Assessment of chimerism in epithelial cancers in transplanted patients.

Cancer is now the most severe complication in the long term in transplant recipients. As most solid-organ or hematopoietic stem-cell transplantations are allogeneic, chimerism studies can be performed on cancers occurring in recipients. We summarize here the different methods used to study chimerism in cancers developing in allogeneic-transplant recipients, analyze their respective advantages and report the main results obtained from these studies. Chimerism analyses of cancers in transplant recipients require methods suited to tissue samples. In the case of gender-mismatched transplantation, the XY chromosomes can be explored using fluorescent in situ hybridization on whole-tissue sections or Y-sequence-specific PCR after the laser microdissection of tumor cells. For cancers occurring after gender-matched transplantation, laser microdissection of tumor cells enables studies of microsatellite markers and high-resolution melting analysis of mitochondrial DNA on genes with marked polymorphism, provided these are different in the donor and the recipient. The results of different studies address the cancers that develop in both recipients and in transplants. The presence of chimeric cells in these two types of cancer implies an exchange of progenitor/stem-cells between transplant and recipient, and the plasticity of these progenitor/stem-cells contributes to epithelial cancers. The presence of chimeric cells in concomitant cancers and preneoplastic lesions implies that the oncogenesis of these cancers progresses through a multistep process.

Erratum: Cuceu, C., et al. Independent Mechanisms Lead to Genomic Instability in Hodgkin Lymphoma: Microsatellite or Chromosomal Instability. Cancers 2018, 10, 233.

The authors wish to make the following corrections to this paper [...].

Independent Mechanisms Lead to Genomic Instability in Hodgkin Lymphoma: Microsatellite or Chromosomal Instability †.

Background: Microsatellite and chromosomal instability have been investigated in Hodgkin lymphoma (HL). Materials and Methods: We studied seven HL cell lines (five Nodular Sclerosis (NS) and two Mixed Cellularity (MC)) and patient peripheral blood lymphocytes (100 NS-HL and 23 MC-HL). Microsatellite instability (MSI) was assessed by PCR. Chromosomal instability and telomere dysfunction were investigated by FISH. DNA repair mechanisms were studied by transcriptomic and molecular approaches. Results: In the cell lines, we observed high MSI in L428 (4/5), KMH2, and HDLM2 (3/5), low MSI in L540, L591, and SUP-HD1, and none in L1236. NS-HL cell lines showed telomere shortening, associated with alterations of nuclear shape. Small cells were characterized by telomere loss and deletion, leading to chromosomal fusion, large nucleoplasmic bridges, and breakage/fusion/bridge (B/F/B) cycles, leading to chromosomal instability. The MC-HL cell lines showed substantial heterogeneity of telomere length. Intrachromosmal double strand breaks induced dicentric chromosome formation, high levels of micronucleus formation, and small nucleoplasmic bridges. B/F/B cycles induced complex chromosomal rearrangements. We observed a similar pattern in circulating lymphocytes of NS-HL and MC-HL patients. Transcriptome analysis confirmed the differences in the DNA repair pathways between the NS and MC cell lines. In addition, the NS-HL cell lines were radiosensitive and the MC-cell lines resistant to apoptosis after radiation exposure. Conclusions: In mononuclear NS-HL cells, loss of telomere integrity may present the first step in the ongoing process of chromosomal instability. Here, we identified, MSI as an additional mechanism for genomic instability in HL.

Inhibition of the proprotein convertases represses the invasiveness of human primary melanoma cells with altered p53, CDKN2A and N-Ras genes.

BACKGROUND: Altered tumor suppressor p53 and/or CDKN2A as well as Ras genes are frequently found in primary and metastatic melanomas. These alterations were found to be responsible for acquisition of invasive and metastatic potential through their defective regulatory control of metalloproteinases and urokinase genes. METHODOLOGY/PRINCIPAL FINDINGS: Using primary human melanoma M10 cells with altered p53, CDKN2A and N-Ras genes, we found that inhibition of the proprotein convertases (PCs), enzymes involved in the proteolytic activation of various cancer-related protein precursors resulted in significantly reduced invasiveness. Analysis of M10 cells and their gastric and lymph node derived metastatic cells revealed the presence of all the PCs found in the secretory pathway. Expression of the general PCs inhibitor alpha1-PDX in these cells in a stable manner (M10/PDX) had no effect on the mRNA expression levels of these PCs. Whereas, in vitro digestion assays and cell transfection experiments, revealed that M10/PDX cells display reduced PCs activity and are unable to process the PCs substrates proIGF-1R and proPDGF-A. These cells showed reduced migration and invasion that paralleled decreased gelatinase MMP-2 activity and increased expression and secretion of tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2. Furthermore, these cells showed decreased levels of urokinase-type plasminogen activator receptor (uPAR) and increased levels of plasminogen activator inhibitor-1 (PAI-1). CONCLUSIONS: Taken together, these data suggest that inhibition of PCs activity results in decreased invasiveness of primary human melanoma cells despite their altered p53, CDKN2A and N-Ras genes, suggesting that PCs may serve as novel therapeutic targets in melanoma.

Highly sensitive multivariable assay detection of melanocytic differentiation antigens and angiogenesis biomarkers in sentinel lymph nodes with melanoma micrometastases.

OBJECTIVES: To evaluate the prognostic value of melanocytic differentiation antigens and angiogenesis biomarkers in sentinel lymph nodes (SLNs) with melanoma micrometastases. DESIGN: Prognostic study of an inception cohort. SETTING: Academic research. Patients Between July 1, 1999, and July 31, 2002, all patients who had primary cutaneous or mucosal melanomas that have a Breslow depth of 1.5 mm or greater, ulceration, or Clark level IV or V, or had SLN biopsies. MAIN OUTCOME MEASURES: By the use of quantitative reverse transcription-polymerase chain reaction, the expression of the following was analyzed in SLNs: 2 melanocytic differentiation antigens (tyrosinase [P17646] and melanoma antigen recognized by T cells [MART-1; Q16655]) and genes involved in angiogenesis (VEGF [NM_001025366] and VEGFR2 [AF035121]), lymphangiogenesis (VEGFC [NM_005429], VEGFR3 [X68203], LYVE1 [NM_016164], and PROX1 [002763]), and invasion (uPA [NM_002658], PAI1 [NM_00602], and EMMPRIN [L10240]). Outcome measures were the association of these melanocytic differentiation antigens and angiogenesis biomarkers with clinicopathologic characteristics of patients, and an evaluation of the prognostic value for relapse-free survival and overall survival. RESULTS: Ninety-one patients were included, with a median follow-up period of 41 months. Micrometastases were present in 15% (14 of 91) of patients. Tyrosinase (P < .001), MART-1 (P < .001), vascular endothelial growth factor 121 (VEGF(121)) (P = .007), and PAI1 (P = .02) expression was significantly associated with micrometastasis. In univariate analysis, histologic findings and tyrosinase and MART-1 expression were significantly associated with relapse-free survival. Tyrosinase and MART-1 expression was associated with overall survival. A multiple Cox proportional hazards regression model identified negative histologic findings and tyrosinase expression that exceeded 27 copies/copy of TATA box-binding protein (third quartile) as significantly associated with an increased risk of relapse or death. CONCLUSIONS: Quantitative assessment of melanocytic differentiation antigens in SLNs, which has prognostic value, is more specific than qualitative assessment. Prognosis may be more effectively predicted by the combination of quantitative assessment of melanocytic differentiation antigens in SLNs with histologic assessment. A significant association was found between the presence of micrometastases and the expression of angiogenesis biomarkers.

[Analysis of overexpression of vascular endothelial growth factor-C in patients with angioimmunoblastic T-cell lymphoma].

OBJECTIVE: To explore the vascular endothelial growth factor-C (VEGF-C) expression and its clinical significance in malignant lymphoma. METHODS: Lymphoma cells were isolated by laser microdissection. VEGF-C expression in lymphoma tissue and microdissected lymphoma cells was measured by realtime quantitative PCR. Meanwhile, vessel ultrastructure was identified by transmission electron microscopy. RESULTS: Comparing with that in 8 patients with reactive lymphocyte hyperplasia, VEGF-C was overexpressed in angioimmunoblastic T-cell lymphoma, both in lymphoma tissue (n = 18, P = 0.0020) and in microdissected lymphoma cells (n = 10, P < 0.0001). Increased VEGF-C level was associated with bone marrow infiltration (P = 0.0039), skin involvement (P = 0.0046) and high-risk international prognostic index (P = 0.0302). In VEGF-C overexpressed cases, ultrastructural study showed dystrophic vessels, with swelling endothelial cells and absence of pericytes. CONCLUSION: The value of VEGF-C expression might be a biomarker of disease progression in angioimmunoblastic T-cell lymphoma.

Estudios bioquímicos y caracterización molecular de pacientes portadores de ß talasemia en Córdoba, Argentina / Biochemical studies and molecular characterization of ß-thalassemia carriers in Córdoba, Argentina

Las ß talasemias constituyen un grupo heterogéneo de anemias microcíticas, causadas por una producción reducida (ß+) o ausencia (ߺ) de síntesis de cadenas de globina ß. La diferente expresividad clínica de estas patologías resulta de la combinación de ambas posibilidades o de cada una de ellas con el gen normal. Dado que este país ha recibido distintas corrientes migratorias, y que no son pocos los portadores ß talasémicos, se intentó identificar por primera vez qué tipo de mutaciones son las más frecuentes en este medio. Para realizarlo, se contó con doce pacientes provenientes de la ciudad de Córdoba que fueron evaluados bioquímicamente con: hematocrito, hemoglobina, porcentaje de reticulocitos, electroforesis de hemoglobinas a pH alcalino, hierro sérico, transferrina, hemoglobina fetal y morfología de los glóbulos rojos. La obtención de un volumen corpuscular medio inferior a 75 fl como así también cantidades superiores de Hb A2 a 2 por ciento fueron los criterios más importantes para la selección de los pacientes. El uso de PCR con uno de los oligonucleótidos de cada set que posee un segmento adicional rico en GC y una posterior electroforesis en un gel desnaturalizante (DGGE), permitió efectuar la identificación de mutaciones puntuales en el gen de la globulina ß. De esta manera se obtuvieron los siguientes resultados: 41,7 por ciento ߺ talasemia (codón 39 C -> T), 50,0 por ciento ß+ talasemia (IVS1 nucleótico 110 G -> A) y 8,3 por ciento ߺ talasemia (IVS1 nucleótico 1 G ->A)