Expression of c-Kit discriminates between two functionally distinct subsets of human type 2 innate lymphoid cells.

Human type 2 innate lymphoid cells (ILC2) are the only ILC subset that shows heterogeneous expression of the SCF receptor c-Kit (CD117). Despite its use as surface marker to distinguish ILC populations, its influence on ILC2 biology has not been investigated. Here, we show that c-Kit expression of peripheral blood ILC distinguishes two functionally distinct ILC2 subsets (c-Kithi and c-Kitlo ). When examined for their potential for functional plasticity we found that c-Kitlo ILC2 displayed greater potential to produce type 2 cytokines, possibly representing fully mature and lineage committed ILC2. On the other hand, c-Kithi ILC2 coexpressed the ILC3-marker and chemokine receptor CCR6 and were able to mount a significant IL-17A response under ILC3-promoting conditions. In addition, c-Kithi ILC2 produced higher levels of IFN-γ than c-Kitlo ILC2 under ILC1-conditions. Although costimulation with SCF did not further influence ILC2 plasticity, it augmented type 2 cytokine production. We conclude that c-Kit marks distinct subpopulations of ILC2, which has therapeutic implications for conditions in which ILC2 are involved, such as allergy and asthma.

Sertoli cell-conditioned medium restores spermatogenesis in azoospermic mouse testis.

The current study evaluates potential applications of Sertoli cell (SC)-conditioned medium (CM) and explores the effects of the conditioned medium on the spermatogenesis process in azoospermic mice. For this study, 40 adult mice (28-30 g) were divided into 4 experimental groups: (1) control, (2) DMSO 2% (10 µl), (3) busulfan (40 mg/kg single dose) and (4) busulfan/CM (10 µl). SCs were isolated from 4-week-old mouse testes. After using anesthetics, 10 µl of CM was injected over 3-5 min into each testis and subsequently, sperm samples were collected from the tail of the epididymis. Afterward, the animals were euthanized and testis samples were taken for histopathology experiments and RNA extraction in order to examine the expression of c-kit, STRA8 and PCNA genes. The data showed that CM notably increased the total sperm count and the number of testicular cells, such as spermatogonia, primary spermatocytes, round spermatids, SCs and Leydig cells compared with the control, DMSO and busulfan groups. Furthermore, the results showed that expression of c-kit and STRA8 was significantly decreased in the busulfan and busulfan/SC groups at 8 weeks after the last injection (p < 0.001) but no significant difference was found for PCNA compared with the control and DMSO groups (p < 0.05). These findings suggest that the Sertoli cell-conditioned medium may be beneficial as a practical approach for therapeutic strategies in reproductive and regenerative medicine.

Regulation of Kit Expression in Early Mouse Embryos and ES Cells.

Kit is a growth factor receptor that regulates proliferation and/or survival of many embryonic and postnatal stem cell types. When mutated, it can induce malignant transformation of the host cells. To dissect the Kit role in the control of ESC pluripotency, we studied its expression during early mouse embryogenesis and during the process of ESC derivation from inner cell mass (ICM) cells. We followed the in vitro development of early mouse embryos obtained from transgenic mice carrying Kit promoter regions fused to EGFP (Kit-EGFP) and found that they initiate EGFP expression at morula stage. EGFP expression is then maintained in the blastocyst, within the ICM, and its levels increase when cultured in the presence of MAPK and GSK3ß inhibitors (2i) plus LIF compared with the LIF-only condition. Kit-EGFP ESCs showed nonhomogeneous EGFP expression pattern when cultured in LIF condition, but they upregulated EGFP expression, as well as that of Sox2, Nanog, Prdm14, when shifted to 2i-LIF culture. Similarly, primordial germ cells (PGCs) in the process of embryonic germ cell (EGC) conversion showed enhanced EGFP expression in 2i-LIF. Kit expression was affected by manipulating Sox2 levels in ESCs. Chromatin immunoprecipitation experiments confirmed that Sox2 binds Kit regulatory regions containing Sox2 consensus sequences. Finally, Kit constitutive activation induced by the D814Y mutation increased ESC proliferation and cloning efficiency in vitro and in teratoma assays in vivo. Our results identify Kit as a pluripotency-responsive gene and suggest a role for Kit in the regulation of ESC proliferation. Stem Cells 2019;37:332-344.

Low-grade oncocytic tumour of kidney (CD117-negative, cytokeratin 7-positive): a distinct entity?

AIM: To describe a group of distinct low-grade oncocytic renal tumours that demonstrate CD117 negative/cytokeratin (CK) 7-positive immunoprofile. METHODS AND RESULTS: We identified 28 such tumours from four large renal tumour archives. We performed immunohistochemistry for: CK7, CD117, PAX8, CD10, AMACR, e-cadherin, CK20, CA9, AE1/AE3, vimentin, BerEP4, MOC31, CK5/6, p63, HMB45, melan A, CD15 and FH. In 14 cases we performed array CGH, with a successful result in nine cases. Median patient age was 66 years (range 49-78 years) with a male-to-female ratio of 1:1.8. Median tumour size was 3 cm (range 1.1-13.5 cm). All were single tumours, solid and tan-brown, without a syndromic association. On microscopy, all cases showed solid and compact nested growth. There were frequent areas of oedematous stroma with loosely arranged cells. The tumour cells had oncocytic cytoplasm with uniformly round to oval nuclei, but without significant irregularities, and showed only focal perinuclear halos. Negative CD117 and positive CK7 reactivity were present in all cases (in two cases there was focal and very weak CD117 reactivity). Uniform reactivity was found for PAX8, AE1/AE3, e-cadherin, BerEP4 and MOC31. Negative stains included CA9, CK20, vimentin, CK5/6, p63, HMB45, Melan A and CD15. CD10 and AMACR were either negative or focally positive; FH was retained. On array CGH, there were frequent deletions at 19p13.3 (seven of nine), 1p36.33 (five of nine) and 19q13.11 (four of nine); disomic status was found in two of nine cases. On follow-up (mean 31.8 months, range 1-118), all patients were alive with no disease progression. CONCLUSION: Low-grade oncocytic tumours that are CD117-negative/CK7-positive demonstrate consistent and readily recognisable morphology, immunoprofile and indolent behaviour.

Primordial follicle activation is affected by the absence of Sohlh1 in mice.

Previous studies have reported that only primordial follicles and empty follicles can be found in 7.5 days postparturition (dpp) Sohlh1-/- mouse ovaries and females are infertility. There appears to be a defect in follicle development during the primordial-to-primary follicle transition in Sohlh1-/- mouse ovaries. However, detailed analyses of these phenomena have not been performed. In this study, we used Sohlh1-/- transgenic mice to explore the role of Sohlh1 in folliculogenesis. The results showed that only primordial follicles and empty follicles can be observed in Sohlh1-/- ovaries from 0.5 to 23.5 dpp. The expression of Foxo3 and FOXO3 was downregulated; nucleocytoplasmic shuttling of FOXO3 was normal in 7.5-dpp Sohlh1+/+ but not Sohlh1-/- ovaries; and primordial follicle activation (PFA) was not observed in 7.5-dpp Sohlh1-/- mice. The expression levels of KIT, AKT, and P308-AKT were downregulated (p < 0.05), whereas that of P473-AKT was not significantly changed (p > 0.05). The KIT/PI3K/AKT pathway was inhibited. Furthermore, we conducted a dual luciferase assay and chromatin immunoprecipitation. The results showed that SOHLH1 can upregulate the Kit gene by binding to the -3698 bp E-box motif. The absence of Sohlh1 may affect PFA in mouse ovaries via downregulation of Kit and inhibition of the KIT/PI3K/AKT pathway.

Stimulatory Effects of Mesenchymal Stem Cells on cKit+ Cardiac Stem Cells Are Mediated by SDF1/CXCR4 and SCF/cKit Signaling Pathways.

RATIONALE: Culture-expanded cells originating from cardiac tissue that express the cell surface receptor cKit are undergoing clinical testing as a cell source for heart failure and congenital heart disease. Although accumulating data support that mesenchymal stem cells (MSCs) enhance the efficacy of cardiac cKit(+) cells (CSCs), the underlying mechanism for this synergistic effect remains incompletely understood. OBJECTIVE: To test the hypothesis that MSCs stimulate endogenous CSCs to proliferate, migrate, and differentiate via the SDF1/CXCR4 and stem cell factor/cKit pathways. METHODS AND RESULTS: Using genetic lineage-tracing approaches, we show that in the postnatal murine heart, cKit(+) cells proliferate, migrate, and form cardiomyocytes, but not endothelial cells. CSCs exhibit marked chemotactic and proliferative responses when cocultured with MSCs but not with cardiac stromal cells. Antagonism of the CXCR4 pathway with AMD3100 (an SDF1/CXCR4 antagonist) inhibited MSC-induced CSC chemotaxis but stimulated CSC cardiomyogenesis (P<0.0001). Furthermore, MSCs enhanced CSC proliferation via the stem cell factor/cKit and SDF1/CXCR4 pathways (P<0.0001). CONCLUSIONS: Together these findings show that MSCs exhibit profound, yet differential, effects on CSC migration, proliferation, and differentiation and suggest a mechanism underlying the improved cardiac regeneration associated with combination therapy using CSCs and MSCs. These findings have important therapeutic implications for cell-based therapy strategies that use mixtures of CSCs and MSCs.

Establishing and characterizing lacrispheres from human lacrimal gland for potential clinical application.

PURPOSE: Lacrimal gland (LG) dysfunction leading to dry eye syndrome (DES) is an important cause of ocular morbidity. One of the potential and promising long-term management therapies for restoration of LG function could be transplantation of autologous ex vivo expanded stem cells. The present study was aimed at exploring the 2D and 3D cultures of human LG, identifying inherent stem cells and evaluating their secretory potential. METHODS: Fresh human lacrimal gland (HuLG) (n = 5) from patients undergoing therapeutic exenteration was harvested after ethical approval and informed consent. The gland was enzymatically digested and the isolated cells plated in Hepato-STIM media supplemented with l-glutamine, epidermal growth factor, fibroblast growth factor, and N-2 supplement. The native HuLG and the cultured spheres (DIV14-16) were evaluated for presence of stem cells (CD117 expression, quiescence, BrdU label retention, cell cycle, colony forming efficiency) and differentiation (secretion of tear proteins). RESULTS: Under the established culture conditions, suspension 3D cultures of human "lacrispheres" could be maintained and propagated for 3-4 weeks. The spheres consist of both acinar as well as ductal cells with evidence of stem cells (0.8 ± 0.05% CD117+ cells), BrdU label retention (9.31 ± 0.41%), G0/G1 profile similar to native lacrimal cells at isolation (76.9 versus 79.9%) and colony forming units (3.1%). The lacrispheres also secreted quantifiable levels of tear proteins (lysozyme, lactoferrin, scIgA) into the conditioned media. CONCLUSION: The study provides promising, first-of-its-kind evidence for the generation of lacrispheres from fresh HuLG, with enriched population of stem cells and secretory competent differentiated cells. The dual properties of these spheres make them a highly suitable source of transplantable cells for restoring the structure and function of damaged lacrimal gland.

CD117 (KIT) is a useful marker in the diagnosis of plasmablastic plasma cell myeloma.

AIMS: Plasmablastic plasma cell myeloma (PPCM) is a rare morphological presentation of multiple myeloma, and can resemble plasmablastic lymphoma (PBL), a human immunodeficiency virus (HIV)-related lymphoid neoplasm, morphologically and immunophenotypically. The aim of this study was to retrospectively search for factors that could help to differentiate these two entities. METHODS AND RESULTS: We used clinical looking glass, a data mining tool, to identify patients with a diagnosis of either PPCM or PBL with a CD117 test performed. We identified 27 cases from 16 patients in our institution with a diagnosis of either PPCM or PBL. There were 14 cases of PBL from eight patients and 13 cases of PPCM from eight patients. We found that six of eight patients with PPCM were positive for CD117, whereas no patients with PBL (0/8) showed positivity (P = 0.007 for patients). All (8/8) PPCM patients had a paraproteinaemia, whereas only two of eight PBL patients had a paraproteinaemia (P = 0.007). Lytic bone lesions were more indicative of a PPCM, with six of eight patients showing these lesions, and only one of eight PBL patients showing these lesions (P = 0.0406). HIV status was more likely to be positive in PBL patients, with six of seven patients showing positivity, and only one of five PPCM patients showing positivity (P = 0.072). Epstein-Barr virus-encoded RNA in-situ hybridization did not differentiate these two entities, with seven of eight PBL patients showing positivity, and two of five PPCM patients showing positivity (P = 0.2168). CONCLUSIONS: This study reveals that CD117 can be a useful marker to help differentiate PPCM from PBL and give the patient an appropriate prognosis and options for treatment.

Human Salivary Gland Stem Cells Functionally Restore Radiation Damaged Salivary Glands.

Adult stem cells are often touted as therapeutic agents in the regenerative medicine field, however data detailing both the engraftment and functional capabilities of solid tissue derived human adult epithelial stem cells is scarce. Here we show the isolation of adult human salivary gland (SG) stem/progenitor cells and demonstrate at the single cell level in vitro self-renewal and differentiation into multilineage organoids. We also show in vivo functionality, long-term engraftment, and functional restoration in a xenotransplantation model. Indeed, transplanted human salisphere-derived cells restored saliva production and greatly improved the regenerative potential of irradiated SGs. Further selection for c-Kit expression enriched for cells with enhanced regenerative potencies. Interestingly, interaction of transplanted cells with the recipient SG may also be involved in functional recovery. Thus, we show for the first time that salispheres cultured from human SGs contain stem/progenitor cells capable of self-renewal and differentiation and rescue of saliva production. Our study underpins the therapeutic promise of salisphere cell therapy for the treatment of xerostomia.

The Transcription Factor Ehf Is Involved in TGF-ß-Induced Suppression of FcεRI and c-Kit Expression and FcεRI-Mediated Activation in Mast Cells.

FcεRI, which is composed of α, ß, and γ subunits, plays an important role in IgE-mediated allergic responses. TGF-ß1 has been reported to suppress FcεRI and stem cell factor receptor c-Kit expression on mast cell surfaces and to suppress mast cell activation induced by cross-linking of FcεRI. However, the molecular mechanism by which these expressions and activation are suppressed by TGF-ß1 remains unclear. In this study, we found that the expression of Ets homologous factor (Ehf), a member of the Ets family transcriptional factors, is upregulated by TGF-ß/Smad signaling in mouse bone marrow-derived mast cells (BMMCs). Forced expression of Ehf in BMMCs repressed the transcription of genes encoding FcεRIα, FcεRIß, and c-Kit, resulting in a reduction in cell surface FcεRI and c-Kit expression. Additionally, forced expression of Ehf suppressed FcεRI-mediated degranulation and cytokine production. Ehf inhibited the promoter activity of genes encoding FcεRIα, FcεRIß, and c-Kit by binding to these gene promoters. Furthermore, the mRNA levels of Gata1, Gata2, and Stat5b were lower in BMMCs stably expressing Ehf compared with control cells. Because GATA-1 and GATA-2 are positive regulators of FcεRI and c-Kit expression, decreased expression of GATAs may be also involved in the reduction of FcεRI and c-Kit expression. Decreased expression of Stat5 may contribute to the suppression of cytokine production by BMMCs. In part, mast cell response to TGF-ß1 was mimicked by forced expression of Ehf, suggesting that TGF-ß1 suppresses FcεRI and c-Kit expression and suppresses FcεRI-mediated activation through upregulation of Ehf.

MYB, CD117 and SOX-10 expression in cutaneous adnexal tumors.

BACKGROUND: Elevated MYB expression has been documented in adenoid cystic carcinoma (ACC), cylindroma, and spiradenoma, but the specificity of this finding is unknown. CD117 and SOX-10 expression also occurs in some cutaneous adnexal tumors. This study assesses MYB, CD117 and SOX-10 expression in cutaneous adnexal tumors. METHODS: Retrospective analysis of 184 benign adnexal tumors (140 eccrine/apocrine, 40 follicular and 10 sebaceous), and 30 malignant adnexal tumors was performed with MYB, SOX-10 and CD117 immunostaining. RESULTS: In the benign adnexal tumors, 16% (23/140) significantly expressed MYB. MYB expression was limited to cylindromas and to a lesser extent, spiradenomas in the benign cohort. Elevated MYB expression was detected in mucinous carcinoma, endocrine mucin-producing sweat gland carcinoma and 1 and 4 cases of extramammary Paget's disease (EMPD) in the malignant cohort. CD117 and SOX-10 had similar overall positivity rates in benign apocrine and eccrine tumors (45% and 68% respectively), and were generally negative in other benign and malignant adnexal tumors. CONCLUSION: Expression of MYB appears limited to a small number of cutaneous adnexal tumors, including cylindromas, spiradenomas, ACCs, mucinous carcinoma, endocrine mucin-producing sweat gland carcinoma and some cases of EMPD.

13C-Acetic Acid Breath Test Monitoring of Gastric Emptying during Disease Progression in Diabetic Rats.

Gastric motility disturbance is commonly found in long-standing hyperglycemia. Both delayed and rapid gastric emptying has been reported in diabetes. However, very few studies have followed the changes in gastric emptying during disease progression in diabetes because of technical limitations. 13C-Acetic acid breath test is a validated method which is non-invasive and can be used repeatedly or serially to evaluate gastric emptying changes in animal. We investigated the gastric emptying changes in different stages of diabetes using 13C-acetic acid breath test, as well as its related mechanisms involving interstitial cells of Cajal (ICCs), and stem cell factor (SCF) in streptozotocin-induced diabetic rats. The results showed that gastric emptying was accelerated at the early stage (12 weeks of diabetes) whereas intramuscular ICCs (ICC-IM) networks were not different from normal group. At long-term stage (28 weeks of diabetes), gastric emptying had returned to normal pattern with no delayed. ICC-IM networks were decreased in the diabetic group compared to 12th weeks, and were lower than in the normal group at the same time point. SCF levels were constantly high in the diabetic group than in the normal group. This result indicated that 13C-acetic acid breath test is useful to track the alteration in gastric emptying during disease progression. The change of gastric emptying was not found to be significantly associated with ICC-IM. Elevated SCF may help to preserve ICC-IM, especially in the early phase of diabetes.

Src-like-adaptor protein (SLAP) differentially regulates normal and oncogenic c-Kit signaling.

The Src-like-adaptor protein (SLAP) is an adaptor protein sharing considerable structural homology with Src. SLAP is expressed in a variety of cells and regulates receptor tyrosine kinase signaling by direct association. In this report, we show that SLAP associates with both wild-type and oncogenic c-Kit (c-Kit-D816V). The association involves the SLAP SH2 domain and receptor phosphotyrosine residues different from those mediating Src interaction. Association of SLAP triggers c-Kit ubiquitylation which, in turn, is followed by receptor degradation. Although SLAP depletion potentiates c-Kit downstream signaling by stabilizing the receptor, it remains non-functional in c-Kit-D816V signaling. Ligand-stimulated c-Kit or c-Kit-D816V did not alter membrane localization of SLAP. Interestingly oncogenic c-Kit-D816V, but not wild-type c-Kit, phosphorylates SLAP on residues Y120, Y258 and Y273. Physical interaction between c-Kit-D816V and SLAP is mandatory for the phosphorylation to take place. Although tyrosine-phosphorylated SLAP does not affect c-Kit-D816V signaling, mutation of these tyrosine sites to phenylalanine can restore SLAP activity. Taken together the data demonstrate that SLAP negatively regulates wild-type c-Kit signaling, but not its oncogenic counterpart, indicating a possible mechanism by which the oncogenic c-Kit bypasses the normal cellular negative feedback control.

Mouse Spermatogenesis Requires Classical and Nonclassical Testosterone Signaling.

Testosterone acts though the androgen receptor in Sertoli cells to support germ cell development (spermatogenesis) and male fertility, but the molecular and cellular mechanisms by which testosterone acts are not well understood. Previously, we found that in addition to acting through androgen receptor to directly regulate gene expression (classical testosterone signaling pathway), testosterone acts through a nonclassical pathway via the androgen receptor to rapidly activate kinases that are known to regulate spermatogenesis. In this study, we provide the first evidence that nonclassical testosterone signaling occurs in vivo as the MAP kinase cascade is rapidly activated in Sertoli cells within the testis by increasing testosterone levels in the rat. We find that either classical or nonclassical signaling regulates testosterone-mediated Rhox5 gene expression in Sertoli cells within testis explants. The selective activation of classical or nonclassical signaling pathways in Sertoli cells within testis explants also resulted in the differential activation of the Zbtb16 and c-Kit genes in adjacent spermatogonia germ cells. Delivery of an inhibitor of either pathway to Sertoli cells of mouse testes disrupted the blood-testis barrier that is essential for spermatogenesis. Furthermore, an inhibitor of nonclassical testosterone signaling blocked meiosis in pubertal mice and caused the loss of meiotic and postmeiotic germ cells in adult mouse testes. An inhibitor of the classical pathway caused the premature release of immature germ cells. Collectively, these observations indicate that classical and nonclassical testosterone signaling regulate overlapping and distinct functions that are required for the maintenance of spermatogenesis and male fertility.

Celastrol Induces Cell Apoptosis and Inhibits the Expression of the AML1-ETO/C-KIT Oncoprotein in t(8;21) Leukemia.

Resistance to chemotherapy is a major challenge to improving overall survival in Acute Myeloid Leukemia (AML). Therefore, the development of innovative therapies and the identification of more novel agents for AML are urgently needed. Celastrol, a compound extracted from the Chinese herb Tripterygium wilfordii Hook, exerts anticancer activity. We investigated the effect of celastrol in the t(8;21) AML cell lines Kasumi-1 and SKNO-1. We demonstrated that inhibition of cell proliferation activated caspases and disrupted mitochondrial function. In addition, we found that celastrol downregulated the AML1-ETO fusion protein, therefore downregulating C-KIT kinases and inhibiting AKT, STAT3 and Erk1/2. These findings provide clear evidence that celastrol might provide clinical benefits to patients with t(8;21) leukemia.

Paradoxical and contradictory effects of imatinib in two cell line models of hormone-refractory prostate cancer.

BACKGROUND: Imatinib mesylate is a chemotherapeutic drug that inhibits the tyrosine kinase activity of c-KIT and has been successfully used to treat leukemias and some solid tumors. However, its application for treatment of hormone-refractory prostate cancer (HRPC) has shown modest effectiveness and did not follow the outcomes in cultured cells or animal models. Moreover, the molecular pathways by which imatinib induces cytotoxicity in prostate cancer cells are poorly characterized. METHODS: Two cell line models of HRPC (DU145 and PC3) were exposed to 20 µM of imatinib for 6-72 hr. MTS assay was used to assess cell viability during the course of experiment. Gene expression analysis of c-KIT, cell-cycle and apoptosis regulators, and angiogenic factors was determined by means of real-time PCR, western blot, and/or immunocytochemistry. The enzymatic activity of the apoptosis effector, caspase-3, was determined by a colorimetric assay. RESULTS: Imatinib significantly decreased the viability of DU145 cells but paradoxically augmented the viability of PC3 cells. DU145 cells displayed diminished expression of anti-apoptotic Bcl-2 protein and augmented levels of caspase-8 and -9, as well as, increased enzymatic activity of caspase-3 in response to imatinib. No differences existed on the expression levels of apoptosis-related proteins in PC3 cells treated with imatinib, though the activity of caspase-3 was decreased. The mRNA levels of angiogenic factor VEGF were decreased in DU145-treated cells, whereas an opposite effect was seen in PC3. In addition, it was shown that DU145 and PC3 cells present a differential expression of c-KIT protein variants. CONCLUSION: DU145 and PC3 cells displayed a contradictory behavior in response to imatinib, which was underpinned by a distinct expression pattern (or activity) of target regulators of cell-cycle, apoptosis, and angiogenesis. The paradoxical effect of imatinib in PC3 cells may be related with the differential expression of c-KIT protein variants. Moreover, the present findings helped to understand the discrepancies in the efficacy of imatinib as therapeutic option in HRPC.

High receptor tyrosine kinase (FLT3, KIT) transcript versus anti-apoptotic (BCL2) transcript ratio independently predicts inferior outcome in pediatric acute myeloid leukemia.

OBJECTIVE: In acute myeloid leukemia (AML), simultaneous expression of proliferative (FLT3, KIT) and anti-apoptotic genes (BCL2) is unknown. The aim of the study was to prospectively evaluate proliferative and anti-apoptotic gene transcripts, their interrelationship and impact on the outcome in pediatric AML patients. METHODS: We assessed proliferative and anti-apoptotic gene transcripts by Q-polymerase chain reaction (TaqMan probe) in 64 consecutive pediatric AML patients. Survival data was analyzed by Kaplan-Meier curves followed by log rank test to compare statistical significance between groups. Stepwise multivariable Cox regression method was used to evaluate independent prognostic factors. RESULTS: In univariate analysis, transcript ratio of FLT3/BCL2 and FLT3+KIT/BCL2 significantly predicted event free survival (EFS) (<0.01 and <0.01 respectively) and overall survival (OS) (<0.01 and<0.01 respectively). In stepwise Cox-regression model, high white blood cell count and high FLT3+KIT/BCL2 ratio predicted EFS (HR: 2.2 and 2.3); high hemoglobin and high FLT3+KIT/BCL2 ratio predicted OS (HR: 0.45 and 3.85). Prognostic index (PI) was calculated using the hazard coefficient of independent prognostic factors; at 57.3 months, predicted OS of patients with the highest PI of 1.8 was 8% versus 73% for the lowest PI of -0.3. The mean PI of patients who died was 1.8±0.72 versus 0.54±0.70 for those who are alive, P=0.004. CONCLUSIONS: This first study showed that individual expression of proliferative and anti-apoptotic transcripts is not as important in AML patients, rather their interrelationship and relative level probably determines the outcome.

CD117 expression in breast phyllodes tumors correlates with adverse pathologic parameters and reduced survival.

CD117 (c-kit) is a type III receptor tyrosine kinase encoded by the KIT gene. Deregulation of expression and mutations in the gene are implicated in various tumors. Reports of CD117 expression in phyllodes tumors have been controversial. We aim to investigate the protein expression of CD117 and mutations in the KIT gene in phyllodes tumors, and correlate the findings with pathological parameters and clinical outcome. A total of 272 cases were included in this study. CD117 expression was investigated by immunohistochemistry on tissue microarray sections. Toluidine blue staining was performed to indicate mast cells. Overall, 28 (10%) cases were CD117 positive. CD117 expression was significantly associated with tumor grade (P<0.001), increased stromal hypercellularity (P=0.003), stromal atypia (P=0.01), and stromal mitotic activity (P<0.001), permeative microscopic margins (P=0.002), and presence of hemorrhage (P=0.001). Expression was also associated with poorer overall survival (P=0.003). Nineteen cases were further selected for mutation screening through the Affymetrix OncoScan platform. No mutation of the KIT gene was found. Despite a lack of mutations in the KIT gene, CD117 protein expression is associated with unfavorable pathological parameters and poorer prognosis, suggesting an underlying role in the biology of phyllodes tumors.

Diagnostic procedures for improving of the KIT (CD117) expressed allele burden for the liver metastases from uterus mast cell tumors: prognostic value of the metastatic pattern and tumor biology.

The activating KIT marker plays a central role in the pathogenesis, diagnosis, and targeted treatment of systemic mastocytosis (SM). Recent studies have identified the KIT (CD117) as a marker that distinguishes nonneoplastic from neoplastic mast cells in human systemic mastocytosis. In this study, we conclude that immunohistopathology assays for KIT staining pattern are useful complimentary tools for diagnosis and evaluation of prognosis in uterus mast cell tumor (MCT) metastasis to the liver in 10 patients. Uterine and hepatic cytology revealed mast cell neoplasia, which was confirmed as visceral mast cell tumor on postmortem examination. Histological changes of densely packed, poorly differentiated neoplastic mast cells, sheets of neoplastic round to pleomorphic cells that formed nonencapsulated nodules, high mitotic figures, necrosis, and fibrosis were found. In addition, eosinophils were scattered among the mast cells at the periphery of the nodules. These findings indicate tumors of high-grade malignancy with infiltrative cells resembling the uterus MCT in the intraparenchymal and periparenchymal areas of the liver. Immunohistochemically, tumors were positive for KIT. The histopathologic features coupled with the KIT immunoreactivity led to diagnosis of high-grade uterus MCTs. Taken together, these findings suggest that CD117 may play a critical role in early uterus MCT development and may be a stimulatory factor in grade 3 MCT. Therefore, the result has supported our hypothesis that there was an increased opportunity to observe a higher CD117 staining pattern in high-grade MCTs.

Acquired resistance to EGFR tyrosine kinase inhibitor in A431 squamous cell carcinoma xenografts is mediated by c-Kit pathway transduction.

Epidermal growth factor inhibitors (EGFRIs), the first targeted cancer therapy, are currently an essential treatment for many advance-stage epithelial cancers. These agents have the superior ability to target cancers cells and better safety profile compared to conventional chemotherapies. However, all responding patients eventually developed acquired resistance to EGFRIs and the mechanisms of acquired resistance invariably develops. In the current study, we reported the tumor xenografts of the human A431 squamous cell carcinoma, after 25-week consecutive therapy with EGFR inhibitor (gefitinib) that developed resistance as a result of c-Kit overexpression. Moreover, combined therapeutic inhibition of EGFR and c-Kit may abrogate this acquired mechanism of drug resistance due to an enhanced apoptotic effect in gefitinib-resistant xenograft model. Taken together, the results suggest that at least in the A431 xenograft model displaying acquired resistance to gefitinib can emerge in vivo, at least in part, by mechanisms involving the c-Kit overexpression.