intake of afatinib. Plasma values of afatinib after drug administration, obtained on day 8, increased during the sampling period to 26.7 ng/ml (gMean [range 0.590 ng/ml to 127 ng/ml]) at 3 hours after intake. Trough values were highly variable at a given time SGI-1776 point (gCV 73.6–92.5%), but (overall) remained unchanged during the trial period (Table IX). Plasma levels of BIBF 1120, obtained on day 15, increased during the sampling period to 20.6 ng/ml (gMean [range 0.841 ng/ml to 127 ng/ml]) at 3 hours after intake. Trough values at 12 hours after the last intake of the first 7-day BIBF 1120 dosing period (day 
were highly variable (gCV 118%), with a gMean of 21.4 ng/ml (range 3.93 ng/ml to 124 ng/ml). BIBF 1120 plasma concentrations before the restart of BIBF 1120 intake after 7 days of treatment with afatinib were consistently negligible, with values at or below the
lower limit of quantification for most patients. This trial demonstrates the feasibility of a sequential combination regimen of small-molecule TKIs, an irreversible EGFR/HER2 inhibitor afatinib and a triple angiokinase inhibitor BIBF 1120. No unexpected buy KU-0063794 drug-related toxicities were observed and the anticipated Gl side-effects were well managed. Only two patients discontinued therapy due to apparent intolerance to the regimen: one patient discontinued for diarrhoea that had already been present at baseline; the other suffered from asthenia in the context of tumour progression. Several patients showed increases in liver enzymes that might not be attributed to progression of CRC manifestations, although most of the patients had extensive, progressing liver metastases. The increase of liver enzymes occurred early, and subsequently resolved to at least maximum CTCAE grade 1 levels after dose interruption and/or reduction.
Only some patients had dose reductions of BIBF 1120 to 150 mg twice daily. purchase KU-0063794 Reversible liver enzyme elevations have been observed in BIBF 1120 single-agent studies (41, 42), but not for afatinib. The absorption kinetics obtained for both drugs after 7-day pre-treatment with the respective other combination partner closely resembled those obtained in phase I trials of each drug alone (41, 50). Similarly, trough levels, as well as nadir levels for BIBF 1120, remained unchanged throughout the trial period. These data suggest that PK drug–drug interactions did not occur between these drugs. This first trial combining BIBF 1120 and afatinib was intended as a first step towards a more intense treatment regimen. In particular, angiogenesis inhibition may need to be maintained continuously with the use of BIBF 1120. Ongoing preclinical studies in colon cancer models suggest that continuous exposure to BIBF 1120 will be needed for optimal activity (57). It should be noted that many patients in this trial presented with advanced disease with palliative treatment intent; almost all patients had already received and failed several lines of prior targeted agents. Antibodies were part of the regimens preceding inclusion into this study, and had been discontinued due to progressive disease.
The lack of clinical efficacy in this study in heavily pretreated patients may not be conclusive for the sequential treatment approach of combining an EGFR/HER2 inhibitor with an angiogenesis inhibitor. Of note, two patients remained progression-free for a order KU-0063794 relatively long period of time and had a time to progression that exceeded that observed during the immediately preceding treatment line. Recent evidence links failure to respond to EGFR antibodies to mutations in the downstream effector pathways (58-61). Whether these may also arise during treatment with EGFR inhibitors and preclude efficacy of retreatment with another EGFR inhibitor has not been investigated. Although afatinib differs from EGFR antibodies by irreversibly targeting the intracellular portion of the receptor, it is conceivable that the same resistance mechanisms may affect treatment with afatinib. Efficacy and to