Transcriptional Antagonism by CDK8 Inhibition Improves Therapeutic Efficacy of MEK Inhibitors.

Aberrant RAS/MAPK signaling is a common driver of oncogenesis that can be therapeutically targeted with clinically approved MEK inhibitors. Disease progression on single-agent MEK inhibitors is common, however, and combination therapies are typically required to achieve significant clinical benefit in advanced cancers. Here we focused on identifying MEK inhibitor-based combination therapies in neuroblastoma with mutations that activate the RAS/MAPK signaling pathway, which are rare at diagnosis but frequent in relapsed neuroblastoma. A genome-scale CRISPR-Cas9 functional genomic screen was deployed to identify genes that when knocked out sensitize RAS-mutant neuroblastoma to MEK inhibition. Loss of either CCNC or CDK8, two members of the mediator kinase module, sensitized neuroblastoma to MEK inhibition. Furthermore, small-molecule kinase inhibitors of CDK8 improved response to MEK inhibitors in vitro and in vivo in RAS-mutant neuroblastoma and other adult solid tumors. Transcriptional profiling revealed that loss of CDK8 or CCNC antagonized the transcriptional signature induced by MEK inhibition. When combined, loss of CDK8 or CCNC prevented the compensatory upregulation of progrowth gene expression induced by MEK inhibition. These findings propose a new therapeutic combination for RAS-mutant neuroblastoma and may have clinical relevance for other RAS-driven malignancies. SIGNIFICANCE: Transcriptional adaptation to MEK inhibition is mediated by CDK8 and can be blocked by the addition of CDK8 inhibitors to improve response to MEK inhibitors in RAS-mutant neuroblastoma, a clinically challenging disease.

Nanoengineered Disruption of Heat Shock Protein 90 Targets Drug-Induced Resistance and Relieves Natural Killer Cell Suppression in Breast Cancer.

Drug-induced resistance, or tolerance, is an emerging yet poorly understood failure of anticancer therapy. The interplay between drug-tolerant cancer cells and innate immunity within the tumor, the consequence on tumor growth, and therapeutic strategies to address these challenges remain undescribed. Here, we elucidate the role of taxane-induced resistance on natural killer (NK) cell tumor immunity in triple-negative breast cancer (TNBC) and the design of spatiotemporally controlled nanomedicines, which boost therapeutic efficacy and invigorate "disabled" NK cells. Drug tolerance limited NK cell immune surveillance via drug-induced depletion of the NK-activating ligand receptor axis, NK group 2 member D, and MHC class I polypeptide-related sequence A, B. Systems biology supported by empirical evidence revealed the heat shock protein 90 (Hsp90) simultaneously controls immune surveillance and persistence of drug-treated tumor cells. On the basis of this evidence, we engineered a "chimeric" nanotherapeutic tool comprising taxanes and a cholesterol-tethered Hsp90 inhibitor, radicicol, which targets the tumor, reduces tolerance, and optimally reprimes NK cells via prolonged induction of NK-activating ligand receptors via temporal control of drug release in vitro and in vivo. A human ex vivo TNBC model confirmed the importance of NK cells in drug-induced death under pressure of clinically approved agents. These findings highlight a convergence between drug-induced resistance, the tumor immune contexture, and engineered approaches that consider the tumor and microenvironment to improve the success of combinatorial therapy. SIGNIFICANCE: This study uncovers a molecular mechanism linking drug-induced resistance and tumor immunity and provides novel engineered solutions that target these mechanisms in the tumor and improve immunity, thus mitigating off-target effects.

Predicting clinical response to anticancer drugs using an ex vivo platform that captures tumour heterogeneity.

Predicting clinical response to anticancer drugs remains a major challenge in cancer treatment. Emerging reports indicate that the tumour microenvironment and heterogeneity can limit the predictive power of current biomarker-guided strategies for chemotherapy. Here we report the engineering of personalized tumour ecosystems that contextually conserve the tumour heterogeneity, and phenocopy the tumour microenvironment using tumour explants maintained in defined tumour grade-matched matrix support and autologous patient serum. The functional response of tumour ecosystems, engineered from 109 patients, to anticancer drugs, together with the corresponding clinical outcomes, is used to train a machine learning algorithm; the learned model is then applied to predict the clinical response in an independent validation group of 55 patients, where we achieve 100% sensitivity in predictions while keeping specificity in a desired high range. The tumour ecosystem and algorithm, together termed the CANScript technology, can emerge as a powerful platform for enabling personalized medicine.

Effect of detoxified karanja (Pongamia spp.) cake on testicular architecture and semen production in ram lambs.

The protein-rich non-conventional detoxified karanja cake (dKC) can be used in place of conventional protein supplements like soybean meal (SBM), groundnut meal, etc. in livestock feed. The present study was conducted to assess the effect of two levels of dKC by replacing SBM on testicular architecture, semen quality and expressions of mRNAs encoding luteinizing hormone receptor (LHR) and insulin-like growth factor (IGF-I) in testes of ram lambs. Eighteen ram lambs were randomly divided into three groups (n = 6) and fed different levels (%) of karanja cake (0% replacement--control; 50% replacement--dKC-50 and 75% replacement--dKC-75) for 140 days. After 120 days of feeding, the semen from the animals was collected and analysed. The testes samples were collected on day 140 of feeding for transcripts expression studies. The dKC-50 group had no change in BW, whereas dKC-75 group showed decreased (P < 0.05) BW as compared with control. The number of animals ejaculated semen in dKC-75 group was lower (P < 0.05) than the control group. A reduction (P < 0.05) in LHR expression in dKC-75 was observed, whereas a reduction in IGF-I expression (P < 0.05) was observed in dKC-50 and dKC-75 as compared with control group. The study reveals that in ram lambs, long-term feeding of dKC at 50% replacement of SBM may not affect BW. However, long-term feeding of dKC as a replacement of SBM may affect testicular function.

Inhibition of rapamycin-induced AKT activation elicits differential antitumor response in head and neck cancers.

The PI3K/AKT/mTOR pathway is an important signaling axis that is perturbed in majority of cancers. Biomarkers such as pS6RP, GLUT1, and tumor FDG uptake are being evaluated in patient stratification for mTOR pathway inhibitors. In the absence of a clear understanding of the underlying mechanisms in tumor signaling, the biomarker strategy for patient stratification is of limited use. Here, we show that no discernible correlation exists between FDG uptake and the corresponding Ki67, GLUT1, pS6RP expression in tumor biopsies from patients with head and neck cancer. Correlation between GLUT1 and pS6RP levels in tumors was observed but elevated pS6RP was noticed even in the absence of concomitant AKT activation, suggesting that other downstream molecules of PI3K/AKT and/or other pathways upstream of mTOR are active in these tumors. Using an ex vivo platform, we identified putative responders to rapamycin, an mTOR inhibitor in these tumors. However, rapamycin did not induce antitumor effect in the majority of tumors with activated mTOR, potentially attributable to the observation that rapamycin induces feedback activation of AKT. Accordingly, treatment of these tumors with an AKT inhibitor and rapamycin uniformly resulted in abrogation of mTOR inhibition-induced AKT activation in all tumors but failed to induce antitumor response in a subset. Phosphoproteomic profiling of tumors resistant to dual AKT/mTOR inhibitors revealed differential activation of multiple pathways involved in proliferation and survival. Collectively, our results suggest that, in addition to biomarker-based segregation, functional assessment of a patient's tumor before treatment with mTOR/AKT inhibitors may be useful for patient stratification.

Evaluation of maize grain and polyunsaturated fatty acid (PUFA) as energy sources for breeding rams based on hormonal, sperm functional parameters and fertility.

The objective of the present study was to elucidate the effect of different sources of dietary energy (maize vs polyunsaturated fatty acid (PUFA) on semen functional parameters and fertility of adult rams. Eighteen adult rams were divided into two groups (maize and PUFA, n=9). The main energy source for the rams in the maize group was coarsely ground maize grain, whereas in the PUFA group it was sunflower oil (rich in 18:2 linoleic acid, an omega-6 acid). The ration was fed for a minimum period of 60 days and thereafter semen was collected for evaluation. The proportion of progressive forward motility was significantly (P<0.05) higher in the PUFA group compared with the maize group. Sperm lipid peroxidation as measured by malondialdehyde formation (µM per 1×10(9) spermatozoa) was significantly (P<0.05) higher in the PUFA group compared with the maize group. When the semen was diluted with Tris-egg yolk-citrate buffer and incubated for 24h at 4°C, the proportions of plasmalemma integrity, the sperm subpopulation positive for functional membrane and acrosomal integrities, and mitochondrial membrane potential were significantly (P<0.05) higher in PUFA-fed than in maize-fed animals. The different sources of energy did not influence the serum and seminal plasma IGF-I levels. The cleavage rate (percentage) did not differ significantly between PUFA- (45.4±4.91) and maize- (44.63±6.8) fed animals. In conclusion, PUFA feeding influenced sperm quality by altering or stabilising membrane integrity. The present study indicates that PUFA may improve semen quality but did not improve in vitro fertilisation.