In recent years, phenotypic-based screens have become increasingly popular in drug discovery. A major challenge of this approach is that it does not provide information about the mechanism of action of the hits. This has led to the development of multiple strategies for target deconvolution. Thermal proteome profiling (TPP) allows for an unbiased search of drug targets and can be applied in living cells without requiring compound labeling. TPP is based on the principle that proteins become more resistant to heat-induced unfolding when complexed with a ligand, e.g., the hit compound from a phenotypic screen. The melting proteome is also sensitive to other intracellular events, such as levels of metabolites, post-translational modifications and protein-protein interactions. In this review, we describe the principles of this approach, review the method and its developments, and discuss its current and future applications. While proteomics has generally focused on measuring relative protein concentrations, TPP provides a novel approach to gather complementary information on protein stability not present in expression datasets. Therefore, this strategy has great potential not only for drug discovery, but also for answering fundamental biological questions.

Spontaneous preterm birth (SPTB) is a major factor associating with deaths and with lowered quality of life in humans. Environmental and genetic factors influence the susceptibility. Previously, by analyzing families with recurrent SPTB in linkage analysis, we identified a linkage peak close to the gene encoding CXCR3. Present objectives were to investigate the association of CXCR3 with SPTB in Finnish mothers (n = 443) and infants (n = 747), to analyze CXCR3 expression levels in human placenta and levels of its ligands in umbilical cord blood, and to verify the influence of Cxcr3 on SPTB-associating cytokines in mice. We detected an association between an intronic CXCR3 polymorphism, rs2280964, and SPTB in infants from families with recurrent preterm births (p = 0.009 versus term controls, odds ratio 0.52, 95% confidence interval 0.32-0.86). The minor allele was protective and undertransmitted to SPTB infants (p = 0.007). In the placenta and fetal membranes, the rs2280964 major allele homozygotes had higher expression levels than minor allele homozygotes; decidual trophoblasts showed strong CXCR3 immunoreactivity. Expression was higher in SPTB placentas compared with those from elective deliveries. Concentration of a CXCR3 ligand, CXCL9, was increased in cord blood from SPTB, and the protective rs2280964 allele was associated with low CXCL9. In CXCR3-deficient mice (Mus musculus), SPTB-associating cytokines were not acutely increased in amniotic fluid after preterm birth-inducing dose of maternal LPS. Our results indicate that CXCR3 contributes to SPTB. Activation of CXCR3 signaling may disturb the maternal-fetal tolerance, and this may promote labor.

Understanding of timing of human parturition is incomplete. Therefore, we carried out proteomic analyses of full‐term placentas from uncomplicated pregnancies to identify protein signatures associated with the onset of spontaneous delivery. We found quantitative associations of 10 proteins with spontaneous term birth, evident either in the basal or in the chorionic plates or in both. Additional 18 proteins were associated according to the location within placenta indicating local variations in protein amounts. Calcineurin‐like phosphoesterase domain‐containing 1 (CPPED1), a phosphatase previously suggested dephosphorylating AKT1/PKB, was one of the identified proteins. qRT‐PCR revealed the mRNA level of CPPED1 was higher in elective caesarean deliveries than in spontaneous births, while immunohistochemistry showed CPPED1 in cytotrophoblasts, syncytiotrophoblasts and extravillous trophoblasts. Noteworthy, phosphorylation status of AKT1 did not differ between placentas from elective caesarean and spontaneous deliveries. Additionally, analyses of samples from infants indicated that single‐nucleotide polymorphisms rs11643593 and rs8048866 of CPPED1 were associated with duration of term pregnancy. Finally, post‐transcriptional silencing of CPPED1 in cultured HTR8/SVneo cells by siRNAs affected gene expression in pathways associated with inflammation and blood vessel development. We postulate that functions regulated by CPPED1 in trophoblasts at choriodecidual interphase have a role in the induction of term labour, but it may be independent of AKT1.