Circulating Tumor DNA Testing Supports Rapid Therapeutic Decision-Making in Metastatic Melanoma: A Case Report.

Treatment of metastatic melanoma includes the option of targeted therapy in patients with driver BRAF mutations. BRAF-MEK inhibitor drugs improve survival in the approximately 50% of patients with melanoma that harbor BRAF mutations. As BRAF mutation detection in tissue often takes days to weeks, it is not always possible or timely to obtain BRAF status in tissue using immunohistochemistry or next generation sequencing. Plasma-derived circulating tumor DNA (ctDNA) is a potential alternative analyte in such treatment settings. We present a case of metastatic melanoma that was treated in an emergent setting using therapy supported by rapid PCR-based detection of ctDNA positive for a BRAF V600 mutation. In this rapidly deteriorating 53-year-old male with diffuse melanoma metastases and unknown BRAF mutation status requiring hospital admission, a plasma-based BRAF mutation detection supported treatment with targeted therapy, dabrafenib and trametinib. Same-day initiation of therapy resulted in swift amelioration allowing discharge within a week, followed by substantial clinical improvement over the following weeks. In cases requiring urgent clinical decision making, a plasma-based, near point-of-care detection system is useful in supporting targeted therapy decisions without the need for invasive and time-consuming biopsy.

IL-2 and IL-15 manifest opposing effects on activation of nuclear factor of activated T cells.

IL-2 and IL-15 are cytokines involved in T cell activation and death. Their non-shared receptors, IL-2Ralpha and IL-15Ralpha, are important in the homeostasis of lymphocytes as evidenced by gene deletion studies. How these cytokine/receptor systems affect T cell antigen receptor signaling pathways is poorly understood. Here, we show that the IL-2 and IL-15 cytokine/receptor alpha systems regulate activation of nuclear factor of activated T cells (NF-AT) in opposing ways. IL-15Ralpha increased while IL-2Ralpha decreased basal NF-AT activation status in a Jurkat transient transfection model. The effect of each of the alpha chain receptors on NF-AT activation was further opposed by addition of the respective cytokine. These effects were inhibited by anti-cytokine and anti-cytokine receptor reagents as well as by inhibitors of TCR signaling. These results suggest a novel pathway of cytokine action to regulate T cell signaling, activation, death, and homeostasis.

Oligomerization of IL-2Ralpha.

Interleukin (IL) 2 receptor subunit alpha (IL-2Ralpha) increases the affinity of the IL-2 receptor complex while hetero-association of IL-2Rbeta and gamma(c) chains initiates a proliferative signal. We show here that IL-2Ralpha is necessary for receptor clustering required for augmentation of IL-2 signalling. Cells expressing chimeras incorporating the extracellular domain of IL-2Ralpha demonstrated IL-2 independent homo-association of the IL-2Ralpha chimera. Singly or co-transfected IL-2Rbeta and gamma(c) chimeras showed no spontaneous or IL-2-inducible oligomerization. Co-transfection of IL-2Ralpha and IL-2Rbeta (+/- gamma(c)) chimeras diminished spontaneous IL-2Ralpha chimera oligomerization and permitted IL-2-inducible hetero-oligomerization of receptor components. Homo-association of IL-2Ralpha was also demonstrated by fluorescence resonance energy transfer (FRET). The spontaneous homo-oligomerization property of IL-2Ralpha required the membrane proximal region of the receptor (exon 6) by deletion analysis; the IL-2 inducible oligomerization property of IL-2Ralpha required the second "sushi" domain (exon 4). This work provides insight into the mechanics of this complex receptor system and to other receptor complexes in the immune system that send signals by clustering receptor subunits.