People who inject drugs (PWID) experience sexual and injection-related HIV risks, but uptake of pre-exposure prophylaxis (PrEP) for HIV prevention among PWID has been low. Improving PrEP uptake in this population will require understanding of PrEP knowledge and interest. In 2017, we conducted in-depth, semistructured interviews with HIV-uninfected PWID and key informants (PrEP and harm reduction providers) in the US Northeast. Thematic analysis of coded data explored PrEP knowledge and the factors that influence PrEP interest. Among PWID (n = 33), median age was 36 years, 55% were male, 67% were white, and 24% identified as Hispanic/Latino. Accurate PrEP knowledge among PWID was low, which key informants (n = 12) attributed to PrEP marketing focused on other risk populations, as well as healthcare providers' lack of time and unwillingness to discuss PrEP with PWID. There was a discrepancy between self-reported HIV risk behaviors, which were common, and HIV risk perceptions, which varied and strongly influenced PrEP interest. Most PWID and key informants thought that PrEP would be most beneficial for those who shared syringes, used discarded syringes, engaged in transactional sex, or were homeless. Improving uptake of PrEP for HIV prevention among high-risk PWID will require education to increase PrEP knowledge and addressing factors that negatively influence PrEP interest such as perceptions regarding low HIV risk and the process for obtaining PrEP. This may require specialized PrEP marketing and outreach efforts and improved capacity of healthcare providers to effectively assess HIV risk (and perceptions) and communicate the benefits of PrEP to at-risk PWID.