Construction of novel immune-related signature for prediction of pathological complete response to neoadjuvant chemotherapy in human breast cancer.

BACKGROUND: The aim of this study was to construct a novel prediction model for the pathological complete response (pCR) to neoadjuvant chemotherapy (NAC) using immune-related gene expression data. PATIENTS AND METHODS: DNA microarray data were used to perform a gene expression analysis of tumor samples obtained before NAC from 117 primary breast cancer patients. The samples were randomly divided into the training (n = 58) and the internal validation (n = 59) sets that were used to construct the prediction model for pCR. The model was further validated using an external validation set consisting of 901 patients treated with NAC from six public datasets. RESULTS: The training set was used to construct an immune-related 23-gene signature for NAC (IRSN-23) that is capable of classifying the patients as either genomically predicted responders (Gp-R) or non-responders (Gp-NR). IRSN-23 was first validated using an internal validation set, and the results showed that the pCR rate for Gp-R was significantly higher than that obtained for Gp-NR (38 versus 0%, P = 1.04E-04). The model was then tested using an external validation set, and this analysis showed that the pCR rate for Gp-R was also significantly higher (40 versus 11%, P = 4.98E-23). IRSN-23 predicted pCR regardless of the intrinsic subtypes (PAM50) and chemotherapeutic regimens, and a multivariate analysis showed that IRSN-23 was the most important predictor of pCR (odds ratio = 4.6; 95% confidence interval = 2.7-7.7; P = 8.25E-09). CONCLUSION: The novel prediction model (IRSN-23) constructed with immune-related genes can predict pCR independently of the intrinsic subtypes and chemotherapeutic regimens.

Isolation and characterization of genomic and cDNA clones encoding mouse senescence marker protein-30 (SMP30).

We isolated and characterized genomic and cDNA clones encoding mouse senescence marker protein-30 (SMP30), the protein amounts of which are known to decrease with aging in the livers of rats. This decrease in the expression of SMP30 is independent of androgen. SMP30 is a calcium binding protein also called regucalcin. The expression of SMP30 in aged mouse liver and 5' flanking sequence of the genome were also characterized. The cDNA contained an open reading frame encoding 299 amino acids with a calculated molecular weight of 33-404. The amino acid sequence of mouse SMP30 showed 94% similarity to rat SMP30 and 89% to human SMP30. Northern blot analysis specifically detected mouse SMP30 transcript in the liver and also confirmed its significant decrease with aging. Analysis of the murine genomic clone revealed that SMP30 was organized by seven exons and six introns, spanning approx. 17.5 kb. Primer extension analysis revealed that two major transcription initiation sites were localized at 101 bp and 102 bp upstream from ATG translation initiation codon. The nucleotide (nt) sequence of 5' flanking region showed a TATA-like sequence, a CAAT box, and SP-1 sites at nt -29, -72 and -169 in the promoter region, respectively. Interestingly, we found two classes of C/EBP sites which are highly and constantly expressed in the liver, in addition to AP-2, AP-1, GATA-1, AP-1/GRE and GAGA sites. These results provide important clues for understanding the regulatory mechanism of SMP30 gene expression and its relationship to aging.

Isolation of cDNA clone encoding human homologue of senescence marker protein-30 (SMP30) and its location on the X chromosome.

We have isolated and characterized a cDNA clone encoding human homologue of senescence marker protein-30 (SMP30), a calcium binding protein also called regucalcin (RC). This clone (pHSMP6) has 1356 base pairs (bp) and contains an open reading frame of 897 bp, which encodes 299 amino acids. The estimated molecular weight of the deduced polypeptide is 33,250 and pI is 5.836. The homology of amino acid sequences between human homologue and rat SMP30 is 88.6%. Using pHSMP6 as a probe, the chromosomal location of the human homologue of SMP30 gene was determined. The results of regional mapping using a panel of 11 rodent-human somatic hybrids indicated that the gene is located in the p11.3-q11.2 segment of the X chromosome. This gene thus could be a candidate for one of the X-linked diseases mapped to this regions.

Protein tyrosine kinases expressed in glomeruli and cultured glomerular cells: Flt-1 and VEGF expression in renal mesangial cells.

Protein tyrosine kinases play an important role in cellular proliferation and differentiation of various cell types. To identify potential tyrosine kinases involved in glomerular functions we have utilized the polymerase chain reaction (PCR) and degenerate oligonucleotides for isolation of such genes from isolated glomeruli, cultured mesangial cell, and glomerular endothelial cells. Sequence analysis of PCR-amplified cDNAs resulted in the isolation of 24 tyrosine kinases. Here we describe for the first time the constitutive expression of 15 tyrosine kinases, tyro-1, tyro-4, tyro-6, hyk, Ptk-3, Ryk, tie, yes, lyn, tec, Jak1, Jak2, Jak3, c-abl, and flk, in renal glomeruli. In addition, Flt-1, an endothelial cell-specific receptor for vascular endothelial growth factor (VEGF), is expressed in renal mesangial cells and its gene expression is up-regulated upon the stimulation of platelet-derived growth factor (PDGF) with the concomitant up-regulation of VEGF. These data suggest that possible involvement of VEGF/Flt-1 system in cytokine-induced mesangial cell proliferations.