Search

The switch/sucrose non-fermentable (SWI/SNF) complex has a crucial role in chromatin remodelling1 and is altered in over 20% of cancers2,3. Here we developed a proteolysis-targeting chimera (PROTAC) degrader of the SWI/SNF ATPase subunits, SMARCA2 and SMARCA4, called AU-15330. Androgen receptor (AR)+ forkhead box A1 (FOXA1)+ prostate cancer cells are exquisitely sensitive to dual SMARCA2 and SMARCA4 degradation relative to normal and other cancer cell lines. SWI/SNF ATPase degradation rapidly compacts cis-regulatory elements bound by transcription factors that drive prostate cancer cell proliferation, namely AR, FOXA1, ERG and MYC, which dislodges them from chromatin, disables their core enhancer circuitry, and abolishes the downstream oncogenic gene programs. SWI/SNF ATPase degradation also disrupts super-enhancer and promoter looping interactions that wire supra-physiologic expression of the AR, FOXA1 and MYC oncogenes themselves. AU-15330 induces potent inhibition of tumour growth in xenograft models of prostate cancer and synergizes with the AR antagonist enzalutamide, even inducing disease remission in castration-resistant prostate cancer (CRPC) models without toxicity. Thus, impeding SWI/SNF-mediated enhancer accessibility represents a promising therapeutic approach for enhancer-addicted cancers., PROTAC degrader–induced SWI/SNF inactivation abolishes DNA accessibility at enhancer elements of oncogenes and also tempers supra-physiologic expression of driver transcription factors, resulting in potent inhibition of tumour growth in mouse models.

Metastatic neuroendocrine prostate cancer (NEPC) is a highly aggressive disease, whose incidence is rising. Long noncoding RNAs (lncRNAs) represent a large family of disease‐ and tissue‐specific transcripts, most of which are still functionally uncharacterized. Thus, we set out to identify the highly conserved lncRNAs that play a central role in NEPC pathogenesis. To this end, we performed transcriptomic analyses of donor‐matched patient‐derived xenograft models (PDXs) with immunohistologic features of prostate adenocarcinoma (AR+/PSA+) or NEPC (AR−/SYN+/CHGA+) and through differential expression analyses identified lncRNAs that were upregulated upon neuroendocrine transdifferentiation. These genes were prioritized for functional assessment based on the level of conservation in vertebrates. Here, LINC00261 emerged as the top gene with over 3229‐fold upregulation in NEPC. Consistently, LINC00261 expression was significantly upregulated in NEPC specimens in multiple patient cohorts. Knockdown of LINC00261 in PC‐3 cells dramatically attenuated its proliferative and metastatic abilities, which are explained by parallel downregulation of CBX2 and FOXA2 through distinct molecular mechanisms. In the cell cytoplasm, LINC00261 binds to and sequesters miR‐8485 from targeting the CBX2 mRNA, while inside the nucleus, LINC00261 functions as a transcriptional scaffold to induce SMAD‐driven expression of the FOXA2 gene. For the first time, these results demonstrate hyperactivation of the LINC00261‐CBX2‐FOXA2 axes in NEPC to drive proliferation and metastasis, and that LINC00261 may be utilized as a therapeutic target and a biomarker for this incurable disease., Neuroendocrine prostate cancer (NEPC) is a highly aggressive disease with high incidence of metastasis. Here, we identified LINC00261 among the top upregulated transcripts in NEPC. Inside the nucleus, LINC00261 promoted anchorage‐independent growth and metastatic gene programs by recruiting the SMAD2/3 transcriptional machinery to FOXA2 cis‐regulatory elements. Cytoplasmic LINC00261 blocked the binding of miR‐8485 to CBX2 and induced a CBX2‐driven proliferative gene program. Our study demonstrates the role of LINC00261‐CBX2‐FOXA2 axis in proliferation and metastasis of NEPC.

T cell receptor (TCR) signal strength is a key determinant of T cell responses. We developed a cancer mouse model in which tumor-specific CD8 T cells (TST cells) encounter tumor antigens with varying TCR signal strength. High-signal-strength interactions caused TST cells to up-regulate inhibitory receptors (IRs), lose effector function, and establish a dysfunction-associated molecular program. TST cells undergoing low-signal-strength interactions also up-regulated IRs, including PD1, but retained a cell-intrinsic functional state. Surprisingly, neither high- nor low-signal-strength interactions led to tumor control in vivo, revealing two distinct mechanisms by which PD1hi TST cells permit tumor escape; high signal strength drives dysfunction, while low signal strength results in functional inertness, where the signal strength is too low to mediate effective cancer cell killing by functional TST cells. CRISPR-Cas9–mediated fine-tuning of signal strength to an intermediate range improved anti-tumor activity in vivo. Our study defines the role of TCR signal strength in TST cell function, with important implications for T cell–based cancer immunotherapies.

In mammalian cells, DNA is wrapped around histone octamers (collectively referred to as nucleosomes) which form a physical barrier to all DNA-based processes. The switch/sucrose non-fermentable (SWI/SNF) is a multi-subunit chromatin remodeling complex that uses energy from ATP hydrolysis to reposition or eject nucleosomes at non-coding regulatory elements, thereby enabling access to the underlying DNA for transcriptional activation. Notably, the SWI/SNF complex plays a crucial role in chromatin remodeling and is recurrently altered in over 20% of human cancers, with the revised complex in cancer cells enabling central oncogenic gene programs. Yet, no studies have assessed the therapeutic efficacy of complete SWI/SNF inactivation across human cancers. Here, we developed a proteolysis targeting chimera (PROTAC) degrader of ATPase subunits of the SWI/SNF complex, SMARCA2 and SMARCA4. In a panel with over 90 normal and cancer cell lines from 18 different lineages, we found MYC-driven multiple myeloma and androgen receptor (AR)/forkhead box A1 (FOXA1)-positive prostate and breast cancers to be preferentially sensitive to dual SMARCA2 and SMARCA4 degradation relative to benign prostate as well as other cancer cell lines, including cancer cell lines with inactivating SMARCA4 mutations. We found complete SWI/SNF ATPase degradation to instantaneously compact the cis-regulatory elements that are bound and activated by transcription factors that drive cancer proliferation, namely MYC, IRF4, TCF3, AR, FOXA1, and ERG. This ensued in parallel untethering of these oncogenic drivers from the chromatin, with subsequent chemical decommissioning of their core enhancer circuitry and attenuation of downstream gene programs. Furthermore, using chromatin conformation assays we found SWI/SNF inactivation to disrupt super-enhancer and promoter DNA looping interactions that wire supra-physiologic expression of the MYC, AR, ERG, IRF4, and TCF3 oncogenes themselves, thereby tempering their expression in cancer cells. Treatment with the SMARCA2/4 degrader alone induced potent inhibition of tumor growth in cell line-derived xenograft models of multiple myeloma, as well as prostate cancer, and synergized with AR antagonists, inducing disease remission in several drug-resistant disease models. Notably, no major toxicities were seen in mice upon prolonged treatment with the SMARCA2/4 degrader, including no indications of thrombocytopenia, gastrointestinal goblet cell depletion, or germ cell degeneration—all being major toxicities associated with the BRD4-targeting therapeutics. To our knowledge, this study is the first preclinical proof of concept that targeted obstruction of chromatin accessibility at non-coding regulatory elements can be a potent therapeutic strategy in enhancer-addicted tumors, warranting the safety and efficacy assessments of SWI/SNF inhibitors and degraders in human clinical trials. Citation Format: Abhijit Parolia, Lanbo Xiao, Yuanyuan Qiao, Pushpinder Bawa, Sanjana Eyunni, Eleanor Young, Rahul Mannan, Sandra E. Carson, Yu Chang, Yuping Zhang, James George, Mustapha Jaber, Fengyun Su, Rui Wang, Sanjita Sasmal, Leena Khare, Subhendu Mukerjee, Chandrasekhar AbbinenI, Kiran Aithal, Xuhong Cao, Yuzhuo Wang, Susanta Samajdar, Murali Ramachandra, Arul M. Chinnaiyan. Targeting SWI/SNF ATPases in enhancer-addicted human cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3592.

The switch/sucrose non-fermentable (SWI/SNF) complex plays a crucial role in chromatin remodeling and is recurrently altered in over 20% of human cancers. Here, we developed a proteolysis targeting chimera (PROTAC) degrader of ATPase subunits of the SWI/SNF complex, SMARCA2 and SMARCA4. Intriguingly, we found androgen receptor (AR)/forkhead box A1 (FOXA1)-positive prostate cancer to be exquisitely sensitive to dual SMARCA2 and SMARCA4 degradation relative to benign prostate as well as other cancer cell lines, including those with inactivating SMARCA4 mutations. Mechanistically, SWI/SNF inhibition rapidly compacts the cis-regulatory elements that are bound and activated by transcription factors that drive cancer proliferation, namely AR, FOXA1, ERG, and MYC. This ensues in chromatin untethering of these oncogenic drivers, chemical decommissioning of their core enhancer circuitry, and attenuation of downstream gene programs. Furthermore, we found SWI/SNF inhibition to disrupt super-enhancer and promoter DNA looping interactions that wire supra-physiologic expression of the AR, FOXA1, and MYC oncogenes, thereby tempering their expression in cancer cells. Monotherapy with the SMARCA2/4 degrader induced potent inhibition of tumor growth in cell line-derived xenograft models of prostate cancer and remarkably synergized with AR antagonists, inducing disease remission in models of castration-resistant prostate cancer. We also found the combinatorial treatment to significantly inhibit the growth of enzalutamide resistant disease using in vitro as well as patient-derived xenograft models. Notably, no major toxicities were seen in mice upon prolonged treatment with the SMARCA2/4 degrader, including no indications of thrombocytopenia, gastrointestinal goblet cell depletion, or germ cell degeneration. Taken together, these results suggest that impeding enhancer accessibility through SWI/SNF ATPase inactivation represents a novel therapeutic approach in enhancer addicted human cancers. Citation Format: Lanbo Xiao, Abhijit Parolia, Yuanyuan Qiao, Pushpinder Bawa Pushpinder, Sanjana Eyunni, Rahul Mannan, Sandra E. Carson, Yu Chang, Xiaoju Wang, Yuping Zhang, Josh Vo, Steven Kregel, Stephanie A. Simko, Andrew D. Delekta, Mustapha Jaber, Heng Zheng, Ingrid Apel, Lisa McMurry, Fengyun Su, Rui Wang, Sylvia Wang, Sanjita Sasmal, Leena K. Satyam, Subhendu Mukherjee, Chandrasekhar AbbinenI, Kiran Aithal, Mital S. Bhakta, Jay Ghurye, Xuhong Cao, Nora M. Navone, Alexey Nesvizhskii, Rohit Mehra, Ulka Vaishampayan, Marco Blanchette, Yuzhuo Wang, Susanta Samajdar, Murali Ramachandra, Arul M. Chinnaiyan. Targeting SWI/SNF ATPases in enhancer-addicted prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5469.

Forkhead box A1 (FOXA1) is a pioneer transcription factor that is essential for the normal development of several endoderm-derived organs, including the prostate gland. FOXA1 independently binds to and de-compacts condensed chromatin to reveal the binding sites of partnering nuclear hormone receptors. In prostate luminal epithelial cells, FOXA1 delimits tissue-specific enhancers that are transcriptionally activated by the androgen receptor (AR). Thus, it is intriguing that FOXA1 is frequently mutated in hormone-receptor-driven prostate, breast, bladder and salivary-gland tumors. In a recent study, leveraging an aggregate cohort of 1,546 prostate cancers, we classified FOXA1 alterations into three novel structural classes that diverge in clinical incidence and genetic co-alteration profiles, with a collective prevalence of 35%. Class1 mutations comprise of missense and in-frame indels within the Wing2 secondary structure and originate in early prostate cancer without other driver alterations in genes such as ETS or SPOP. Contrarily, class2 mutations comprise of frameshift mutations which truncate the C-terminal end of the protein and are clonally acquired in the metastatic disease. Finally, class-3 genomic rearrangements comprise of duplications and translocations that reposition a conserved enhancer element to drive overexpression of FOXA1 or other oncogenes. Here, we mechanistically expand upon the neomorphic class2 mutants and show that they centrally enable prostate cancer metastasis. Remarkably, we found truncated class2 mutants to entirely displace their wild-type counterpart from the chromatin - a characteristic we termed as “cistromic dominance” - due to higher DNA affinity. Through proteomic assays, we identified the C-terminal regulatory domain of wild-type FOXA1 to interact with and recruit a bonafide transcriptional WNT-repressor called TLE3 to the chromatin. Notably, class2 mutants had lost this interaction and dominantly displace TLE3 from chromatin to de-repress WNT signaling. Accordingly, we found CRISPR-engineered prostate cancer cells harboring class2 FOXA1 mutations to show higher invasive and metastatic ability both in vitro and in vivo. Also, the increase in the metastatic ability of class2-mutant cells was completely reversed upon treatment with a WNT inhibitor. Overall, our study reaffirms the central role of FOXA1 in mediating AR-driven oncogenesis and provides mechanistic insights into how distinct classes of FOXA1 alterations drive initiation or metastatic progression of prostate cancer. These results reveal a novel FOXA1-TLE3-WNT signaling axis and rationalize its therapeutic co-inhibition in advanced prostate cancer. Citation Format: Abhijit Parolia, Marcin Cieslik, Sandra Carson, Shih-Chun Chu, Lanbo Xiao, Takahiro Ouchi, Mustapha Jaber, Yuping Zhang, Pankaj Vats, Xuhong Cao, Arul Chinnaiyan. Cistromically-dominant class2 mutants of FOXA1 de-repress WNT signaling to drive prostate cancer metastasis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3437.