Circulating innate immune markers and outcomes in treatment-naïve advanced non-small cell lung cancer patients.

INTRODUCTION: Innate immunity represents the first step of activation of the immune system and dictates the quality of adaptive immune responses. Studies have reported links between systemic inflammatory or innate immune markers and prognosis in patients with lung cancer. To our knowledge, the prospective and concomitant study of these systemic markers has never been performed. METHODS: Advanced treatment-naive non-small cell lung cancer (NSCLC) patients eligible for first-line platinum-based chemotherapy were prospectively included from December 2012 to July 2015 (N = 148). Blood samples of patients were collected before the first cycle for fresh NK cell phenotyping. Peripheral blood mononuclear cells were cryopreserved for natural cytotoxicity receptor (NCR) genotyping as well as sera for NCR's ligand quantification. Data on leukocytes, neutrophils and monocyte counts and lactate dehydrogenase (LDH) levels were extracted from electronic medical records. RESULTS: Among all studied markers, monocytosis, neutrophilia, leucocytosis, high LDH and sBAG6 levels and reduced levels of NCR3 transcripts were associated with poor overall survival (OS) in univariate analysis. The levels of NCR3 transcripts was linked to age, number of metastatic sites, monocyte counts, LDH and sBAG6 levels. Neutrophilia was associated to high sBAG6 levels. NCR3 was the unique innate immune parameter that remained as an independent factor associated with both OS (P = 0.003) and progression-free survival (P = 0.009) in the multivariate analysis. CONCLUSION: This study brought evidence that these biomarkers are entangled; parameters associated with an inflammatory process were related to reduced levels of NCR3 transcripts. Finally, the level of NCR3 transcripts was independently associated with outcomes in treatment-naive patients with advanced NSCLC.

Development of anti-CD30 radioimmunoconstructs (RICs) for treatment of Hodgkin's lymphoma. Studies with cell lines and animal studies.

OBJECTIVES: Comparison of the binding affinity to a CD30-positive Hodgkin lymphoma (HL) cell line and biodistribution in HL bearing mice of new anti-CD30 radioimmunoconjugates (RICs) of varying structure and labelling nuclides. METHODS: The antibodies Ki-4 and 5F11 were radioiodinated by the chloramine T method or labelled with (111)In via p-NCS-Benzyl-DOTA. In addition, the Ki-4-dimer was investigated in the iodinated form. The RICs were analyzed for retained immunoreactivity by immunochromatography. In-vitro binding studies were performed on CD30-positive L540 cell lines. For in-vivo biodistribution studies, SCID mice bearing human HL xenografts were injected with the various radioimmunoconjugates. After 24 h, activities in the organs and tumour were measured for all 5 RICs. Tumour-free animals were studied in the same way with (131)I- Ki-4 24 h p. i. The three RICs with the highest tumour/background ratios 24 h p.i. ((131)I-Ki-4, (131)I-5F11, (111)In-bz-DOTA-Ki-4) were analysed further at 48 h and 72 h. RESULTS: All the RICs were successfully labelled with high specific activities (28-47 TBq/mmol) and sufficient radiochemical yields (>80%). Scatchard plot analysis proved high tumour affinity (KD = 20-220 nmol/l). In-vivo tumour accumulation in % of injected dose per g tissue (%ID/g) lay between 2.6 ((131)I-5F11) and 12.3 % ID/g ((131)I-Ki-4) with permanently high background in blood. Tumour/blood-ratios of all RICs were below one at all time points. CONCLUSIONS: In-vitro tumour cell affinities of all RICs were promising. However, in-vivo biokinetics tested in the mouse model did not meet expectations. This highlights the importance of developing and testing further new anti-CD30 conjugates.

ERH (enhancer of rudimentary homologue), a conserved factor identical between frog and human, is a transcriptional repressor.

Drosophila enhancer of rudimentary [e(r)] interacts genetically with the rudimentary gene, which encodes a protein possessing the first three enzymatic activities of the pyrimidine biosynthesis pathway. A regulatory or enzymatic activity of e(r) in pyrimidine biosynthesis and the cell cycle has been suggested, but nothing is known about its molecular function. The factor is evolutionarily highly conserved since homologues exist in plants and mammals. We cloned the Xenopus enhancer of rudimentary homologue (XERH) as an interaction partner of DCoH/PCD (dimerisation cofactor of HNF1/pterin-4alpha-carbinolamine dehydratase) in the yeast two-hybrid assay. DCoH/PCD is a multifunctional factor originally identified as a positive cofactor of the HNF1 homeobox transcription factors. XERH is a 104 amino acid protein that is identical to its mammalian homologues. The mRNA is expressed maternally, enriched in ectodermal derivatives during development and ubiquitously detectable in the adult. Fused to the DNA binding region of the GAL4 transcription factor domain, XERH represses the activity of a GAL4 responsive reporter in HeLa, but not in NIH3T3 cells. Furthermore, the DCoH/PCD coactivation of a HNF1 responsive reporter is inhibited by XERH. We propose that XERH is a cell type-specific transcriptional repressor, probably interfering with HNF1-dependent gene regulation via DCoH/PCD.

The mutated human gene encoding hepatocyte nuclear factor 1beta inhibits kidney formation in developing Xenopus embryos.

The transcription factor hepatocyte nuclear factor 1beta (HNF1beta) is a tissue-specific regulator that also plays an essential role in early development of vertebrates. In humans, four heterozygous mutations in the HNF1beta gene have been identified that lead to early onset of diabetes and severe primary renal defects. The degree and type of renal defects seem to depend on the specific mutation. We show that the frameshift mutant P328L329fsdelCCTCT associated with nephron agenesis retains its DNA-binding properties and acts as a gain-of-function mutation with increased transactivation potential in transfection experiments. Expression of this mutated factor in the Xenopus embryo leads to defective development and agenesis of the pronephros, the first kidney form of amphibians. Very similar defects are generated by overexpressing in Xenopus the wild-type HNF1beta, which is consistent with the gain-of-function property of the mutant. In contrast, introduction of the human HNF1beta mutant R137-K161del, which is associated with a reduced number of nephrons with hypertrophy of the remaining ones and which has an impaired DNA binding, shows only a minor effect on pronephros development in Xenopus. Thus, the overexpression of both human mutants has a different effect on renal development in Xenopus, reflecting the variation in renal phenotype seen with these mutations. We conclude that mutations in human HNF1beta can be functionally characterized in Xenopus. Our findings imply that HNF1beta not only is an early marker of kidney development but also is functionally involved in morphogenetic events, and these processes can be investigated in lower vertebrates.

The embryonic expression of the tissue-specific transcription factor HNF1alpha in Xenopus: rapid activation by HNF4 and delayed induction by mesoderm inducers.

The tissue-specific transcription factor HNF1alpha is expressed in kidney, liver, intestine and stomach of Xenopus. We show that the HNF1alpha gene is transcriptionally activated at the onset of zygotic gene transcription and that this transcription is maintained throughout development. Ectodermal explants of blastulae (animal caps) express HNF1alpha mRNA upon stimulation with the mesoderm inducers activin A and BMP4 as well as on overexpression of Smad2 and Smad1, the corresponding members of the intracellular TGF-beta signal transducers, respectively. Beside these factors that mediate their response through serine/threonine kinase receptors, bFGF, which acts via tyrosine kinase receptors, leads to HNF1alpha expression, too. These embryonic inducers result in a delayed appearance of HNF1alpha mRNA, excluding a direct activation of HNF1alpha. In contrast, the maternally expressed nuclear receptors HNF4alpha and HNF4beta activate the initial HNF1alpha transcription, since overexpression of HNF4 leads to a rapid expression of HNF1alpha mRNA in animal caps. Similarly, in entire neurulae HNF4 overexpression results in increased HNF1alpha transcription. Therefore, we assume that the initial activation is dependent on maternal HNF4alpha and HNF4beta transcription factors whereas HNF1alpha induction by growth factors reflects the property of these factors to induce the differentiation of mesodermal and entodermal cell types expressing HNF1alpha.