Recombinant Mouse G-protein coupled receptor 87 (Gpr87)

What is GPR87 and what is its expression pattern in normal versus diseased tissues?

GPR87 is a G protein-coupled receptor that shows specific expression in tumor cells, including lung, pancreatic, bladder, skin, and cervical cancers, while being rarely expressed in normal cells . This tumor-specific expression pattern makes it a promising target for cancer-specific therapies. Studies have demonstrated that high GPR87 expression in lung and pancreatic cancers correlates with worse prognosis for patients . In pulmonary fibrosis, GPR87 is highly expressed in basal and aberrant basaloid cells, with expression levels correlating with disease severity .

To properly characterize GPR87 expression, researchers typically employ quantitative RT-PCR, western blotting, and immunohistochemistry across tissue panels. Single-cell RNA sequencing has recently provided more granular insights into cell-type specific expression patterns, particularly in identifying the specific cell populations that express GPR87 in complex tissues like fibrotic lungs .

What signaling pathways does GPR87 participate in?

GPR87 participates in several important signaling pathways that contribute to disease progression:

• JAK2/STAT3 pathway: GPR87 forms a positive feedback loop with JAK2 and STAT3 to promote the expansion of pancreatic ductal adenocarcinoma stem cells. STAT3 directly binds to the promoter of GPR87 to increase its expression, while GPR87 activates JAK2, which in turn activates STAT3 .

• PI3K/mTOR pathway: In pulmonary fibrosis models, GPR87 signals through the PI3K and mTOR pathways, with stimulation or inhibition of the PI3K pathway mimicking GPR87 activation or inhibition responses, respectively .

• TNF/NFκB pathway: Research indicates that GPR87 also signals through the TNF/NFκB pathway in fibrotic disease contexts .

• KRAS and c-Myc expression: Inhibition of GPR87 expression leads to significant decreases in KRAS and c-Myc expression in lung cancer cell lines, suggesting a regulatory relationship between GPR87 and these oncogenes .

How does GPR87 contribute to cancer stem cell maintenance and expansion?

GPR87 plays a critical role in promoting cancer stem cell properties, particularly in pancreatic ductal adenocarcinoma (PDA). Research has demonstrated that GPR87 significantly enhances sphere formation ability, increases side population (SP) cell number, elevates expression of PDA stem cell markers (CD133, EpCAM, CD24, CD44, and MET), and increases tumor initiation ability in vivo .

The mechanism involves a positive feedback loop between GPR87, JAK2, and STAT3. STAT3 directly binds to the GPR87 promoter to increase its expression, while GPR87 activates JAK2, which subsequently activates STAT3. This loop creates a self-reinforcing circuit that maintains and expands the cancer stem cell population .

Experimental evidence supporting this role includes:

• Sphere formation assays showing GPR87 overexpression increases sphere size and generation frequency (~6% vs. ~1.6% in knockdown cells)

• Side population assays demonstrating GPR87 overexpression increases stem cell-enriched SP-positive population

• qPCR analysis showing GPR87 overexpression upregulates stem cell markers CD133, EpCAM, CD24, CD44, and MET

• In vivo tumor initiation assays revealing that as few as 1,000 GPR87-overexpressing cells can generate tumors, while the same number of GPR87 knockdown cells fails to do so

What is the role of GPR87 in pulmonary fibrosis pathogenesis?

Recent research has uncovered a significant role for GPR87 in the development of idiopathic pulmonary fibrosis (IPF). GPR87 is highly expressed in basal and aberrant basaloid cells in IPF lungs, with expression levels correlating with disease severity .

Several lines of evidence demonstrate GPR87's profibrotic effects:

• In vivo studies: GPR87 knockout (GPR87-/-) mice are protected against bleomycin-induced pulmonary fibrosis, suggesting that GPR87 signaling is necessary for fibrotic development .

• Ex vivo studies: GPR87 knockdown is protective against fibrosis development in normal precision-cut lung slices (PCLS) treated with fibrotic cocktail and leads to fibrosis regression in IPF PCLS .

• In vitro studies: In induced pluripotent stem cell-derived airway basal cells (iBC), GPR87 knockdown leads to decreased expression of fibrosis-related genes, proteins, and microRNAs, while GPR87 stimulation with lysophosphatidic acid (LPA) produces opposite results .

The main downstream pathways involved in GPR87-mediated fibrosis are PI3K, mTOR, and TNF/NFκB. Importantly, stimulation or inhibition of the PI3K pathway mimics GPR87 stimulation or inhibition responses, respectively, identifying a potential therapeutic intervention point .

What approaches are effective for targeting GPR87 in cancer therapy?

Several approaches have been developed to target GPR87 in cancer therapy:

• Near-infrared photoimmunotherapy (NIR-PIT): This novel cancer therapy combines a photosensitizer (IRDye700DX) with anti-GPR87 antibodies to specifically destroy GPR87-expressing cancer cells when irradiated with near-infrared light. To avoid human anti-mouse antibody (HAMA) responses, humanized anti-GPR87 antibodies (huGPR87ab) have been developed from mouse anti-GPR87 antibodies (moGPR87ab) .

• RNA interference: Short hairpin RNA (shRNA) targeting GPR87 has been delivered via adenoviral vectors (Ad-shGPR87) to effectively downregulate GPR87 expression. This approach significantly inhibits cell proliferation in GPR87-overexpressing lung cancer cell lines H358 and PC9, and exerts significant antitumor effects against GPR87-expressing H358 xenografts in vivo .

• Recombinant antibodies: Recombinant mouse antibodies against GPR87, such as TAB-002CT, have been developed with high purity (>95% as determined by SDS-PAGE and SEC-HPLC) and specificity, offering consistent research tools for GPR87 targeting. These antibodies are available in different formats, including full-length antibodies, scFv fragments, and Fab fragments, providing flexibility for different targeting approaches .

What methods are available for detecting and quantifying GPR87 expression?

Researchers have employed several complementary techniques to detect and quantify GPR87 expression:

• Quantitative RT-PCR: Used to measure GPR87 mRNA expression levels, this technique is valuable for comparing expression across cell lines, tissues, or in response to treatments. In studies examining GPR87's role in cancer stem cells, qPCR has been used to correlate GPR87 expression with stem cell marker expression .

• Western blot analysis: Utilized to detect GPR87 protein expression, western blotting has been instrumental in confirming the effects of GPR87 knockdown or overexpression at the protein level. This technique has also been used to examine downstream signaling effects, such as changes in KRAS and c-Myc expression following GPR87 inhibition .

• Immunohistochemistry: Essential for visualizing GPR87 expression in tissue samples, this method helps identify cell-specific expression patterns, particularly in heterogeneous tissues like tumors or fibrotic lungs .

• RNA sequencing: Both bulk and single-cell RNA sequencing have been employed to comprehensively analyze GPR87 expression patterns. Single-cell RNA sequencing has been particularly valuable in identifying specific cell populations that express GPR87 in complex tissues, such as the basal and aberrant basaloid cells in IPF lungs .

• ELISA and Dot Blot analysis: For antibody characterization and validation, ELISA and Dot Blot analyses have been used to confirm antibody specificity against recombinant GPR87 proteins .

What are effective approaches for manipulating GPR87 expression in experimental models?

Researchers have employed several strategies to manipulate GPR87 expression:

• Genetic knockdown: Short interfering RNAs (siRNAs) targeting GPR87 have been used in cell culture models, particularly in precision-cut lung slices (PCLS) from normal and IPF tissues to study the role of GPR87 in fibrosis .

• Viral vector-mediated knockdown: Adenoviral vectors expressing short hairpin RNAs targeting GPR87 (Ad-shGPR87) have been developed for more efficient delivery and sustained knockdown. This approach has been effective both in vitro in cancer cell lines and in vivo in xenograft models .

• Genetic knockout: GPR87 knockout mice (GPR87-/-) have been generated using targeted gene editing, providing valuable tools for studying the role of GPR87 in vivo. These knockout mice have demonstrated protection against bleomycin-induced pulmonary fibrosis, confirming GPR87's role in fibrotic disease progression .

• Overexpression systems: Plasmid-based overexpression of GPR87 in cell lines with low endogenous expression has been used to study the functional consequences of GPR87 upregulation, particularly in the context of cancer stem cell properties .

• Pharmacological modulation: Lysophosphatidic acid (LPA) has been used as a ligand to stimulate GPR87 signaling, while pathway inhibitors like AG490 (a JAK2/STAT3 pathway inhibitor) have been used to block downstream signaling .

How are recombinant GPR87 antibodies produced and validated for research applications?

Recombinant GPR87 antibodies are produced through several steps:

• Expression in mammalian cells: Recombinant mouse antibodies against GPR87, such as TAB-002CT, are expressed in mammalian cells using chemically defined culture media to ensure proper folding and post-translational modifications .

• Purification: These antibodies are purified using affinity chromatography to achieve high purity (>95% as determined by SDS-PAGE and SEC-HPLC) .

• Validation through multiple methods:

  • SDS-PAGE analysis under both non-reduced and reduced conditions to confirm purity and integrity

  • SEC-HPLC (Size Exclusion Chromatography-High-Performance Liquid Chromatography) for purity assessment

  • ELISA analysis using Human GPR87 Full Length Protein as coating antigen

  • Dot Blot analysis with appropriate positive and negative controls

• Humanization: For therapeutic applications, mouse anti-GPR87 antibodies have been humanized to reduce the human anti-mouse antibody (HAMA) response. This process involves replacing mouse-specific regions while maintaining antigen specificity .

The available formats include:

• Full-length antibodies

• scFv (single-chain variable fragment) for applications requiring smaller binding molecules

• Fab (antigen-binding fragment) for specific research applications

These recombinant antibodies offer several advantages over traditional monoclonal antibodies, including increased sensitivity, confirmed specificity, high repeatability, excellent batch-to-batch consistency, sustainable supply, and animal-free production .

What are the major unresolved questions about GPR87 biology?

Despite significant progress in understanding GPR87, several key questions remain unresolved:

• Natural ligand controversy: While lysophosphatidic acid (LPA) has been used to stimulate GPR87 in experimental settings, the natural ligand of GPR87 remains controversial and not fully established . Identifying the definitive ligand would provide important insights into physiological regulation.

• Cell type-specific functions: While GPR87 is expressed in multiple cancer types and in specific cell populations in fibrotic lungs, the precise functions in different cellular contexts remain to be fully elucidated. For example, its role in the aberrant basaloid cells in IPF requires further investigation .

• Crosstalk with other signaling pathways: Although GPR87 has been linked to JAK2/STAT3, PI3K/mTOR, and TNF/NFκB pathways, the full extent of its signaling network, including potential crosstalk with other GPCR pathways, remains to be characterized .

• Regulation of GPR87 expression: While STAT3 has been identified as a transcriptional regulator of GPR87, and H3.3 as an activator for GPR87 transcription , the complete regulatory framework controlling GPR87 expression in different physiological and pathological contexts is not fully understood.

What emerging therapeutic strategies target GPR87?

Several promising therapeutic strategies targeting GPR87 are emerging:

• Near-infrared photoimmunotherapy (NIR-PIT): This approach combines humanized anti-GPR87 antibodies with photosensitizers to specifically destroy GPR87-expressing cancer cells when irradiated with near-infrared light. This method offers potential for localized treatment with minimal damage to surrounding normal tissues .

• RNA interference-based therapies: Adenoviral vectors expressing shRNAs targeting GPR87 (Ad-shGPR87) have shown efficacy in preclinical models, significantly inhibiting proliferation of GPR87-overexpressing cancer cells both in vitro and in vivo .

• Pathway inhibitors: Given GPR87's involvement in signaling through PI3K/mTOR and JAK2/STAT3 pathways, inhibitors of these pathways may offer therapeutic potential, particularly in combination with direct GPR87 targeting .

• Epithelial-specific targeting in pulmonary fibrosis: Recent findings on GPR87's role in basal and basaloid cells in pulmonary fibrosis suggest potential for epithelial-specific targeting as a therapeutic strategy in fibrotic lung diseases .

• Antibody-drug conjugates: The tumor-specific expression and cell surface location of GPR87 make it a potential target for antibody-drug conjugates, which could deliver cytotoxic payloads specifically to GPR87-expressing cancer cells .