Sunitinib does not attenuate contractile force following a period of ischemia in isolated human cardiac muscle.

Concerns have been raised about the development of heart failure in patients treated for cancer with angiogenesis inhibitors, such as the tyrosine kinase inhibitor sunitinib. Patients with previous coronary artery disease and hypertension have an increased risk of developing heart failure. Therefore, we studied the effect of sunitinib on the contractility of isolated human atrial trabeculae and the effect on recovery after ischemic stimulation. After informed consent, the atrial appendage of patients undergoing cardiac surgery was harvested and isolated trabeculae were placed in an organ bath with a force transducer. During electrical stimulation, contractile force was measured during normal pacing or after simulated ischemia. Of each patient, one trabecula was perfused with control and one with sunitinib. Contractile force (expressed as percentage of baseline force) declined over time to 57 ± 8 and 73 ± 20% after 150 min of stimulation for solvent- and sunitinib-treated trabeculae, respectively (mean ± SE; n = 8; p > 0.1). After simulated ischemia and reperfusion, contractile force was 40 ± 6% in the control compared to 39 ± 6% in the sunitinib-treated trabeculae during the last final 5 min of reperfusion (n = 12; p > 0.1). Sunitinib at low, but clinically relevant, concentrations does not have a direct effect on function of human atrial cardiomyocytes nor does it attenuate the recovery in contractile force of atrial cardiomyocytes after a period of ischemia. A direct and acute toxic effect on cardiomyocytes does not explain the development of heart failure in patients treated with sunitinib.

Aberrant expression of the hematopoietic-restricted minor histocompatibility antigen LRH-1 on solid tumors results in efficient cytotoxic T cell-mediated lysis.

CD8(+) T cells recognizing minor histocompatibility antigens (MiHA) on solid tumor cells may mediate effective graft-versus-tumor (GVT) reactivity after allogeneic stem cell transplantation (SCT). Previously, we identified LRH-1 as a hematopoietic-restricted MiHA encoded by the P2X5 gene. Here, we report that LRH-1 is aberrantly expressed on solid tumor cells. P2X5 mRNA expression is demonstrated in a significant portion of solid tumor cell lines, including renal cell carcinoma (RCC), melanoma, colorectal carcinoma, brain cancer and breast cancer. Importantly, P2X5 gene expression was also detected in a subset of primary solid tumor specimens derived from RCC, brain cancer and breast cancer patients. Furthermore, P2X5 expressing solid tumor cells can be effectively targeted by LRH-1-specific cytotoxic T lymphocytes under inflammatory conditions. The expression of HLA-B7 and CD54 on tumor cells increases upon cytokine stimulation resulting in improved T cell activation as observed by higher levels of degranulation and enhanced tumor cell lysis. Overall, hematopoietic-restricted MiHA LRH-1 is aberrantly expressed on solid tumor cells and may be used as target in GVT-specific immunotherapy after SCT.

Myeloid leukemic progenitor cells can be specifically targeted by minor histocompatibility antigen LRH-1-reactive cytotoxic T cells.

CD8(+) T cells recognizing minor histocompatibility antigens (MiHAs) on leukemic stem and progenitor cells play a pivotal role in effective graft-versus-leukemia reactivity after allogeneic stem cell transplantation (SCT). Previously, we identified a hematopoiesis-restricted MiHA, designated LRH-1, which is presented by HLA-B7 and encoded by the P2X5 purinergic receptor gene. We found that P2X5 is significantly expressed in CD34(+) leukemic subpopulations from chronic myeloid leukemia (CML) and acute myeloid leukemia (AML) patients. Here, we demonstrate that LRH-1-specific CD8(+) T-cell responses are frequently induced in myeloid leukemia patients following donor lymphocyte infusions. Patients with high percentages of circulating LRH-1-specific CD8(+) T cells had no or only mild graft-versus-host disease. Functional analysis showed that LRH-1-specific cytotoxic T lymphocytes (CTLs) isolated from 2 different patients efficiently target LRH-1-positive leukemic CD34(+) progenitor cells from both CML and AML patients, whereas mature CML cells are only marginally lysed due to down-regulation of P2X5. Furthermore, we observed that relative resistance to LRH-1 CTL-mediated cell death due to elevated levels of antiapoptotic XIAP could be overcome by IFN-gamma prestimulation and increased CTL-target ratios. These findings provide a rationale for use of LRH-1 as immunotherapeutic target antigen to treat residual or persisting myeloid malignancies after allogeneic SCT.

The B cell antigen receptor controls AP-1 and NFAT activity through Ras-mediated activation of Ral.

Signaling by the BCR involves activation of several members of the Ras superfamily of small GTPases, among which is Ras itself. Ras can control the activity of multiple effectors, including Raf, PI3K, and guanine nucleotide exchange factors for the small GTPase Ral. Ras, Raf, and PI3K have been implicated in a variety of processes underlying B cell development, differentiation, and function; however, the role of Ral in B lymphocytes remains to be established. In this study, we show that Ral is activated upon BCR stimulation in human tonsillar and mouse splenic B lymphocytes and in B cell lines. Using signaling molecule-deficient B cells, we demonstrate that this activation is mediated by Lyn and Syk, Btk, phospholipase C-gamma2, and inositol-1,4,5-trisphosphate receptor-mediated Ca(2+) release. In addition, although Ral can be activated by Ras-independent mechanisms, we demonstrate that BCR-controlled activation of Ral is dependent on Ras. By means of expression of the dominant-negative mutants RasN17 and RalN28, or of RalBPDeltaGAP, a Ral effector mutant which sequesters active Ral, we show that Ras and Ral mediate BCR-controlled transcription of c-fos. Furthermore, while not involved in NF-kappaB activation, Ras and Ral mediate BCR-controlled activation of JUN/ATF2 and NFAT transcription factors. Taken together, our data show that Ral is activated upon BCR stimulation and mediates BCR-controlled activation of AP-1 and NFAT transcription factors. These findings suggest that Ral plays an important role in B cell development and function.