Joseph Broucek

High-dose IL-2 induces rapid albumin uptake by endothelial cells through Src-dependent caveolae-mediated endocytosis

High-dose interleukin-2 (HDIL2) treatment of patients with metastatic melanoma and renal cell carcinoma is associated with durable responses, but therapy is accompanied by significant toxicity related to vascular leak syndrome (VLS). Currently, the cause of VLS is not well defined; however, based on the role of endothelial cell (EC) permeability in VLS and the commonly observed hypoalbuminemia in patients receiving HDIL2 therapy, we established an in vitro approach utilizing primary human pulmonary microvascular ECs to monitor the effect of HDIL2 therapy on albumin uptake. We found that HDIL2 treatment of ECs results in albumin colocalization with caveolin-1 leading to albumin uptake by ECs. This albumin uptake occurs through caveolae-mediated but not clathrin-mediated endocytosis and is abrogated with inhibition of the Src tyrosine kinase pathway. These findings provide insight into how IL-2 induces VLS and may help identify potential targets for prevention of toxicity without affecting the therapeutic activity of HDIL2.

Interleukin-2 alters distribution of CD144 (VE-cadherin) in endothelial cells

BackgroundHigh-dose IL-2 (HDIL2) is approved for the treatment of metastatic melanoma and renal cell carcinoma, but its use is limited in part by toxicity related to the development of vascular leak syndrome (VLS). Therefore, an understanding of the mechanisms that underlie the initiation and progression of HDIL2-induced increases in endothelial cell (EC) permeability leading to VLS are of clinical importance.MethodsWe established a novel ex vivo approach utilizing primary human pulmonary microvascular ECs to evaluate EC barrier dysfunction in response to IL-2.ResultsComplementary in vitro studies using exogenous IL-2 and ex vivo studies using serum from patients treated with IL-2 demonstrate that HDIL2 induces VLS through CD144 (vascular endothelial (VE)-cadherin) redistribution.ConclusionsThese findings provide new insight into how IL-2 induces VLS and identifies VE-cadherin as a potential target for preventing IL-2-related VLS.

Metastasectomy following incomplete response to high-dose interleukin-2

To evaluate our experience with metastasectomy following partial response or stable disease after treatment with high‐dose interleukin‐2 (HD IL‐2).

Combination immunotherapy with anti-CTLA-4 and interleukin-2 redirects regulatory T cells into tumor-draining lymph nodes and expands anti-tumor CD8+ T cells in the tumor microenvironment

Meeting abstracts Monotherapy with Ipilimumab (anti-CTLA-4 antibody) and monotherapy with IL-2 (T cell stimulating cytokine) are approved for the treatment of metastatic melanoma. Combination immunotherapy has been suggested as a more potent regimen but has not been sufficiently investigated. We

Role of dose selection in successful interleukin-2 immunotherapy: solving the Goldilock’s Complex

Background The results of multiple IL-2 clinical trials (using a range of doses) have shown similar trends, in which IL-2 immunotherapy leads to an objective response in 15-20% of patients and complete response in 5-10%. Importantly, the majority (> 80%) of patients with a complete response maintain long-term responses. However, why only a small proportion of patients attain a complete response at any dose is unknown. We hypothesized that the dose of IL-2 utilized, differentially targets CD8+ effector versus CD4+ regulatory T cells based on patient-specific characteristics, and that an optimal IL-2 dose may be needed to achieve therapeutic benefit.

PD-1 blockade reverses viral infection-induced loss of anti-tumor CD8+ T cell responses

Background Emerging epidemiologic studies describe an increased prevalence of tumors in patients with non-oncogenic chronic viral disease (e.g., non-AIDS-defining cancers in HIV and non-hepatic cancers in HCV). However, there is a lack of basic understanding by what mechanism infections result in increased unrelated cancer formation (i.e., of cancers in tissues not associated with the infection), and what role immunotherapy may have in the rescue of immune responses under this condition. We hypothesized that viral infections establish an immunological environment that leads to the preferential loss of anti-tumor (i.e., anti-self antigen) CD8+ T cells, resulting in the inability of the host to avert tumor growth, and further that immunotherapy can reverse this detrimental loss. Methods

Combination immunotherapy with Interleukin-2 and CTLA-4 blockade decreases tumor growth and improves overall survival

Background Combination immunotherapy is quickly gaining attention in the field of cancer treatment. IL-2, a cytokine which activates T cells, and ipilimumab, a monoclonal antibody that blocks CTLA-4, are both approved as monotherapy for metastatic melanoma. To date, combination immu- notherapy with IL-2 and anti-CTLA-4 has not been adequately investigated. We hypothesized that that this combination may work synergistically owing to its dis- tinct but complementary mechanisms of T cell activation. mean tumor area with combination IL-2 and anti-CTLA- 4 was 2mm2, while in the anti-CTLA-4 only, IL-2 only, and placebo groups it was 14, 29, and 68 mm2, respec- tively (P<0.01 for all comparisons, except IL-2 only ver- sus anti-CTLA-4 only (not significant)). At day 30, the overall survival with combination IL-2 and anti-CTLA-4 was 50%, while with IL-2 only and anti-CTLA-4 only it was 10% and 20%, respectively (P<0.01 for all compari- sons). All animals treated in the placebo group suc- cumbed to their tumors by day 19. These findings were confirmed in a second experiment with similar results. Conclusions Combination immunotherapy with IL-2 activation and CTLA-4 blockade significantly decreases tumor growth and increases overall survival when compared to IL-2 or anti-CTLA-4 monotherapy or placebo. The role of CD8+ effector versus CD4+ regulatory T cells in the success of this immunotherapy is ongoing and will be included in our formal presentation. Ultimately, we aim to translate this work into a combination immunotherapy clinical trial.