Preclinical Evaluation of JAB-2485, a Potent AURKA Inhibitor with High Selectivity and Favorable Pharmacokinetic Properties.

As a critical mitotic regulator, Aurora kinase A (AURKA) is aberrantly activated in a wide range of cancers. Therapeutic targeting of AUKRA is a promising strategy for the treatment of solid tumors. In this study, we evaluated the preclinical characteristics of JAB-2485, a small-molecule inhibitor of AURKA currently in Phase I/IIa clinical trial in the US (NCT05490472). Biochemical studies demonstrated that JAB-2485 is potent and highly selective on AURKA, with subnanomolar IC50 and around 1500-fold selectivity over AURKB or AURKC. In addition, JAB-2485 exhibited favorable pharmacokinetic properties featured by low clearance and good bioavailability, strong dose-response relationship, as well as low risk for hematotoxicity and off-target liability. As a single agent, JAB-2485 effectively induced G2/M cell cycle arrest and apoptosis and inhibited the proliferation of small cell lung cancer, triple-negative breast cancer, and neuroblastoma cells. Furthermore, JAB-2485 exhibited robust in vivo antitumor activity both as monotherapy and in combination with chemotherapies or the bromodomain inhibitor JAB-8263 in xenograft models of various cancer types. Together, these encouraging preclinical data provide a strong basis for safety and efficacy evaluations of JAB-2485 in the clinical setting.

A novel tuberculosis antigen identified from human tuberculosis granulomas.

Tuberculosis is a global infectious disease caused by Mycobacterium tuberculosis (Mtb). Although novel Mtb biomarkers from both the pathogen and host have been studied, more breakthroughs are still needed to meet different clinic requirements. In an effort to identify Mtb antigens, chaperone-peptide complexes were purified from TB infected lungs using free-solution isoelectric focusing combined with high resolution LTQ Orbitrap Velos mass spectrometry. Antigen specific cellular immune responses in vitro were then examined. Those efforts led to the identification of six Mtb peptides only identified in Tuberculosis lung samples and that were not found in the control samples. Additionally, antigen-specific IFN-γ secretion, T-cell proliferation, cytokine expression, and a cytotoxic assay were also evaluated. Among the peptides isolated, we identified a 34 amino acid peptide named PKAp belonging to a serine/threonine-protein kinase, as being able to generate Mtb-specific cellular immune responses as noted by elevated antigen-specific cytokine secretion levels, increased CD8(+) T-cell proliferation and a strong cytotoxic lymphocyte (CTL) response. Moreover, the immune stimulating abilities of PKAp were further validated in vivo, with target peptide immunized mice showing an increased cellular IFN-γ in both the lungs and spleen without causing immunopathogenesis. In conclusion, we identified novel functional Mtb antigens directly from the granulomatous lesions of Tuberculosis patients, inducing not only significant antigen-specific IFN-γ secretion but also a marked cytotoxic lymphocyte functional response. These findings indicated that PKAp has potential as a novel antigen biomarker for vaccine development.

Growth factor receptor bound protein 2-associated binder 2, a scaffolding adaptor protein, negatively regulates host immunity against tuberculosis.

Cell-mediated immunity is indispensable for host protection against tuberculosis (TB). Growth factor receptor bound protein 2-associated binder (Gab) 2, a scaffolding adaptor protein, negatively regulates signaling pathways critical for T cell-mediated immunity. We sought to investigate the clinical significance and immunological role of Gab2 in Mycobacterium tuberculosis infection. We evaluated Gab2 protein and messenger RNA (mRNA) expression in human patients with pulmonary TB and determined the correlation of the mRNA expression pattern with antigen-specific IFN-γ secretion. Subsequently, we carried out M. tuberculosis infection in Gab2-deficient and wild-type control mice to explore the immunological role of Gab2 by examining bacterial load, histological changes, cytokine secretion, and gene expression of immune-associated transcription factors. mRNA levels of Gab2 and its correlated family member, Gab1, were markedly decreased in untreated patients with pulmonary TB compared with healthy control subjects. Importantly, this decreased Gab2 expression to normal levels after bacterial load in the patient's sputum became undetectable under the standard anti-TB treatment, which negatively correlated with the level of M. tuberculosis antigen-specific IFN-γ secretion. In the M. tuberculosis infection mouse model, infected Gab2-deficient mice exhibited decreased bacterial load and milder lung pathological damage compared with infected wild-type mice, accompanied by decreased production of IL-2, IL-6, and granulocyte/macrophage colony-stimulating factor proinflammatory cytokines, and an increased T-cell-specific T-box transcription factor/GATA binding protein 3 expression ratio. Overall, our study indicates that down-regulation of Gab2 relates to a protective function during M. tuberculosis infection, revealing a potential negative regulatory role for Gab2 in immunity to TB.

Different patterns of cytokines and chemokines combined with IFN-γ production reflect Mycobacterium tuberculosis infection and disease.

BACKGROUND: IFN-γ is presently the only soluble immunological marker used to help diagnose latent Mycobacterium tuberculosis (M.tb) infection. However, IFN-γ is not available to distinguish latent from active TB infection. Moreover, extrapulmonary tuberculosis, such as tuberculous pleurisy, cannot be properly diagnosed by IFN-γ release assay. As a result, other disease- or infection-related immunological biomarkers that would be more effective need to be screened and identified. METHODOLOGY: A panel of 41 soluble immunological molecules (17 cytokines and 24 chemokines) was tested using Luminex liquid array-based multiplexed immunoassays. Samples, including plasma and pleural effusions, from healthy donors (HD, n = 12) or patients with latent tuberculosis infection (LTBI, n = 20), pulmonary tuberculosis (TB, n = 12), tuberculous pleurisy (TP, n = 15) or lung cancer (LC, n = 15) were collected and screened for soluble markers. Peripheral blood mononuclear cells (PBMCs) and pleural fluid mononuclear cells (PFMCs) were also isolated to investigate antigen-specific immune factors. PRINCIPAL FINDINGS: For the 41 examined factors, our results indicated that three patterns were closely associated with infection and disease. (1) Significantly elevated plasma levels of IL-2, IP-10, CXCL11 and CXCL12 were present in both patients with tuberculosis and in a sub-group participant with latent tuberculosis infection who showed a higher level of IFN-γ producing cells by ELISPOT assay compared with other latently infected individuals. (2) IL-6 and IL-9 were only significantly increased in plasma from active TB patients, and the two factors were consistently highly secreted after M.tb antigen stimulation. (3) When patients developed tuberculous pleurisy, CCL1, CCL21 and IL-6 were specifically increased in the pleural effusions. In particular, these three factors were consistently highly secreted by pleural fluid mononuclear cells following M.tb-specific antigen stimulation. In conclusion, our data imply that the specific secretion of soluble immunological factors, in addition to IFN-γ, may be used to evaluate M.tb infection and tuberculosis disease.

Increased levels of BAFF and APRIL related to human active pulmonary tuberculosis.

BACKGROUND: Despite great efforts to improve diagnosis and treatment, tuberculosis (TB) remains a major health problem worldwide, especially in developing countries. Lack of concrete immune markers is still the obstacle to properly evaluate active TB. Therefore, identification of more validated biomarkers and phenotypic signatures is imperative. In particular, T cell-related biomarkers are more significant. METHODOLOGY: To understand the nature of CD4(+) T cell-derived signatures involved in infection and disease development, we examined and analyzed whole genome expression profiles of purified CD4(+) T cells from healthy individuals (HD), two distinct populations with latent infection (with low or high IFN-γ levels, LTB(L)/LTB(H)) and untreated TB patients. Following, we validated the expression profiles of genes in the peripheral CD4(+) T cells from each group and examined secretion levels of distinct cytokines in serum and pleural effusion. PRINCIPAL FINDINGS: Our bio-informatic analyses indicate that the two latent populations and clinical TB patients possess distinct CD4(+) T cell gene expression profiles. Furthermore, The mRNA and protein expression levels of B cell activating factor (BAFF), which belongs to the TNF family, and a proliferation-inducing ligand (APRIL) were markedly up-regulated at the disease stage. In particular, the dramatic enhancement of BAFF and APRIL in the pleural effusion of patients with tuberculosis pleurisy suggests that these proteins may present disease status. In addition, we found that the BAFF/APRIL system was closely related to the Th1 immune response. Our study delineates previously unreported roles of BAFF and APRIL in the development of tuberculosis, and these findings have implications for the diagnosis of the disease. Our study also identifies a number of transcriptional signatures in CD4(+) T cells that have the potential to be utilized as diagnostic and prognostic tools to combat the tuberculosis epidemic.