Chronic infection with hepatitis C virus (HCV) is a leading cause of liver disease, cirrhosis, and hepatocellular carcinoma, resulting in 475,000 deaths annually [1]. Estimates of prevalence based on seropositivity range between 1.3 and 2.1%, or between 92 and 149 million individuals globally [2]. Though direct-acting antiviral (DAA) therapy is largely curative, only a minority of chronic infections (~ 20%) have been diagnosed, with even fewer treated (~ 3%) [1]. Persistent challenges in screening, diagnosis, access to affordable DAA, and the risk for re-infection in vulnerable populations aggravates elimination efforts [2]. A prophylactic vaccine for HCV is therefore still urgently needed.
The extreme diversity of the virus, with billions of related but distinct variants circulating in each infected person, has been a major barrier to vaccine development [3]. Early candidate vaccines using mammalian expressed E2 glycoprotein (E2), a meditator of viral entry expressed on the surface of mature virions, were successful in inducing protective immunity in chimpanzee against homologous challenge [4]. However, progression to chronic infection was observed following heterologous challenge, with escape mutations subsequently mapped to the highly variable N-terminus of E2, termed HVR1 [4, 5]. Since then, cohort and in vitro analysis has consistently identified HVR1 as the principle target of neutralizing antibodies (nAb) in natural infection [6].
Recently, the immunodominant HVR1 has been hypothesized to divert the humoral response from conserved neutralizing epitopes [7, 8]. Some groups have therefore sought to elicit cross-neutralizing Ab targeting conserved epitopes by amputating HVR1 from E2 immunogens [9]. It was shown that HVR1-deleted E2 was an inferior immunogen, failing to elicit even homologous nAb following immunization [9]. Variations in HVR1 have since been implicated in resistance to extra HVR1 targeting neutralizing antibodies, underscoring the crucial role of the anti-HVR1 response in any potential HCV vaccine [10]. To understand how HCV variation mediates immune escape, global networks of HVR1 cross-reactivity have been elaborated [11]. Consistent with cohort analysis, chimpanzee vaccination, and in vitro neutralization assays, the sequence similarity between two HVR1 peptides was predictive of cross-reactivity [5, 6, 11]. However, cross-reactive pairs with low sequence similarity were also observed, indicating a more complex relationship between HVR1 variability and immune evasion [11]. Moreover, these studies have not established if the observed association between cross-reactivity and sequence similarity applies to cross-neutralization. Given the critical implications of these questions for HCV vaccine design, we sought to clarify these dynamics by synthesizing high-Hamming distance HVR1 peptides, immunizing mice, and evaluating how sequence similarity associated with cross-neutralization. We hypothesized that intrinsic physicochemical features of HVR1 sequences contributing to secondary structure might influence resistance to neutralization.