Abstract 5913: Genetic landscape changes in metastatic prostate cancer after sequential steps of androgen targeted therapy
Authors: Raittinen Paavo V. H. , Pennanen Pasi , Syvälä Heimo , Bläuer Merja , Ikonen Niina , Nätkin Reetta , Nykter Matti , Tammela Teuvo , Ilmonen Pauliina , Murtola Teemu T. 
Published: September 2020

Abstract
Metastatic prostate cancer is non-curable fatal disease requiring active treatment. Prostate cancer cell proliferation requires androgen hormones mediated by androgen receptor (AR). Typical metastatic prostate cancer treatment exploits this dependency by systemic androgen deprivation and AR inhibition. While androgen deprivation therapy (ADT) effectively halts cancer cell proliferation, the cells adapt to the limited androgen supply over time by up-regulating AR, making ADT ineffective and creating castration-resistant prostate cancer (CRPC). Androgen receptor signaling inhibitors (ARSIs) such as enzalutamide remain effective even after CRPC development. These drugs have effectively replaced usage of AR inhibitors such as bicalutamide in clinical management of CRPC. However, over time cancer cells undergo further genetic transformations leading to ARSI resistance. Some genetic adaptation mechanisms behind this are known, e.g., AR over-expression and alternative splicing of AR. However, detailed genetic landscape shift is unknown. Moreover, it is unknown whether different AR inhibitors or their sequencing may affect cancer cell genotype and phenotype similarly.
We compare genetic profiles of two different multiresistant VCaP cell populations achieved by long-term cell culture. Two treatment paths are: A. ADT, bicalutamide (BIC), and enzalutamide (ENZA) sequentially; and B. ADT and ENZA sequentially, without bicalutamide. Each consequent treatment is continued as long as it takes for the cells to regain cellular proliferation despite the treatment in order to induce adaptive genetic transformations required for treatment resistance. Once cell proliferation resumes, the obtained resistant cell population is sequenced for mRNA expression levels and splice variant distribution. The genetic profiles of treatment path A and B are compared by analyzing differentially expressed genes and splice variant expression distributions. At the end of the treatment paths, both cell populations are exposed to variable doses of docetaxel to investigate whether ADT- and ARSI-resistant genetic profile affects vulnerability to subsequent chemotherapy.
There was no difference in AR expression or AR splice variant distribution between the treatment paths A and B. AR splicing variants are observed after ARSI resistance. However, multiple other cancer-related genes are differentially expressed between the treatment paths, including FOS, CCND1, IGF1R, and AGR2 which were down-regulated by BIC and heavily up-regulated by ENZA. These genetic alterations also translate into proteomic changes. Remarkably, cells treated with BIC prior to ENZA responded significantly better to docetaxel than ENZA-only treated cells (100 nM dose treatment left 73.75% and 64.55 % cells dead, respectively).
ENZA treatment drives genetic landscape shift in cancer cells when achieving treatment resistance. However, this differs by treatment sequencing; observed genetic transformations differ by whether ENZA treatment was preceded by BIC exposure. Importantly, the differences in the genetic profile lead to greater chemoresistance in cells not treated with BIC before ENZA as compared to cells receiving both treatments. This observation may have implications in optimal sequencing of treatment in CRPC.
Citation Format: Paavo V. H. Raittinen, Pasi Pennanen, Heimo Syvälä, Merja Bläuer, Niina Ikonen, Reetta Nätkin, Matti Nykter, Teuvo Tammela, Pauliina Ilmonen, Teemu T. Murtola. Genetic landscape changes in metastatic prostate cancer after sequential steps of androgen targeted therapy [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 5913.