild, mainly limited to fatigue and injection site reactions. PSA declined following therapy in 7 of 16 evaluable men (2 of 5 mCRPC). Flow cytometry revealed an average increase in absolute NK cell count of 107% at week 4 and 117% at the end of cycle 1. NK cell percent change had an inverse relationship with PSA (2-tailed Pearson correlation coefficient r = -0.46; P= 0.13). The actual vs. predicted survival for 5 of the 6 men with mCRPC disease was greater than predicted by the Halabi nomogram (Spearman correlation analysis r=0.29; P=0.57, i.e. nomogram did not predict survival). Conclusions The combination of J591 with low-dose IL-2 was well tolerated in patients with biochemical relapse and mCRPC. In addition, combination therapy expanded NK cell fraction which correlated with PSA decline. Interestingly, long term follow up reveals that 83% of the mCRPC group lived significantly longer than predicted, mostly in an era prior to the approval of life-prolonging therapy. These data support further testing of the hypothesis that PSMA-directed immunotherapy influences outcome in men with advanced PC. Citation Format: Yulian Khagi, Gurveen Kaur, Paul Christos, Naveed H. Akhtar, David M. Nanus, Neil H. Bander, Scott T. Tagawa. Anti-prostate-specific membrane antigen (PSMA) monoclonal antibody J591 plus low-dose interleukin-2 (IL-2) in patients with recurrent prostate cancer (PC). [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr CT304. doi:10.1158/1538-7445.AM2014-CT304

Dendritic cell-based vaccine in advanced melanoma

Dendritic cells (DCs) are unique specialized antigen-presenting cells capable of priming naive T cells and inducing antigen-specific cytotoxic T lymphocytes. This study presents an update of clinical results from a DC-based phase I-II clinical vaccine trial in stage IV melanoma. From 2003 to 2010, 27 patients with metastatic melanoma were treated with mature DCs pulsed with autologous tumor lysate and keyhole limpet hemocyanin and with subcutaneous low-dose interleukin-2. Delayed-type hypersensitivity (DTH) tests for in-vivo immunomonitoring were performed at baseline and every four vaccinations thereafter. Two complete, two mixed and six partial responses, and five stable diseases were observed (overall response, 37.0%; clinical benefit, 55.5%). All 15 responders showed DTH positivity. A median overall survival of 22.9 months [95% confidence interval (CI): 13.4-61.3] for DTH-positive patients (19) and 4.8 months (95% CI: 3.9-11.9) for DTH-negative patients (8; log rank=7.26; P=0.007) was observed. The overall median overall survival was 16 months (95% CI: 9-33). Our results would seem to highlight a relationship between positive-DTH test and an improved survival.

Interleukin-2 and Regulatory T Cells in Graft-versus-Host Disease

Dysfunction of regulatory T (Treg) cells has been detected in diverse inflammatory disorders, including chronic graft-versus-host disease (GVHD). Interleukin-2 is critical for Treg cell growth, survival, and activity. We hypothesized that low-dose interleukin-2 could preferentially enhance Treg cells in vivo and suppress clinical manifestations of chronic GVHD. In this observational cohort study, patients with chronic GVHD that was refractory to glucocorticoid therapy received daily low-dose subcutaneous interleukin-2 (0.3×10(6), 1×10(6), or 3×10(6) IU per square meter of body-surface area) for 8 weeks. The end points were safety and clinical and immunologic response. After a 4-week hiatus, patients with a response could receive interleukin-2 for an extended period. A total of 29 patients were enrolled. None had progression of chronic GVHD or relapse of a hematologic cancer. The maximum tolerated dose of interleukin-2 was 1×10(6) IU per square meter. The highest dose level induced unacceptable constitutional symptoms. Of the 23 patients who could be evaluated for response, 12 had major responses involving multiple sites. The numbers of CD4+ Treg cells were preferentially increased in all patients, with a peak median value, at 4 weeks, that was more than eight times the baseline value (P<0.001), without affecting CD4+ conventional T (Tcon) cells. The Treg:Tcon ratio increased to a median of more than five times the baseline value (P<0.001). The Treg cell count and Treg:Tcon ratio remained elevated at 8 weeks (P<0.001 for both comparisons with baseline values), then declined when the patients were not receiving interleukin-2. The increased numbers of Treg cells expressed the transcription factor forkhead box P3 (FOXP3) and could inhibit autologous Tcon cells. Immunologic and clinical responses were sustained in patients who received interleukin-2 for an extended period, permitting the glucocorticoid dose to be tapered by a mean of 60% (range, 25 to 100). Daily low-dose interleukin-2 was safely administered in patients with active chronic GVHD that was refractory to glucocorticoid therapy. Administration was associated with preferential, sustained Treg cell expansion in vivo and amelioration of the manifestations of chronic GVHD in a substantial proportion of patients. (Funded by a Dana-Farber Dunkin' Donuts Rising Star award and others; ClinicalTrials.gov number, NCT00529035.).

Pilot study of granulocyte-macrophage colony-stimulating factor and interleukin-2 as immune adjuvants for a melanoma peptide vaccine

Thus far, peptide vaccines used to stimulate tumor-specific immune responses in patients with melanoma have been largely unsuccessful. Granulocyte-macrophage colony-stimulating factor and interleukin-2 are immune-potentiating cytokines that have improved vaccine responses in preclinical models. We hypothesized that higher doses of granulocyte-macrophage colony-stimulating factor and addition of low-dose interleukin-2 might augment responses to vaccine antigens. Patients with resected stage II, III, or IV melanoma were treated with vaccines containing three melanoma-associated peptides [MART-1a, gp100(207-217), and survivin], along with 300 or 500 mcg granulocyte-macrophage colony-stimulating factor in Montanide ISA. Cohorts of patients received low-dose subcutaneous interleukin-2 on days 7-20 after vaccination. Induction of a response was defined as either doubling of cytotoxic T lymphocyte frequency from baseline or increase in frequency from undetectable (<0.05%) to detectable. Leukocyte subsets and plasma cytokines were analyzed before and after vaccination. Cytotoxic T lymphocyte responses to MART-1a, gp100(207-217), and survivin were induced in 11, 16, and 14 of 19 patients, respectively. Responses were not higher in patients receiving 500 mcg granulocyte-macrophage colony-stimulating factor or low-dose interleukin-2 than in patients receiving 300 mcg granulocyte-macrophage colony-stimulating factor only. Interleukin-2 treatment (in nine patients) led to increases in natural killer cells and T regulatory cells compared with no interleukin-2 treatment (nine patients). Multiple plasma cytokines were transiently induced during vaccination. Neither increasing the dose of granulocyte-macrophage colony-stimulating factor nor addition of low-dose interleukin-2 resulted in an increase in the frequency of vaccine-specific cytotoxic T lymphocytes to a melanoma peptide vaccine. The increase in T regulatory cells associated with interleukin-2 treatment suggests that interleukin-2 may be immunosuppressive in this setting.

Maintenance immunotherapy in recurrent ovarian cancer: Long term follow-up of a phase II study

Objectives Vascular endothelial growth factor (VEGF), a mediator of tumor-associated immunodeficiency, plays a key role in angiogenesis and is a prognostic factor in advanced ovarian cancer (AOC). Previously, we showed that low-dose interleukin-2 (IL-2) and 13-cis-retinoic acid (RA) improved the tumor-associated immunodeficiency and decreased VEGF in patients with AOC. Here, we report long term follow-up of a group of patients with platinum-sensitive AOC who were treated with IL-2 and RA. Methods Sixty-five patients with AOC who had a clinical benefit from second line chemotherapy and elevated serum levels of VEGF were entered into the study from 04/98 to 04/05. Therapy consisted of low-dose subcutaneous IL-2 and oral RA, administered on intermittent schedules for up to 5 years. Results A statistically significant improvement in lymphocyte and NK counts and a decrease in VEGF levels were observed with respect to baseline values among the 65 evaluable patients. Five-year progression-free survival and overall survival rate were 29% and 38%, respectively. Conclusions These data show that patients treated with low-dose IL-2 and RA have a statistically significant improvement in their lymphocyte and NK counts, a decrease in VEGF, and seem to have an improved clinical outcome.

Dendritic cell vaccination in melanoma patients: Update and subgroup analysis of clinical response to post-vaccine treatment

9042 Background: Dendritic cells (DCs) play a crucial role in the interplay between innate and adaptive immune response towards cancer. The combination of immunotherapies with standard treatments for cancer could represent a further chance for advanced melanoma patients. In the literature, higher response rates than those normally obtained have been reported after second-line chemotherapy in patients with non small cell lung cancer pre-treated with vaccines and in patients with follicular B-cell lymphoma vaccinated with an anti-idiotype vaccine whilst in remission. On the basis of this data, we reviewed and updated the clinical results of our dendritic cell based vaccine clinical trial in stage IV melanoma patients. Methods: From December 2002 to 2007, 24 pre-treated metastatic melanoma patients were vaccinated with mature DCs (mDCs) pulsed with autologous tumor lysate (ATL) and keyhole limpet hemocyanin (KLH) followed by a 5-day treatment with low-dose subcutaneous Interleukin-2. Results: We observed 2 complete response (CR), 2 mixed response (MR), 5 partial response (PR), 4 stable disease (SD) and 11 progressive disease (PD) (overall response (OS) 37.5%; clinical benefit 54.1%). All 13 responders had delayed-type hypersensitivity (DTH) positivity to KLH, of whom 10 also showed positivity to the lysate. Eleven (45.8%) of the 24 patients underwent further lines of treatment (5 chemotherapy [CT], 3 surgery [S], 4 biotherapy, 2 radiotherapy [RT] and 4 biochemotherapy [BioCT]) after stopping vaccination (8 due to progression and 3, in SD, because all of their lysate had been used). Of these 11 patients, 2 obtained CR (1 RT, 1 S), 5 PR (3 BioCT, 2 S) for an OR of 63.6%, 1 SD (BioCT) and 3 showed PD as the best response to subsequent therapies, with a median OS of 30 months (range 16–52). Of the 3 SD patients who were forced to stop vaccine treatment, 1 had CR following RT and 2 progressed. Conclusions: Metastatic melanoma responds poorly to standard therapy, in particular after first-line treatment. Vaccination could enhance clinical response to subsequent third- or fourth-line therapies, thus prolonging overall survival. No significant financial relationships to disclose.

Vaccination of renal cell cancer (RCC) patients with modified vaccinia Ankara (MVA) delivering tumor antigen 5T4 administered alone or with interleukin 2 (IL-2) or interferon-alpha (IFN)

3026 Background: The attenuated vaccinia virus (MVA) has been engineered to deliver the tumor antigen 5T4 (TroVax). More than 90% of RCCs overexpress the 5T4 antigen. A series of clinical trials were conducted to evaluate the effectiveness of MVA 5T4 as a single agent or in combination with either IL-2 or IFN. Methods: Eligibility: Pathologic diagnosis of clear cell or papillary RCC, progressive measurable metastases, any prior therapy, adequate physiological parameters, Karnofsky performance status ≥ 80%, and no active CNS involvement. A regimen of MVA 5T4 alone or in combination with IL-2 or IFN was given. Results: A total of 53 pts received MVA 5T4 alone or in combination with IL-2 or IFN. 13 pts received MVA 5T4 alone, 25 pts received low dose subcutaneous IL-2, and 15 pts received IFN. Clinical responses were assessed by measuring changes in tumor burden via computed tomography or magnetic resonance imaging scan. 5T4-specific cellular and humoral responses were monitored throughout the study. 5T4 was well tolerated with no serious adverse events attributed to vaccination. Of 48 intention-to-treat pts, 43 mounted 5T4-specific antibody responses. 2 pts showed a complete response for &gt; 36 months and 2 other pts had a partial response for &gt; 24 months and 12 months respectively. 20 pts demonstrated disease stabilization for ≥ 3 months. Median progression-free survival and overall survival for all pts was 3.6 months (range 0.8–29.7) and 13.2 months (range 1–38) respectively. A significant relationship was detected between the magnitude of 5T4-specific antibody response and overall survival. Conclusions: 5T4, whether administered alone or in combination, was well tolerated. High frequency of 5T4-specific immune responses and associated enhanced patient survival is encouraging and warrants further investigation. [Table: see text]

Italian multicentric trial comparing chemotherapy with or without low-dose interleukin-2 (IL-2) in advanced non-small cell lung cancer: Results from a phase III randomized study

8019 Background: Non small cell lung cancer (NSCLC) is associated with an IL-2-dependent cell-mediated immunodeficiency, and lymphocyte count is considered an independent prognostic factor as it seems to correlate with overall survival. A phase III randomized Italian multicentric trial was conducted to evaluate the efficacy of subcutaneous low-dose IL-2 added to standard CT on overall survival (OS) in advanced NSCLC patients. Methods: Histologically/cytologically confirmed stage IIIb or IV non operable NSCLC patients with measurable disease, ECOG PS 0–2, age 18–70 years and adequate bone marrow, renal and liver function were eligible for the study. Randomization was stratified by center, ECOG PS, stage of disease and percentage of weight loss. All patients received gemcitabine (1000mg/m2) on days 1 and 8 + cisplatin (100mg/m2) on day 2 every 21 days for a maximum of 6 cycles (CT arm). In the CT+IL-2 arm, patients also received low-dose subcutaneous IL-2 3,000,000 IU/die on days 3–5; 9–11; 15–17. The study had 90% power to detect a 20% absolute increase in 1-year OS with 120 patients per arm. Results: From June 2000 to October 2004, 241 patients were randomized (arm A/B: 127/114). At a median follow up of 32 months, 1-year OS was 45% for the CT+IL-2 arm vs. 51% for the CT arm (p=0.456 logrank). Median progression-free survival was 6.6 months in the CT+IL-2 arm vs. 6.9 months in the CT arm (p=0.573, logrank). Conclusions: The study did not show any relevant difference in clinical outcome by the addition of IL-2 to CT. Safety and subgroup analyses are ongoing to verify the efficacy of IL-2 as a function of clinical and biological characteristics of patients and tumors. Future studies to investigate the role of T-regulators in chemoimmunotherapy strategies, unknown when the study was originally planned, could be conducted. No significant financial relationships to disclose.

Predicting Prostate Cancer Biochemical Recurrence Using a Panel of Serum Proteomic Biomarkers

The pathological state of the prostate may be reflected by serum proteome in a man. We hypothesized that biomarkers are present in preoperative serum, which may be used to predict the probability of biochemical recurrence following radical prostatectomy. Mass spectrometry analysis was used to compare 52 men who experienced biochemical recurrence with 52 who remained biochemical recurrence-free for approximately 5 years after radical retropubic prostatectomy. A total of 30 matched pairs of recurrent and nonrecurrent serum samples were randomly selected as a training set for biomarker discovery and model development. Selected mass spectrometry peaks were combined with pre-radical retropubic prostatectomy prostate specific antigen in a multivariate algorithm to predict recurrence. The algorithm was evaluated using the remaining 22 recurrent and 22 nonrecurrent subjects as test samples. Protein identities of the selected mass spectrometry peaks were investigated. Two serum biomarkers for recurrence, P1 and P2, were combined with preoperative prostate specific antigen to predict biochemical recurrence. The ROC AUC for prostate specific antigen and the predicted outcome was 0.606 and 0.691 in the testing data, respectively. Using a single cutoff the samples were divided into 2 groups that were predictive of biochemical recurrence (p = 0.026). In contrast, preoperative prostate specific antigen did not differ between recurrent and nonrecurrent cases (Wilcoxon matched pairs test p = 0.07). The protein identity of P1 was determined to be a truncated form of C4a (C4a des-Arg). Preliminary data indicated that P2 was an N-terminal fragment of protein C inhibitor. In the current study population, which was matched on Gleason score and TNM staging, pre-radical retropubic prostatectomy prostate specific antigen retained no independent power to predict recurrence. However, by adding 2 proteomic biomarkers to preoperative prostate specific antigen the combined model demonstrated statistically significant value for predicting prostate cancer recurrence in men who underwent radical retropubic prostatectomy.

Adoptive immunotherapy for indolent non-Hodgkin lymphoma and mantle cell lymphoma using genetically modified autologous CD20-specific T cells

AbstractAdoptive immunotherapy with T cells expressing a tumor-specific chimeric T-cell receptor is a promising approach to cancer therapy that has not previously been explored for the treatment of lymphoma in human subjects. We report the results of a proof-of-concept clinical trial in which patients with relapsed or refractory indolent B-cell lymphoma or mantle cell lymphoma were treated with autologous T cells genetically modified by electroporation with a vector plasmid encoding a CD20-specific chimeric T-cell receptor and neomycin resistance gene. Transfected cells were immunophenotypically similar to CD8+ effector cells and showed CD20-specific cytotoxicity in vitro. Seven patients received a total of 20 T-cell infusions, with minimal toxicities. Modified T cells persisted in vivo 1 to 3 weeks in the first 3 patients, who received T cells produced by limiting dilution methods, but persisted 5 to 9 weeks in the next 4 patients who received T cells produced in bulk cultures followed by 14 days of low-dose subcutaneous interleukin-2 (IL-2) injections. Of the 7 treated patients, 2 maintained a previous complete response, 1 achieved a partial response, and 4 had stable disease. These results show the safety, feasibility, and potential antitumor activity of adoptive T-cell therapy using this approach. This trial was registered at www.clinicaltrials.gov as #NCT00012207.

Maintenance Immunotherapy in Recurrent or Metastatic Squamous Cell Carcinoma of the Head and Neck

The purpose of this study was to determine the efficacy and safety of a maintenance immunotherapy regimen administered to patients with recurrent/metastatic squamous cell carcinoma of the head and neck (RMHN) who showed clinical benefit from docetaxel, ifosfamide, and cisplatin chemotherapy (DIP). Every 4 weeks, patients with RMHN received 60 mg/m docetaxel on day 1, and 1200 mg/m ifosfamide and 20 mg/m cisplatin on days 1 to 4. Low-dose subcutaneous interleukin-2 and oral 13-cis-retinoic acid were administered as maintenance immunotherapy to patients who showed a clinical benefit (complete or partial response, disease stability). The primary end point was response; secondary end points were progression-free survival, overall survival, toxicity, and evaluations of lymphocytes, natural killer cells, and serum vascular endothelial growth factor (VEGF). After a median follow-up of 22 months, 263 courses of chemotherapy were administered to the 54 patients. The overall response rate was 59%. Forty-two patients (78%) had a clinical benefit and received 185 courses of maintenance immunotherapy. Median progression-free survival and overall survival were 11.1 and 21.8 months, respectively. Statistically significant, progressive increases in lymphocytes and natural killer cells and a decrease in VEGF were observed in patients treated with maintenance immunotherapy. The toxicity was relatively well tolerated and caused no death. Outpatient administration of DIP, followed by low-dose interleukin-2 and 13-cis-retinoic acid, was generally well tolerated and showed promising activity against RMHN. Longitudinal changes in lymphocytes, natural killer cells, and VEGF might be useful biomarkers for response and survival.