This study established a recommended phase II dose for PR-104 of 770 mg/m2 for combination with standard clinical doses of docetaxel (60 to 75 mg/m2)
, both given on day one with prophylactic G-CSF on day two of a three-week treatment cycle. At the recommended phase II dose, PR-104-docetaxel-G-CSF combination therapy appeared to be adequately tolerated without any DLT in a cohort of 12 patients treated at this dose-level. In contrast, two of three patients experienced DLT on cycle one of PR-104-docetaxel-G-CSF combination treatment at the next highest dose-level above the recommended phase II dose (PR-104 1100 mg/m2). At the recommended phase II dose-level, clinical benefit was apparent in five of a cohort of 12 patients either as partial tumour responses in three patients or as stable disease maintained for at least six treatment cycles (18 weeks) in a further two patients. It was not possible from our study to determine what contribution PR-104 may have made to this apparent clinical benefit relative to that made by docetaxel. However, the current study has provided a starting dose for phase II trials to address whether PR-104 enhances the therapeutic efficacy of docetaxel and to further evaluate the clinical safety of PR-104-docetaxel-G-CSF combination therapy.
Our study also found that combining PR-104 with gemcitabine or docetaxel in the absence of prophylactic G-CSF was not clinically feasible due to severe myelotoxicity. With the PR-104-gemcitabine combination, dose-limiting thrombocytopenia was encountered in two of three patients at the starting dose-level (275 mg/m2). Prophylactic G-CSF was not added to the PR-104-gemcitabine study group and the combination was not evaluated further due to thrombocytopenia being dose-limiting. With the PR-104-docetaxel combination given without G-CSF, dose-limiting febrile neutropenia occurred in three of six patients treated at or below the starting dose-level (400 mg/m2). The PR104-docetaxel combination was evaluated further with the addition of prophylactic G-CSF as dose-limiting neutropenia prohibited dose escalation of PR-104 without haematological growth factor support.
The unexpectedly severe and dose-limiting myelotoxicity encountered in this trial of PR-104-based combination chemotherapy appeared to have been due to a pharmacodynamic interaction, rather than a pharmacokinetic interaction, between PR-104 and gemcitabine or docetaxel. The plasma pharmacokinetics of docetaxel, gemcitabine, its major metabolite (difluorodeoxyuridine) and PR-104 and its metabolites (PR-104A, PR-104G, PR-104H, PR-104M and PR-104S1) determined in patients treated with PR-104 combined with gemcitabine or docetaxel appeared similar to published reports of the pharmacokinetics of PR-104
[7, 8], gemcitabine
[22, 23] or docetaxel
[19–21] given alone. Gemcitabine and docetaxel are known to cause blood cytopenias as single agents at standard clinical doses, presumably due to their antiproliferative effects against bone marrow blood progenitor cells occurring as a result of their main pharmacological actions that involve the inhibition of DNA synthesis in S-phase cells and disruption of microtubule assembly in the mitotic spindle of M-phase cells, respectively
[9, 10]. PR-104 is also myelotoxic as a single agent in clinical trials
[7, 8], via mechanisms that are currently unclear but that may involve its metabolic activation by hypoxia or AKR1C3 in normal bone marrow (personal communications, J. Down and K. Parmar) followed by DNA cross-linking and cytotoxicity to blood progenitor cells by mechanisms analogous to other myelosuppressive nitrogen mustards
. These considerations point to the possibility of the unexpectedly severe and dose-limiting blood cytopenias of the PR-104-chemotherapy combinations evaluated in this trial having been due to the overlapping myelotoxicity of PR-104, docetaxel and gemcitabine.
Dose-limiting myelotoxicity may have restricted PR-104 dose-escalation and PR-104A systemic exposure in this clinical study. When PR-104 was combined with gemcitabine or docetaxel without prophylactic G-CSF, for example, the MTD for PR-104 was less than 200 mg/m2 and PR-104A AUC values were ≤ 3 μg*hr/ml. Addition of prophylactic G-CSF to the combination of PR-104 and docetaxel 60 to 75 mg/m2 permitted escalation of PR-104 to a dose of 770 mg/m2 that achieved PR-104A AUC values ranging from about 5 to 20 μg*hr/ml. These PR-104A AUC values achieved at the recommended phase II dose for the PR-104-docetaxel-G-CSF combination were similar to the values from earlier clinical pharmacokinetic studies of PR-104 at this dose-level but lower than those achieved at its single agent MTD in phase Ia studies
[7, 8]. In contrast, mice appear to be able to tolerate both higher PR-104A systemic exposure, in the order of 50 μg*hr/ml
, and the combination of PR-104 with gemcitabine or docetaxel at doses that achieved significant preclinical antitumour activity
. Furthermore, in human tumour xenograft murine models, the therapeutic antitumour activity of PR-104 alone or in combination with gemcitabine or docetaxel appears to be associated with doses
 that achieve systemic exposures to PR-104A that were higher than those achieved by most patients treated in the current clinical study.
This was one of the first studies that we are aware of that incorporated hypoxia imaging with F MISO or an equivalent PET imaging agent into a multicentre early-phase trial of a novel chemotherapy combination in a broad oncology patient population. Previously, hypoxia imaging in therapeutic trials has been limited to only a few studies of mainly chemo-radiation protocols that were often restricted to head and neck cancer
. Several issues related to our experience are worthy of comment. Firstly, hypoxia imaging was successfully carried out in less than half of the study patients at baseline and very few scans were obtained post-treatment, pointing to issues with its feasibility in the context of multicentre early-phase oncology trials. Tumour hypoxia was detected in most but not all subjects undergoing FMISO scans and across a range of tumour types, suggesting that tumour hypoxia is very common among a broad phase I oncology patient population and unrestricted to any particular tumour site or specific histopathological diagnosis. No correlation was apparent in our study between the presence or absence of tumour hypoxia at baseline and subsequent tumour response to PR-104-based combination chemotherapy. However, the small sample size, heterogeneous patient population, range of PR-104 dose-levels and incomplete data-set for FMISO scans in our study may have contributed to the apparent lack of correlation between tumour hypoxia and therapeutic outcome from PR-104-combination chemotherapy. Further studies of hypoxia imaging in early-phase multicentre oncology trials are required to further evaluate its predictive value and feasibility.
It is interesting to compare the results of the current study with those recently reported for TH-302
[27–29], a hypoxia-activated prodrug of the cytotoxin bromoisophosphoramide mustard. Like PR-104, the mechanism of action of TH-302 involves its activation in hypoxia, via NADPH-cytochrome P450 oxidoreductases and other one-electron reductases, into reactive mustard species that crosslink DNA. In phase Ib and II clinical trials, the dose-limiting toxicities of TH-302 combined with gemcitabine and docetaxel primarily involved the haematological system, similar to PR-104. However, combining TH-302 with docetaxel and gemcitabine appeared not to require G-CSF support and increased tumour response rates and progression free survival, particularly in patients with advanced pancreatic cancer. These findings are very promising and further investigations of TH-302 and other hypoxia-activated prodrugs combined with conventional chemotherapy are awaited with interest.