diff --git a/cwltool/pathmapper.py b/cwltool/pathmapper.py
index c89fac5f4..7ed4437e6 100644
--- a/cwltool/pathmapper.py
+++ b/cwltool/pathmapper.py
@@ -178,6 +178,16 @@ def setup(self, referenced_files: List[CWLObjectType], basedir: str) -> None:
                 staged=True,
             )
 
+        targets: Dict[str, str] = {}
+        for k, v in self._pathmap.items():
+            if v.target in targets:
+                raise ValidationException(
+                    "Name conflict: both %s and %s have been assigned to target %s"
+                    % (k, targets[v.target], v.target)
+                )
+            if v.type != "Directory":
+                targets[v.target] = k
+
     def mapper(self, src: str) -> MapperEnt:
         if "#" in src:
             i = src.index("#")