Why does "start debugging" cause an immediate HTTP404 error?

Setup

I recently changed computers, and pulled down a codebase that had been working before. I created the application using the "Create Virtual Directory" button in Visual Studio, and I can confirm that the application shows up in IIS as an application. I know this codebase works because it's in production; I've just changed development environments.

The Problem: HTTP 404.0

When I run the application, the debugger launches and a browser tab opens to HTTP 404.0

The problem resembles this unanswered question, except I'm using IIS 10 instead of 8.5: https://serverfault.com/questions/702235/iis-8-5-instant-404-0-error-on-mvc-site

Details:
Detailed Error Information:
Module             IIS Web Core
Notification       MapRequestHandler
Handler            StaticFile
Error Code         0x80070002
Requested URL      http://localhost:80/OURMVCSITE
Physical Path      D:\OURMVCSITE
Logon Method       Anonymous
Logon User         Anonymous

What I've tried

Many StackOverflow answers point to issues with the RouteConfig or Global.asax.cs. I've set breakpoints in both; neither class is ever executed.

The app pool is the default app pool running in Integrated mode under the Network Service user, and Network Service has access to every file and folder in the directory.

Searching for answers in general is somewhat difficult because most results are about creating custom 404 error pages for when users incorrectly enter a URL.