UNIQUE REPORTER PROTEINS MANUFACTURED AND SUPPLIED BY PhaRNA, LLC

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Reporter proteins are detectable and measurable indicators of gene expression or cellular activity that allows investigators to monitor and analyze different biological processes. A marker gene or reporter gene is a gene that is introduced into cells in vitro or in vivo to identify and select those that have successfully acquired the genetic material (pDNA or mRNA) encoding that gene and translate it into the corresponding reporter protein.

To optimize procedures for modifying a cell outside and inside of organism with a gene of interest, a gene encoding one of the reporter proteins is often used in preliminary experiments. In other settings to introduce a reporter gene into an organism, scientists place the reporter gene and the gene of interest in the same mRNA bicistronic construct to be inserted into the cell or organism. It is important to use a reporter gene that is not natively expressed in the cell or organism under study, since the expression of the reporter is being used as a marker for successful uptake of the gene of interest.

Commonly used reporter genes that induce visually identifiable characteristics usually involve Fluorescent and Luminescent proteins. For example, mRNA encoding Jellyfish GFP (green fluorescent protein) and Discosoma mCherry (red fluorescent protein) proteins causes cells that express it to glow accordingly green and red under ultraviolet light, while the mRNA encoding the enzyme Luciferase induces catalyzes a reaction with substrate luciferin to produce light.  A common reporter in bacteria is the lacZ gene of E. coli, which encodes the enzyme beta-galactosidase, causing cells expressing this gene to appear blue when grown in media containing an analogue of the substrate X-gal.

PhaRNA, LLC offer several premade synthetic mRNAs encoding for some of the most common reporter proteins (https://www.pharna.com/reporter-proteins/) including mRNA of the Jellyfish GFP (green fluorescent protein), Discosoma mCherry (red fluorescent protein), firefly Luciferase (fLuc), renilla Luciferase (rLuc), gaussia Luciferase (gLuc) and beta-galactosidase (beta-Gal). In addition to these synthetic mRNAs encoding reporter proteins localized in the cellular cytosol, PhaRNA offers unique mRNAs encoding GFP-NLS (https://www.pharna.com/product/1500201/) and mCherry-NLS (https://www.pharna.com/product/1500801/) localized after translation in the nucleus. Such reporter proteins, which, after translation of the synthetic mRNA encoding them, are transferred into the nucleus, serve as the best control for those researchers who study structural, enzymatic and regulatory proteins functioning in the cell nucleus.       

Each synthetic mRNA encoding for reporter proteins localized in cellular cytosol or in nucleus can be readily assayed after translation and therefore used as control markers for mRNA packaging and delivery optimization in vitro and in vivo. These mRNAs are synthesized on error free sequence verified plasmid DNA based (non-PCR product) template with built-in constant poly-157A tail and enzymatically Cap1 caped, providing close to 100% capping efficiency and therefore superior mRNA stability and translatability.

PhaRNA – Redefining mRNA Therapeutics

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Since 2016, PhaRNA has been a trusted name in the synthetic RNA space, providing best-in-class, innovative synthetic RNA products to the scientific community and pharmaceutical and cosmeceutical organizations. The Company focuses on design and production of in vitro transcribed (IVT)-RNA constructs encoding for messenger RNA (mRNA), long non-coding RNA (lncRNA), and other RNA species of 150 nucleotides or longer. PhaRNA  established world’s largest catalog off-the-shelf synthetic mRNA (ready-to-be-delivered)  products (https://www.PhaRNA.com/mrna_catalog/) and provides a broad range of one-stop shop services including customized Research Grade and GMP Grade synthetic mRNA production, making it an ideal solution for preclinical and clinical applications.

PhaRNA produces high-fidelity synthetic mRNA generated from an error-free sequence-verified pDNA template which introduced with codon optimized coding sequence (CDS) of the gene of interest, 5’ and 3’ stabilizing Untranslated Regions (UTRs), and builds in 157 residues poly-A track. Each synthetic mRNA is capped with enzymatically introduced translation-boosting Cap 1 structure, terminated with extra-long 157 residues constant poly-A tail, and synthesized as modified mRNA with 100% replacement of Uridine-5′-triphosphate (UTP) on chemically modified UTP analogs (5-Methoxy-UTP, N1-Methylpseudo-UTP) to ensure generation of the synthetic mRNA species with improved pharmacological parameters such as stability, translational efficiency, and toxicity.

“Our unique combination of empirical and in-silico platforms allowing generation of a cell specific synthetic mRNA prototypes encoding gene of interest is a game-changer in synthetic mRNA design and optimization field, which is the result of our team’s extensive scientific and industry experience and expertise in producing of IP protected synthetic mRNA drug products.” says Eduard Yakubov, Co-founder and Chief Scientific Officer at PhaRNA.

“PhaRNA’s cutting-edge technologies, laboratory facilities, and scientists with extensive innovation histories have helped the company garner client success stories”, Yakubov states. In one instance, PhaRNA assisted a client in witnessing the release of synthetic mRNA encoding growth factor (HGF) from scaffolds in a temporally controlled fashion in vivo with extended translation of active HGF protein. When sutured to the ligated porcine femoral artery, such modified by synthetic HGF mRNA scaffolds enhanced vascular regeneration.

Moving ahead, PhaRNA aspires to continue its legacy as a leader in the synthetic mRNA field and expand its employee base with new talent. With its best-in-class material and human resources, the company intends sequentially advance synthetic mRNA-based drug research and serve the increased needs of scientific, pharmaceutical, and cosmeceutical companies.

Growth Factor Receptors

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Growth factors, also known as trophic factors, bind to cell-surface receptors to initiate signaling cascades that lead to cell development and differentiation. Their influence on cell growth is emphasized in cancer studies.

Defining Growth Factor Receptors 

Transmembrane proteins, known as growth factor receptors, attach to specific growth factors and communicate the signals those factors convey to the intracellular environment.

As the name implies, growth factor receptors bind to growth factors and serve to regulate growth factors. Growth factor receptor signaling triggers cell division and differentiation. Activation of GFR is the first step in the division or enlargement of a cell, and this process begins with the binding of growth factors. Growth factors bind to these cells. The signaling pathways MAP kinase, JAK / STAT, and PI3 kinase may all be involved in this receptor activation.

It has been found that these receptors are mainly found in cells and in high density on cell surfaces. A significant proportion of GFR, such as those of insulin-like growth factor (IGF), fibroblast growth factor (FGF), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), and platelet-derived growth factor (PDGF), contain a cysteine-rich region involved in signal transduction by receptor tyrosine kinases. For some receptors, such as the transforming growth factor receptor, serine-threonine kinase facilitates signal transduction as a consequence of its action.

Various Growth Factor Receptor Examples

Cell proliferation, development, and differentiation are tightly regulated by a range of growth factors and their receptors.

In addition to Wnt receptors, ties, neurotrophin receptors, ephrine receptors, insulin-like growth factor receptors, epidermal growth factor receptors, fibroblast growth factor receptors, platelet-derived growth factor receptors, and vascular endothelial growth factor receptors, GFR also includes insulin-like growth factor receptors.

Significance of GFRs in Cancer Therapy

ErbBs, a group of receptor tyrosine kinases belonging to the epidermal growth factor family, are crucial for controlling cell motility, differentiation, and proliferation. The ErbB receptors play both redundant and specialized roles in the maintenance of tissues in mature mammals and in mammals’ development.

GFs are essential for basement membrane disruption, cancer cell invasion into nearby tissues, the vascular or lymphatic systems, as well as their exit from the bloodstream and subsequent colonisation of remote organs (intravasation, extravasation). Tumor spread is signalled by the transition of densely packed polarised epithelial cells into individual, motile cells, known as the epithelial-mesenchymal transition (EMT).  

Hepatocellular carcinoma is one type of cancer where growth factors and their corresponding receptors are frequently overexpressed and/or dysregulated. (HCC). Clinical studies show that growth factor receptors and the associated signalling pathways play significant roles in the development and progression of HCC cancer. This makes them viable targets for cutting-edge cancer treatments. Tyrosine kinase inhibitors (also known as “small molecule inhibitors”), antisense oligonucleotides, and monoclonal antibodies have all been tested for their ability to block the activity and subsequent signalling pathways of these receptors in HCC. 

Preclinical trials are presently being conducted to research HCC. A further potential strategy for treating HCCs is to inhibit tumour vessel growth by interfering with the VEGF/VEGFR system. This is because HCCs are hypervascularized neoplasms. 

Other Medical Relevance 

Growth factors play a crucial role in both physiologically normal processes like wound repair and physiologically abnormal processes like cancer and diabetic retinopathy. Diabetic retinopathy causes blindness because abnormal retinal blood vessel growth causes the disease. 

In order to target cancer treatment, research currently focuses on GFRs. Receptors for epidermal growth factors play a significant role in cancer activity. An internal signal transduction system ensures cell functions after growth factors bind to their receptors. Cancerous cells, however, may never switch the pathway on or off. Furthermore, it has been found that certain cancers frequently exhibit overexpression of receptors (like RTKs), which is consistent with unchecked cell growth and differentiation. Tyrosine receptors are often a focus in cancer therapy.

Conclusion 

An overview of growth factor receptors is given in this text. If you’re interested in reading more about these subjects, subscribe to our blog and leave a comment below. You can subscribe to our site if you’re interested in reading more about these subjects.

An Introduction to Cytokine Receptors

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Cytokine receptors are a diverse family of proteins that play a critical role in the immune system. They regulate a variety of cellular processes, including cell survival, proliferation, differentiation, and migration. Dysregulation of cytokine receptor signaling can lead to a variety of diseases, including autoimmune and inflammatory diseases and cancer. In this blog, we will explore the basic introduction, structure, function, and therapeutic potential of these receptors.

Types
These receptors can be classified into several different categories based on their structure and function. For example, some receptors are composed of single transmembrane domains and bind to cytokines that have a four-helix bundle structure, while others are composed of multiple transmembrane domains and bind to cytokines that are structurally distinct.

Structure
We can say that these receptors are typically transmembrane proteins with an extracellular ligand-binding domain and an intracellular domain responsible for transmitting signals to the cell. The extracellular domain of these receptors is composed of several distinct structural domains, including fibronectin type III, cytokine receptor homology, and Ig-like domains. These domains play a critical role in ligand binding and receptor activation.

Function
These receptors transmit signals from cytokines, which are small secreted proteins that regulate cellular processes. Upon binding to their respective cytokines, these receptors undergo conformational changes that activate downstream signaling pathways. These pathways can lead to changes in gene expression, protein production, and cellular behavior.

Importance
Dysregulation of cytokine receptor signaling can lead to a variety of diseases, including autoimmune and inflammatory diseases and cancer. In recent years, targeting these receptors and their downstream signaling pathways has become a promising strategy for developing new therapies for these diseases. The development of cytokine receptor-targeted therapies has already led to significant improvements in autoimmune and inflammatory diseases. Ongoing research is likely to uncover additional targets for drug development in the future.

Therapeutic Potential
Cytokine receptor-targeted therapies have been approved for autoimmune and inflammatory diseases. Tocilizumab, for example, is a monoclonal antibody that targets the IL-6 receptor. IL-6 is a pro-inflammatory cytokine implicated in rheumatoid arthritis and other autoimmune diseases. Tocilizumab binds to the IL-6 receptor, preventing IL-6 from binding and activating downstream signaling pathways. Another example of a cytokine receptor-targeted therapy is ustekinumab, a monoclonal antibody that targets the IL-12/IL-23 receptor. IL-12 and IL-23 are pro-inflammatory cytokines implicated in psoriasis and other autoimmune diseases. Ustekinumab binds to the IL-12/IL-23 receptor, preventing IL-12 and IL-23 from activating downstream signaling pathways. This reduces inflammation and improves disease symptoms.

Clinical applications
The drugs targeting these particular receptors have become an important class of therapeutics for a variety of diseases, including rheumatoid arthritis, inflammatory bowel disease, and psoriasis.
Understanding the mechanisms by which these drugs work can provide insights into potential new therapeutic targets for these and other diseases.

Challenges and future directions
Despite the success of cytokine receptor-targeted therapies, there are still many challenges and unanswered questions in this field. For example, it is not always clear why some patients respond well to certain therapies while others do not, and there may be unexpected side effects associated with long-term use of these drugs. In-depth research will be required to better understand the complex interactions between cytokines, cytokine receptors, and the immune system, and to develop new and more effective therapies for autoimmune and inflammatory diseases.

Conclusion
These receptors are critical components of the immune system, responsible for regulating a variety of cellular processes. Cancer and autoimmune diseases can be caused by dysregulation of cytokine receptor signaling. Targeting these receptors and their downstream signaling pathways has become a promising strategy for developing new therapies for these diseases. The development of cytokine receptor-targeted therapies has already led to significant improvements in autoimmune and inflammatory diseases. Ongoing research is likely to uncover additional targets for drug development in the future.
Reach out to PhaRNA to get detailed information regarding available products and therapies.