An open-label, pilot study of veliparib and lapatinib in patients with metastatic, triple-negative breast cancer.

BACKGROUND: Poly (ADP-ribose)-polymerase inhibitors (PARPi) have been approved for cancer patients with germline BRCA1/2 (gBRCA1/2) mutations, and efforts to expand the utility of PARPi beyond BRCA1/2 are ongoing. In preclinical models of triple-negative breast cancer (TNBC) with intact DNA repair, we have previously shown an induced synthetic lethality with combined EGFR inhibition and PARPi. Here, we report the safety and clinical activity of lapatinib and veliparib in patients with metastatic TNBC. METHODS: A first-in-human, pilot study of lapatinib and veliparib was conducted in metastatic TNBC (NCT02158507). The primary endpoint was safety and tolerability. Secondary endpoints were objective response rates and pharmacokinetic evaluation. Gene expression analysis of pre-treatment tumor biopsies was performed. Key eligibility included TNBC patients with measurable disease and prior anthracycline-based and taxane chemotherapy. Patients with gBRCA1/2 mutations were excluded. RESULTS: Twenty patients were enrolled, of which 17 were evaluable for response. The median number of prior therapies in the metastatic setting was 1 (range 0-2). Fifty percent of patients were Caucasian, 45% African-American, and 5% Hispanic. Of evaluable patients, 4 demonstrated a partial response and 2 had stable disease. There were no dose-limiting toxicities. Most AEs were limited to grade 1 or 2 and no drug-drug interactions noted. Exploratory gene expression analysis suggested baseline DNA repair pathway score was lower and baseline immunogenicity was higher in the responders compared to non-responders. CONCLUSIONS: Lapatinib plus veliparib therapy has a manageable safety profile and promising antitumor activity in advanced TNBC. Further investigation of dual therapy with EGFR inhibition and PARP inhibition is needed. TRIAL REGISTRATION: ClinicalTrials.gov , NCT02158507 . Registered on 12 September 2014.

WNT pathway signaling is associated with microvascular injury and predicts kidney transplant failure.

Microvascular injury is associated with accelerated kidney transplant dysfunction and allograft failure. Molecular pathology can identify new mechanisms of microvascular injury while improving on the diagnostic and prognostic capabilities of traditional histology. We conducted a case-control study of archived kidney biopsy specimens stored up to 10 years with microvascular injury (n = 50) compared with biopsy specimens without histologic injury (n = 45) from patients of similar age, race, and sex. We measured WNT gene expression with a multiplex quantification platform by using digital barcoding, given the importance of WNT reactivation to the response to wounding in the kidney microvasculature and other compartments. Of 210 genes from a commercial WNT panel, 71 were associated with microvascular injury and 79 were associated with allograft failure, with considerable overlap of genes between each set. Molecular pathology identified 46 biopsy specimens with molecular evidence of microvascular injury; 18 (39%) were either C4d negative, donor-specific antibody negative, or had no microvascular injury by histology. The majority of cases with molecular evidence of microvascular injury had poor long-term outcomes. We identified novel WNT pathway genes associated with microvascular injury and allograft failure in residual clinical biopsy specimens obtained up to 10 years earlier. Further mechanistic studies may identify the WNT pathway as a new diagnostic and therapeutic target.

CHK1/2 Inhibitor Prexasertib Suppresses NOTCH Signaling and Enhances Cytotoxicity of Cisplatin and Radiation in Head and Neck Squamous Cell Carcinoma.

Platinum-based chemoradiotherapy is a mainstay of organ-preserving therapy for patients with head and neck squamous cell carcinoma cancer (HNSCC). However, the disease eventually becomes resistant to treatment necessitating new therapies. Checkpoint kinase 1 and 2 (CHK1/2) are serine/threonine kinases that activate cell-cycle checkpoints and serve a critical role in the DNA-damage response (DDR). As resistance to cisplatin and radiation may involve a heightened DDR, we hypothesized that prexasertib, an inhibitor of CHK1/2, may enhance the cytotoxicity induced by cisplatin and irradiation in HNSCC. In this study, we found that combining prexasertib with cisplatin and radiation significantly decreased the in vitro survival fraction in HNSCC cell lines both with and without radiotherapy. Reduced survival was accompanied by inhibition of DNA repair checkpoint activation, which resulted in persistent DNA damage and increased apoptosis. In addition, NanoString analysis with the PanCancer Pathways Panel revealed that prexasertib downregulated NOTCH signaling target genes (NOTCH1, NOTCH2, and NOTCH3) and their associated ligands (JAG1, JAG2, SKP2, MAML2, and DLL1). Prexasertib also reduced NOTCH1, NOTCH3 and HES1 protein expression. Importantly, a significant tumor growth delay was observed in vivo in both human papillomavirus (HPV)-positive UM-SCC47 and HPV-negative UM-SCC1 cell line xenografts treated with prexasertib, cisplatin, and radiotherapy without increased toxicity as measured by mouse body weight. Taken together, prexasertib reduced NOTCH signaling and enhanced the in vitro and in vivo response of HNSCCs to cisplatin and radiation, suggesting combination therapy may increase clinical benefit. A clinical trial has recently completed accrual (NCT02555644).

Therapeutically actionable PAK4 is amplified, overexpressed, and involved in bladder cancer progression.

Muscle-invasive bladder carcinomas (MIBCs) are aggressive genitourinary malignancies. Metastatic urothelial carcinoma of the bladder is generally incurable by current chemotherapy and leads to early mortality. Recent studies have identified molecular subtypes of MIBCs with different sensitivities to frontline therapy, suggesting tumor heterogeneity. We have performed multi-omic profiling of the kinome in bladder cancer patients with the goal of identify therapeutic targets. Our analyses revealed amplification, overexpression, and elevated kinase activity of P21 (RAC1) activated kinase 4 (PAK4) in a subset of Bladder cancer (BLCA). Using bladder cancer cells, we confirmed the role of PAK4 in BLCA cell proliferation and invasion. Furthermore, we observed that a PAK4 inhibitor was effective in curtailing growth of BLCA cells. Transcriptomic analyses identified elevated expression of another kinase, protein tyrosine kinase 6 (PTK6), upon treatment with a PAK4 inhibitor and RNA interference of PAK4. Treatment with a combination of kinase inhibitors (vandetanib and dasatinib) showed enhanced sensitivity compared with either drug alone. Thus, PAK4 may be therapeutically actionable for a subset of MIBC patients with amplified and/or overexpressed PAK4 in their tumors. Our results also indicate that combined inhibition of PAK4 and PTK6 may overcome resistance to PAK4. These observations warrant clinical investigations with selected BLCA patients.

Early Time-Restricted Feeding Improves 24-Hour Glucose Levels and Affects Markers of the Circadian Clock, Aging, and Autophagy in Humans.

Time-restricted feeding (TRF) is a form of intermittent fasting that involves having a longer daily fasting period. Preliminary studies report that TRF improves cardiometabolic health in rodents and humans. Here, we performed the first study to determine how TRF affects gene expression, circulating hormones, and diurnal patterns in cardiometabolic risk factors in humans. Eleven overweight adults participated in a 4-day randomized crossover study where they ate between 8 am and 2 pm (early TRF (eTRF)) and between 8 am and 8 pm (control schedule). Participants underwent continuous glucose monitoring, and blood was drawn to assess cardiometabolic risk factors, hormones, and gene expression in whole blood cells. Relative to the control schedule, eTRF decreased mean 24-hour glucose levels by 4 ± 1 mg/dl (p = 0.0003) and glycemic excursions by 12 ± 3 mg/dl (p = 0.001). In the morning before breakfast, eTRF increased ketones, cholesterol, and the expression of the stress response and aging gene SIRT1 and the autophagy gene LC3A (all p < 0.04), while in the evening, it tended to increase brain-derived neurotropic factor (BNDF; p = 0.10) and also increased the expression of MTOR (p = 0.007), a major nutrient-sensing protein that regulates cell growth. eTRF also altered the diurnal patterns in cortisol and the expression of several circadian clock genes (p < 0.05). eTRF improves 24-hour glucose levels, alters lipid metabolism and circadian clock gene expression, and may also increase autophagy and have anti-aging effects in humans.

Kinase analysis of penile squamous cell carcinoma on multiple platforms to identify potential therapeutic targets.

Penile squamous cell carcinoma (PSCC) is an orphan malignancy with poorly understood biology and suboptimal systemic therapy. Given that kinases may be drivers and readily actionable, we performed comprehensive multiplatform analysis of kinases in PSCC tumor and normal tissue. Fresh frozen tumors were collected from 11 patients with PSCC. After macrodissection to demarcate tumor from normal tissue, the samples underwent multiplatform analysis of kinases. Next Generation Sequencing (NGS) of 517 kinase genes was performed using Agilent Kinome capture and run on the Illumina MiSeq at PE150bp. The NanoString nCounter® platform analyzed the expression of 519 kinase genes. Kinase activity of tissue lysates was measured using PamStation®12 high-content phospho-peptide substrate microarray system. Network mapping was done with GeneGo MetaCore™ and upstream kinase prediction was performed with BioNavigator and the Kinexus database. Ingenuity pathway analysis was performed to integrate elevated kinase activity and gene over-expression with coexisting missense mutations at DNA level. Top pathways upregulated in both the kinase activity and gene expression platforms were PTEN, STAT3, GNRH, IL-8 and B cell receptor signaling. Potentially relevant missense mutations were seen in 176 kinase genes, with the top altered pathways overlapping with gene overexpression being GNRH, NF-kB and STAT3 signaling. ERBB2, ERBB3 and SYK were altered on NGS and also exhibited elevated kinase activity. To summarize, multiplatform comprehensive analysis of kinases discovered potential drivers of PSCC and actionable therapeutic targets. Translational studies are necessary to validate the functional relevance of our data to make advances in this rare malignancy.

Notch Signaling Activation Is Associated with Patient Mortality and Increased FGF1-Mediated Invasion in Squamous Cell Carcinoma of the Oral Cavity.

UNLABELLED: Oral squamous cell carcinoma (OSCC) is a cancer subtype that lacks validated prognostic and therapeutic biomarkers, and human papillomavirus status has not proven beneficial in predicting patient outcomes. A gene expression pathway analysis was conducted using OSCC patient specimens to identify molecular targets that may improve management of this disease. RNA was isolated from 19 OSCCs treated surgically at the University of Alabama at Birmingham (UAB; Birmingham, AL) and evaluated using the NanoString nCounter system. Results were confirmed using the oral cavity subdivision of the Head and Neck Squamous Cell Carcinoma Cancer (HNSCC) study generated by The Cancer Genome Atlas (TCGA) Research Network. Further characterization of the in vitro phenotype produced by Notch pathway activation in HNSCC cell lines included gene expression, proliferation, cell cycle, migration, invasion, and radiosensitivity. In both UAB and TCGA samples, Notch pathway upregulation was significantly correlated with patient mortality status and with expression of the proinvasive gene FGF1 In vitro Notch activation in HNSCC cells increased transcription of FGF1 and induced a marked increase in cell migration and invasion, which was fully abrogated by FGF1 knockdown. These results reveal that increased Notch pathway signaling plays a role in cancer progression and patient outcomes in OSCC. Accordingly, the Notch-FGF interaction should be further studied as a prognostic biomarker and potential therapeutic target for OSCC. IMPLICATIONS: Patients with squamous cell carcinoma of the oral cavity who succumb to their disease are more likely to have upregulated Notch signaling, which may mediate a more invasive phenotype through increased FGF1 transcription. Mol Cancer Res; 14(9); 883-91. ©2016 AACR.

Kinase Gene Expression Profiling of Metastatic Clear Cell Renal Cell Carcinoma Tissue Identifies Potential New Therapeutic Targets.

Kinases are therapeutically actionable targets. Kinase inhibitors targeting vascular endothelial growth factor receptors (VEGFR) and mammalian target of rapamycin (mTOR) improve outcomes in metastatic clear cell renal cell carcinoma (ccRCC), but are not curative. Metastatic tumor tissue has not been comprehensively studied for kinase gene expression. Paired intra-patient kinase gene expression analysis in primary tumor (T), matched normal kidney (N) and metastatic tumor tissue (M) may assist in identifying drivers of metastasis and prioritizing therapeutic targets. We compared the expression of 519 kinase genes using NanoString in T, N and M in 35 patients to discover genes over-expressed in M compared to T and N tissue. RNA-seq data derived from ccRCC tumors in The Cancer Genome Atlas (TCGA) were used to demonstrate differential expression of genes in primary tumor tissue from patients that had metastasis at baseline (n = 79) compared to those that did not develop metastasis for at least 2 years (n = 187). Functional analysis was conducted to identify key signaling pathways by using Ingenuity Pathway Analysis. Of 10 kinase genes overexpressed in metastases compared to primary tumor in the discovery cohort, 9 genes were also differentially expressed in TCGA primary tumors with metastasis at baseline compared to primary tumors without metastasis for at least 2 years: EPHB2, AURKA, GSG2, IKBKE, MELK, CSK, CHEK2, CDC7 and MAP3K8; p<0.001). The top pathways overexpressed in M tissue were pyridoxal 5'-phosphate salvage, salvage pathways of pyrimidine ribonucleotides, NF-kB signaling, NGF signaling and cell cycle control of chromosomal replication. The 9 kinase genes validated to be over-expressed in metastatic ccRCC may represent currently unrecognized but potentially actionable therapeutic targets that warrant functional validation.