Mechanism of c-Met and EGFR tyrosine kinase inhibitor resistance through epithelial mesenchymal transition in non-small cell lung cancer.

According to currently available estimates from Cancer Research UK, 14.1 million new lung cancer cases were diagnosed and a staggering 8.2 million people worldwide died from lung cancer in 2012. EGFR and c-Met are two tyrosine kinase receptors most commonly overexpressed or mutated in Non-small Cell Lung Cancer (NSCLC) resulting in increased proliferation and survival of lung cancer cells. Tyrosine kinase inhibitors (TKIs), such as erlotinib, approved by the FDA as first/second line therapy for NSCLC patients have limited clinical efficacy due to acquired resistance. In this manuscript, we investigate and discuss the role of epithelial mesenchymal transition (EMT) in the development of resistance against EGFR and c-Met TKIs in NSCLC. Our findings show that Zeb-1, a transcriptional repressor of E-Cadherin, is upregulated in TKI-resistant cells causing EMT. We observed that TKI-resistant cells have increased gene and protein expression of EMT related proteins such as Vimentin, N-Cadherin, ß-Catenin and Zeb-1, while expression of E-Cadherin, an important cell adhesion molecule, was suppressed. We also confirmed that TKI-resistant cells display mesenchymal cell type morphology, and have upregulation of ß-Catenin which may regulate expression of Zeb-1, a transcriptional repressor of E-Cadherin in TKI-resistant NSCLC cells. Finally, we show that down-regulating Zeb-1 by inducing miR-200a or ß-Catenin siRNA can increase drug sensitivity of TKI-resistant cells.

B cells require licensing by dendritic cells to serve as primary antigen-presenting cells for plasmid DNA.

DNA vaccines have been an attractive approach for treating cancer patients, however have demonstrated modest immunogenicity in human clinical trials. Dendritic cells (DCs) are known to cross-present DNA-encoded antigens expressed in bystander cells. However, we have previously reported that B cells, and not DCs, serve as primary antigen-presenting cells (APCs) following passive uptake of plasmid DNA. Here we sought to understand the requirements for B cells to present DNA-encoded antigens, to ultimately increase the immunogenicity of plasmid DNA vaccines. Using ovalbumin-specific OT-1 CD8+ T cells and isolated APC populations, we demonstrated that following passive uptake of plasmid DNA, B cells but not DC, can translate the encoded antigen. However, CD8 T cells were only activated by B cells when they were co-cultured with DCs. We found that a cell-cell contact is required between B cells and DCs. Using MHCI KO and re-purification studies, we demonstrated that B cells were the primary APCs and DCs serve to license this function. We further identified that the gene expression profiles of B cells that have been licensed by DCs, compared to the B cells that have not, are vastly different and have signatures similar to B cells activated with a TLR7/8 agonist. Our data demonstrate that B cells transcribe and translate antigens encoded by plasmid DNA following passive uptake, however require licensing by live DC to present antigen to CD8 T cells. Further study of the role of B cells as APCs will be important to improve the immunological efficacy of DNA vaccines.

Vaccines as treatments for prostate cancer.

Prostate cancer is a leading cause of death in men worldwide. For over 30 years, growing interest has focused on the development of vaccines as treatments for prostate cancer, with the goal of using vaccines to activate immune cells capable of targeting prostate cancer to either eradicate recurrent disease or at least delay disease progression. This interest has been prompted by the prevalence and long natural history of the disease and by the fact that the prostate is an expendable organ. Thus, an immune response elicited by vaccination might not need to target the tumour uniquely but could theoretically target any prostate tissue. To date, different vaccine approaches and targets for prostate cancer have been evaluated in clinical trials. Overall, five approaches have been assessed in randomized phase III trials and sipuleucel-T was approved as a treatment for metastatic castration-resistant prostate cancer, being the only vaccine approved to date by the FDA as a treatment for cancer. Most vaccine approaches showed safety and some evidence of immunological activity but had poor clinical activity when used as monotherapies. However, increased activity has been observed when these vaccines were used in combination with other immune-modulating therapies. This evidence suggests that, in the future, prostate cancer vaccines might be used to activate and expand tumour-specific T cells as part of combination approaches with agents that target tumour-associated immune mechanisms of resistance.

Genetic diversity of medically important scorpions of the genus Centruroides (Buthidae) from Panama including two endemic species.

With more than 33,000 sting cases and 47 deaths recorded between 2000 and 2016, Panama is the country with the highest incidence of envenomations by scorpions in Central America. Species in the genus Centruroides are responsible for most scorpion sting reports, however, identification at the species level is complicated because the genus has considerable intraspecific morphological variation. To date no molecular data have been reported from Panama that would help to estimate their genetic diversity and validate morphometric identification methods. We provide here the first genetic diversity data of the two endemic species (C. granosus and C. panamensis) and other two species reported in Panama (C. bicolor and C. limbatus). A total of 41 specimens were sequenced for COI and 16S rDNA mitochondrial genes. The phylogenetic concatenated analysis separates the Panamanian samples into four well-supported clades represented by C. bicolor, C. granosus and (C. panamensis + C. limbatus). The two endemic species are not the closest relatives in the tree. Low diversity in combination with its very narrow distribution suggest that C. panamensis is susceptible to environmental degradation. A single specimen of Coiba island is intermediate in the tree structure between C. bicolor and C. panamensis and may represent an early stage of speciation. The haplotype network is also consistent with the phylogenetic trees.

Role of B cells as antigen presenting cells.

B cells have been long studied for their role and function in the humoral immune system. Apart from generating antibodies and an antibody-mediated memory response against pathogens, B cells are also capable of generating cell-mediated immunity. It has been demonstrated by several groups that B cells can activate antigen-specific CD4 and CD8 T cells, and can have regulatory and cytotoxic effects. The function of B cells as professional antigen presenting cells (APCs) to activate T cells has been largely understudied. This, however, requires attention as several recent reports have demonstrated the importance of B cells within the tumor microenvironment, and B cells are increasingly being evaluated as cellular therapies. Antigen presentation through B cells can be through antigen-specific (B cell receptor (BCR) dependent) or antigen non-specific (BCR independent) mechanisms and can be modulated by a variety of intrinsic and external factors. This review will discuss the pathways and mechanisms by which B cells present antigens, and how B cells differ from other professional APCs.

Treatment Combinations with DNA Vaccines for the Treatment of Metastatic Castration-Resistant Prostate Cancer (mCRPC).

Metastatic castration-resistant prostate cancer (mCRPC) is a challenging disease to treat, with poor outcomes for patients. One antitumor vaccine, sipuleucel-T, has been approved as a treatment for mCRPC. DNA vaccines are another form of immunotherapy under investigation. DNA immunizations elicit antigen-specific T cells that cause tumor cell lysis, which should translate to meaningful clinical responses. They are easily amenable to design alterations, scalable for large-scale manufacturing, and thermo-stable for easy transport and distribution. Hence, they offer advantages over other vaccine formulations. However, clinical trials with DNA vaccines as a monotherapy have shown only modest clinical effects against tumors. Standard therapies for CRPC including androgen-targeted therapies, radiation therapy and chemotherapy all have immunomodulatory effects, which combined with immunotherapies such as DNA vaccines, could potentially improve treatment. In addition, many investigational drugs are being developed which can augment antitumor immunity, and together with DNA vaccines can further enhance antitumor responses in preclinical models. We reviewed the literature available prior to July 2020 exploring the use of DNA vaccines in the treatment of prostate cancer. We also examined various approved and experimental therapies that could be combined with DNA vaccines to potentially improve their antitumor efficacy as treatments for mCRPC.

Mechanism of Resistance and Novel Targets Mediating Resistance to EGFR and c-Met Tyrosine Kinase Inhibitors in Non-Small Cell Lung Cancer.

Tyrosine kinase inhibitors (TKIs) against EGFR and c-Met are initially effective when administered individually or in combination to non-small cell lung cancer (NSCLC) patients. However, the overall efficacies of TKIs are limited due to the development of drug resistance. Therefore, it is important to elucidate mechanisms of EGFR and c-Met TKI resistance in order to develop more effective therapies. Model NSCLC cell lines H1975 and H2170 were used to study the similarities and differences in mechanisms of EGFR/c-Met TKI resistance. H1975 cells are positive for the T790M EGFR mutation, which confers resistance to current EGFR TKI therapies, while H2170 cells are EGFR wild-type. Previously, H2170 cells were made resistant to the EGFR TKI erlotinib and the c-Met TKI SU11274 by exposure to progressively increasing concentrations of TKIs. In H2170 and H1975 TKI-resistant cells, key Wnt and mTOR proteins were found to be differentially modulated. Wnt signaling transducer, active ß-catenin was upregulated in TKI-resistant H2170 cells when compared to parental cells. GATA-6, a transcriptional activator of Wnt, was also found to be upregulated in resistant H2170 cells. In H2170 erlotinib resistant cells, upregulation of inactive GSK3ß (p-GSK3ß) was observed, indicating activation of Wnt and mTOR pathways which are otherwise inhibited by its active form. However, in H1975 cells, Wnt modulators such as active ß-catenin, GATA-6 and p-GSK3ß were downregulated. Additional results from MTT cell viability assays demonstrated that H1975 cell proliferation was not significantly decreased after Wnt inhibition by XAV939, but combination treatment with everolimus (mTOR inhibitor) and erlotinib resulted in synergistic cell growth inhibition. Thus, in H2170 cells and H1975 cells, simultaneous inhibition of key Wnt or mTOR pathway proteins in addition to EGFR and c-Met may be a promising strategy for overcoming EGFR and c-Met TKI resistance in NSCLC patients.

Current Molecularly Targeting Therapies in NSCLC and Melanoma.

Surgery, radiation therapy, and chemotherapy are the traditional options to control tumor progression. However, these strategies are fraught with harmful side effects and are ineffective in metastatic and advanced cancers. Biomarkers that are overexpressed in cancers and are involved in cell growth, proliferation, migration, and survival have recently become the focus of new molecular targeting therapies. Novel therapies targeting biomarkers have roles in tumorigenesis that are overexpressed in cancers may be more efficacious and less toxic in comparison to traditional therapies. These therapies include the use of tyrosine kinase inhibitors and monoclonal antibodies for the treatment of cancer. However, the efficacy of these therapies is limited due to the development of drug resistance after prolonged treatment. Current research is focused on understanding mechanisms of resistance to overcome the barriers limiting the use of these targeting therapies in the treatment of cancer. In this review, we will discuss the clinical status of tyrosine kinase inhibitors and monoclonal antibodies against several prevalent biomarkers that are candidates for therapy in non-small cell lung cancer (NSCLC) and melanoma.