RHPN1-AS1 Antibody

What is RHPN1-AS1 and how is it classified?

RHPN1-AS1 is a long non-coding RNA (lncRNA) that has been identified as a key regulatory molecule in multiple cancers. As a non-protein coding transcript, it functions primarily through RNA-RNA and RNA-protein interactions rather than through translation into protein. RHPN1-AS1 belongs to the antisense lncRNA category, transcribed from the opposite strand of the RHPN1 gene. Current research has established its critical role in modulating gene expression through mechanisms such as competing endogenous RNA (ceRNA) activity, whereby it can sponge microRNAs and prevent their binding to target mRNAs .

Which cancer types show altered RHPN1-AS1 expression?

RHPN1-AS1 has been found to be significantly upregulated in multiple cancer types. Based on current literature, elevated RHPN1-AS1 expression has been documented in retinoblastoma (RB), epithelial ovarian cancer (EOC), and breast cancer (BC). The upregulation pattern appears consistent across these malignancies, suggesting a common oncogenic role. In breast cancer specifically, analysis of The Cancer Genome Atlas (TCGA) dataset revealed significantly higher expression of RHPN1-AS1 in tumor tissues compared to non-tumor samples . Similarly, studies in epithelial ovarian cancer showed that RHPN1-AS1 was one of the most upregulated lncRNAs in EOC tissues compared to para-cancerous control tissues .

How does RHPN1-AS1 function as a competing endogenous RNA?

RHPN1-AS1 functions primarily as a competing endogenous RNA (ceRNA) by sponging specific microRNAs, thereby preventing these microRNAs from binding to their target mRNAs. This mechanism has been validated through multiple experimental approaches. In retinoblastoma, RHPN1-AS1 sponges miR-3133, counteracting miR-3133-mediated JAK2/STAT3 suppression . In epithelial ovarian cancer, RHPN1-AS1 acts as a ceRNA for miR-596, consequently increasing LETM1 expression and activating the FAK/PI3K/Akt signaling pathway . These interactions have been confirmed through RNA immunoprecipitation (RIP) assays, which showed that endogenous RHPN1-AS1 was enriched in Ago2-RIP, and dual-luciferase reporter assays that demonstrated direct binding between RHPN1-AS1 and its target microRNAs.

What regulatory networks are controlled by RHPN1-AS1 in cancer cells?

RHPN1-AS1 regulates several critical oncogenic pathways through its ceRNA activity. In retinoblastoma, the RHPN1-AS1/miR-3133/JAK2/STAT3 axis controls cell proliferation and apoptosis. Silencing RHPN1-AS1 increases proapoptotic factors (Bax and p53) while decreasing antiapoptotic factors (Bcl-2 and Survivin) . In epithelial ovarian cancer, RHPN1-AS1 regulates the miR-596/LETM1/FAK/PI3K/Akt pathway, promoting proliferation, migration, and invasion . In breast cancer, RHPN1-AS1 influences epithelial-to-mesenchymal transition (EMT), with its silencing resulting in decreased expression of EMT markers . These diverse regulatory networks highlight the multifaceted role of RHPN1-AS1 in cancer progression.

What mechanisms regulate RHPN1-AS1 expression itself?

Research has identified epigenetic mechanisms controlling RHPN1-AS1 expression. In epithelial ovarian cancer, N6-methyladenosine (m6A) modification improves the stability of RHPN1-AS1 methylation transcript by reducing RNA degradation, resulting in upregulation of RHPN1-AS1 . This post-transcriptional regulation demonstrates how RNA modifications can contribute to elevated RHPN1-AS1 levels in cancer cells. Understanding these regulatory mechanisms is essential for developing therapeutic strategies targeting RHPN1-AS1 expression.

What techniques are recommended for detecting and quantifying RHPN1-AS1?

Several complementary techniques are recommended for comprehensive RHPN1-AS1 analysis:

• Quantitative Real-Time PCR (qRT-PCR): The gold standard for quantifying RHPN1-AS1 expression, using primers designed to generate short RHPN1-AS1 amplicons (approximately 60 bp) to enhance quantitation efficiency and reliability, particularly when working with paraffin-embedded tissues where nucleic acids may be degraded .

• RNA Fluorescence In Situ Hybridization (FISH): Used to determine the subcellular localization of RHPN1-AS1. Studies have demonstrated that RHPN1-AS1 is primarily localized in the cytoplasm of cancer cells, which is consistent with its role as a ceRNA .

• Microarray Analysis: Useful for initial screening to identify differentially expressed lncRNAs, including RHPN1-AS1, between cancer and control tissues .

How can researchers effectively modulate RHPN1-AS1 expression in experimental models?

For functional studies, researchers can modulate RHPN1-AS1 expression through:

• RNA Interference: Short hairpin RNAs (shRNAs) targeting RHPN1-AS1 have been successfully used to silence its expression. Typically, shRNAs are synthesized and cloned into lentivirus plasmids (e.g., pcDNA-EF2-puromycin), followed by stable transfection and puromycin selection .

• Overexpression Systems: RHPN1-AS1 can be overexpressed using appropriate expression vectors. This approach is valuable for gain-of-function studies to validate the oncogenic effects of RHPN1-AS1 .

• CRISPR-Cas9 Gene Editing: Though not explicitly mentioned in the search results, CRISPR-Cas9 technology represents an advanced approach for manipulating RHPN1-AS1 expression at the genomic level.

What assays are used to validate RHPN1-AS1's interaction with microRNAs?

Multiple complementary techniques are employed to validate the RHPN1-AS1-microRNA interactions:

• Dual-Luciferase Reporter Assay: This assay confirms direct binding between RHPN1-AS1 and microRNAs. Wild-type (WT) and mutant (Mut) RHPN1-AS1 sequences are cloned into luciferase reporter vectors, and changes in luciferase activity upon microRNA overexpression are measured .

• RNA Immunoprecipitation (RIP) Assay: Ago2-RIP is used to demonstrate the enrichment of RHPN1-AS1 in RNA-induced silencing complexes (RISCs), providing evidence for its interaction with microRNAs within these complexes .

• Biotin-Labeled miRNA Pull-Down Assay: This technique directly captures RNA complexes containing the microRNA of interest, allowing for the identification of RHPN1-AS1 as an interacting partner .

How does RHPN1-AS1 affect cancer cell proliferation and apoptosis?

RHPN1-AS1 significantly impacts cancer cell proliferation and apoptosis through multiple mechanisms:

• Proliferation Enhancement: In both retinoblastoma and epithelial ovarian cancer, RHPN1-AS1 promotes cell proliferation. CCK-8 and colony formation assays have demonstrated that RHPN1-AS1 knockdown significantly inhibits cancer cell viability and clonogenic ability, while its overexpression enhances these processes .

• Apoptosis Regulation: In retinoblastoma, silencing RHPN1-AS1 promotes apoptosis by increasing proapoptotic factors (Bax and p53) and decreasing antiapoptotic factors (Bcl-2 and Survivin) . This indicates that RHPN1-AS1 normally functions to suppress apoptotic pathways in cancer cells.

• Cell Cycle Control: Though not explicitly detailed in the search results, the effects on proliferation suggest RHPN1-AS1 may also influence cell cycle progression.

What role does RHPN1-AS1 play in cancer cell migration and invasion?

RHPN1-AS1 promotes cancer cell migration and invasion capabilities:

• Migration Enhancement: Wound healing assays have shown that RHPN1-AS1 depletion represses cancer cell migration, while its overexpression enhances this process. In epithelial ovarian cancer specifically, RHPN1-AS1 silencing significantly reduced the migratory capacity of HEY and ES-2 cancer cell lines .

• Invasion Promotion: Transwell assays have demonstrated that RHPN1-AS1 knockdown inhibits cancer cell invasion, while its overexpression promotes invasive capabilities .

• EMT Regulation: In breast cancer, RHPN1-AS1 silencing is associated with decreased expression of epithelial-to-mesenchymal transition (EMT) markers, suggesting that RHPN1-AS1 promotes EMT, a critical process for cancer metastasis .

How does RHPN1-AS1 influence tumor growth in vivo?

In vivo studies confirm RHPN1-AS1's role in promoting tumor growth:

• Xenograft Models: Studies using xenograft mouse models have demonstrated that RHPN1-AS1 silencing significantly reduces tumor growth in vivo, validating the in vitro findings on cell proliferation .

• Metastatic Potential: The association between high RHPN1-AS1 expression and distant metastasis in clinical samples suggests its role in promoting metastasis in vivo .

Can RHPN1-AS1 serve as a reliable prognostic biomarker?

Current evidence strongly supports RHPN1-AS1's potential as a prognostic biomarker:

What therapeutic potential does targeting RHPN1-AS1 offer?

The therapeutic potential of targeting RHPN1-AS1 is supported by several lines of evidence:

How does RHPN1-AS1 expression vary across cancer subtypes?

RHPN1-AS1 shows differential expression across cancer subtypes:

• Breast Cancer Molecular Subtypes: RHPN1-AS1 is overexpressed in HER2-enriched and Luminal B (LumB) subtypes compared to Luminal A (LumA) and Basal subtypes . This subtype-specific expression pattern may reflect distinct molecular mechanisms and could inform personalized therapeutic approaches.

• Cross-Cancer Analysis: TCGA data analysis suggests variable RHPN1-AS1 expression across different cancer types, including Bladder Urothelial Carcinoma (BLCA), Cholangiocarcinoma (CHOL), Colon cancer (COAD), and others , indicating cancer-specific regulatory mechanisms.

What are the current technical challenges in studying RHPN1-AS1?

Several technical challenges must be addressed when studying RHPN1-AS1:

• RNA Degradation in Clinical Samples: Nucleic acids are extensively degraded in paraffin-embedded tissues, necessitating specialized approaches such as designing primers for short amplicons (approximately 60 bp) to enhance quantitation efficiency and reliability .

• Complex Regulatory Networks: RHPN1-AS1 participates in intricate regulatory networks involving multiple microRNAs and downstream targets, requiring comprehensive approaches to fully elucidate its function.

• Context-Dependent Functions: RHPN1-AS1 may have different roles in different cancer types and even across cancer subtypes, as seen in breast cancer molecular subtypes , necessitating cancer-specific and subtype-specific investigations.

How can conflicting data about RHPN1-AS1 across cancer types be reconciled?

Reconciling potentially conflicting data requires:

• Molecular Context Analysis: Identifying cancer-specific molecular partners of RHPN1-AS1, such as different microRNA targets in different cancers (miR-3133 in retinoblastoma versus miR-596 in epithelial ovarian cancer ).

• Pathway Integration: Determining whether seemingly distinct pathways regulated by RHPN1-AS1 in different cancers converge on common biological processes, such as cell proliferation and apoptosis.

• Comprehensive Multi-Omics Approaches: Integrating transcriptomic, proteomic, and epigenomic data to build a more complete picture of RHPN1-AS1 function across cancer types.

What are emerging directions in RHPN1-AS1 research?

Emerging research directions include:

• Therapeutic Targeting Strategies: Developing methods to specifically target RHPN1-AS1, such as antisense oligonucleotides or RNA interference-based therapeutics, and evaluating their efficacy in preclinical models.

• Combination Therapies: Investigating potential synergistic effects between RHPN1-AS1 targeting and conventional cancer treatments, such as chemotherapy or targeted therapies.

• RNA Modifications: Further exploring the role of N6-methyladenosine (m6A) and other RNA modifications in regulating RHPN1-AS1 stability and function, as highlighted in epithelial ovarian cancer research .