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Updated: Mar 28, 2026
Using RNA-sequencing to Detect Novel Splice Variants Related to Drug Resistance in In Vitro Cancer Models
Published on: December 9, 2016
Lin28 induces resistance to anti-androgens via promotion of AR splice variant generation
Ramakumar Tummala1, Nagalakshmi Nadiminty1, Wei Lou1
1Department of Urology, University of California at Davis, Sacramento, California.
Background:
Prostate cancer (PCa) is androgen-dependent initially and progresses to a castration-resistant state after androgen deprivation therapy. Treatment options for castration-resistant PCa include the potent second-generation anti-androgen enzalutamide or CYP17A1 inhibitor abiraterone. Recent clinical observations point to the development of resistance to these therapies which may be mediated by constitutively active alternative splice variants of the androgen receptor (AR).
Methods:
Sensitivity of LNCaP cells overexpressing Lin28 (LN-Lin28) to enzalutamide, abiraterone, or bicalutamide was compared to that of control LN-neo cells using cell growth assays, proliferation assays using MTT, anchorage-dependent clonogenic ability assays and soft agar assays. Ability of LN-Lin28 cells to maintain AR activation after treatment with enzalutamide, abiraterone, or bicalutamide was tested using immunofluorescence, Western blotting, ChIP assays, and qRT-PCR. Importance of Lin28 in enzalutamide resistance was assessed by the downregulation of Lin28 expression in C4-2B and 22Rv1 cells chronically treated with enzalutamide. Requirement for sustained AR signaling in LN-Lin28 cells was examined by the downregulation of either full length AR or AR-V7 using siRNA.
Results:
We show that Lin28 promotes the development of resistance to currently used targeted therapeutics by enhancing the expression of AR splice variants such as AR-V7. PCa cells overexpressing Lin28 exhibit resistance to treatment with enzalutamide, abiraterone, or bicalutamide. Downregulation of Lin28 resensitizes enzalutamide-resistant PCa cells to enzalutamide treatment. We also show that the upregulation of splicing factors such as hnRNPA1 by Lin28 may mediate the enhanced generation of AR splice variants in Lin28-expressing cells.
Conclusions:
Our findings suggest that Lin28 plays a key role in the acquisition of resistance to AR-targeted therapies by PCa cells and establish the importance of Lin28 in PCa progression.
Insights
Lin28 promotes resistance to prostate cancer therapies by increasing androgen receptor (AR) splice variants like AR-V7. Reducing Lin28 can resensitize resistant cancer cells to treatments such as enzalutamide.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Prostate cancer (PCa) is initially androgen-dependent but progresses to castration-resistant PCa (CRPC) after androgen deprivation therapy.
- CRPC treatments include enzalutamide and abiraterone, but resistance can develop.
- Resistance may involve constitutively active androgen receptor (AR) splice variants.
Purpose of the Study:
- To investigate the role of Lin28 in the development of resistance to AR-targeted therapies in prostate cancer.
- To determine if Lin28 overexpression contributes to resistance against enzalutamide, abiraterone, or bicalutamide.
Main Methods:
- Compared sensitivity of Lin28-overexpressing LNCaP cells (LN-Lin28) versus control cells (LN-neo) to anti-androgens using cell growth, proliferation, and clonogenic assays.
- Assessed AR activation, AR splice variant expression (e.g., AR-V7), and splicing factor levels (e.g., hnRNPA1) via immunofluorescence, Western blotting, ChIP, and qRT-PCR.
- Evaluated Lin28's importance by downregulating its expression in resistant cell lines and examining AR signaling pathway dependence.
Main Results:
- Lin28 overexpression confers resistance to enzalutamide, abiraterone, and bicalutamide in PCa cells.
- Lin28 enhances the expression of AR splice variants, including AR-V7, contributing to therapeutic resistance.
- Downregulating Lin28 resensitizes enzalutamide-resistant PCa cells to enzalutamide treatment.
- Lin28 upregulates splicing factors like hnRNPA1, potentially mediating increased AR splice variant generation.
Conclusions:
- Lin28 plays a critical role in the acquisition of resistance to AR-targeted therapies in prostate cancer.
- Lin28 is important in prostate cancer progression and represents a potential therapeutic target for overcoming treatment resistance.
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