After failure on docetaxel, which has been the standard first line therapy for patients with
      metastatic castration-resistant prostate cancer (mCRPC), several treatment options are
      currently available.

      Two of the treatment options are directed against the androgen receptor (AR), enzalutamide
      and abiraterone. A third option is cabazitaxel, a next generation taxane. No head-to-head
      comparisons have been done for these three therapies in second-line mCRPC and as of yet, the
      optimal choice is unknown.

      Resistance to the AR-targeted therapies is at least in part a consequence of signaling
      through constitutively active AR splice variants (AR-Vs). Because AR splice variants only
      occur after conversion to a castration-resistant tumor, and can be acquired during systemic
      therapy for mCRPC, analysis of the castration-naïve primary tumor is not informative in the
      setting of second-line treatment of mCRPC.

      Circulating tumor cells (CTCs) can be analyzed repetitively and in real-time. Recently, AR-V7
      messenger ribonucleid acid (mRNA) expression in CTCs was shown to be associated with lack of
      response to AR-targeted therapy (reference 1). AR-V7 mRNA expression does not seem to hinder
      response to cabazitaxel in our retrospective pilot study (reference 2) nor in two recently
      published retrospective studies (reference 3 and reference 4).

      Therefore we hypothesize that the mRNA expression of AR-V7 in CTCs assessed before start of
      second-line treatment for mCRPC does not affect prostate-specific antigen (PSA) response to
      cabazitaxel in patients who have progressed to docetaxel.

      Patients who are eligible to undergo second or third line treatment will be asked to undergo
      prescreening consisting of a CTC count and, in case ≥3 CTCs are detected, AR V7
      determination. Patients with ≥3 CTCs with AR-V7 expression will be asked to sign a second
      informed consent to enter the treatment study. In this study they will receive Cabazitaxel 25
      mg/m² every 3 weeks plus prednisone 10 mg daily, and undergo repeated blood sampling for
      biomarker sample collection.

      During the prescreening in all patients, 2 x 10 mL blood will be drawn for enumeration and
      isolation of CTCs. All patients with ≥3 CTCs with AR-V7 expression will be asked to sign
      consent for the treatment study.

      All patients included in the treatment study will be administrated cabazitaxel intravenously
      at a dose of 25 mg/m², during a one-hour infusion every 3 weeks, as well as continuous
      treatment with prednisone 5 mg orally twice daily, or 10 mg once daily. In the treatment
      study patients, an additional 2 x 10 mL blood will be drawn after the third cycle of
      treatment for CTC enumeration and isolation. An additional 10 mL blood will be drawn for
      storage of plasma at baseline and before start of every cycle (i.e., every 3 weeks) for
      analysis of cell-free DNA (cfDNA). Moreover, 4 x 5 mL blood (baseline; end of infusion, 2 and
      6 hours after end of the first cabazitaxel infusion) will be drawn for pharmacokinetic
      studies, in order to explore a cabazitaxel exposure effect relation.