A Downward Trend of the Ratio of Influenza RNA Copy Number to Infectious Viral Titer in Hospitalized Influenza A-Infected Patients.

Although influenza frequently results in a self-limited respiratory illness, in most cases it can cause severe complications leading to hospitalization and death, especially in high-risk groups including elderly patients, immunocompromised persons, pregnant women, very young children, and persons with underlying medical conditions [1]. Worldwide, the annual influenza epidemics are estimated to result in approximately 3 to 5 million cases of severe illness and approximately 250 000 to 500 000 deaths [2]. Among hospitalized patients, antiviral therapy is most efficacious if started within 48 hours of influenza illness onset [1, 3–5]. Nonetheless, some studies have demonstrated that antiviral treatment is still beneficial in hospitalized patients when started 4 and 5 days after illness onset [6–12]. Current available antiviral treatments include the neuraminidase inhibitors ([NAIs] inhaled zanamivir, oral oseltamivir, and intravenous peramivir) and M2 ion channel blockers (amantadine and rimantadine). However, all currently circulating strains of influenza are adamantane-resistant, and therefore this class is not recommended for the prevention or treatment of influenza [13]. The NAI peramivir is given intravenously, and so it offers potential advantages in the management of severe hospitalized influenza. Indeed, NAI peramivir was used to treat critically ill patients during the 2009 pandemic under emergency use authorization by the Food and Drug Administration (FDA) [14, 15] and, it received FDA approval to treat acute uncomplicated influenza patients [16–18]. Until now, little or no resistance to zanamivir has been observed [13], and only a small percentage of A(H1N1)pdm09 viruses had highly reduced peramivir inhibition [19]. Although oseltamivir resistance was found in virtually all former seasonal A(H1N1) viruses, the frequency of oseltamivir resistance in A(H1N1)pdm09 viruses has remained low on a global scale. However, with the appearance of permissive mutations in A(H1N1)pdm09 viruses, the risk that oseltamivir-resistant viruses may spread globally is increasing [13]. With the appearance of permissive mutations in A(H1N1)pdm09 viruses, the risk that oseltamivir-resistant or even multidrug-resistant viruses may spread globally is increasing [13, 20], and there is a need for new antivirals with other mechanisms of action. One of the key challenges in designing influenza clinical trials is to define relevant efficacy endpoints. Although such efficacy endpoints are well established for acute uncomplicated influenza, there is a lack of consensus on the optimal endpoint to use for individuals hospitalized with influenza. Several antiviral drugs have been studied in hospitalized influenza patients, but none has clearly demonstrated to be universally effective in predicting how patients feel, function, and recover from the infection [6, 9, 11, 16, 17]. Proposed primary clinical endpoints in clinical trials for hospitalized influenza patients could include symptoms (eg fever, cough, sore throat), duration of hospitalization, time to normalization of vital signs and oxygenation, requirements for supplemental oxygen, need for admission to the intensive care unit (ICU), or assisted ventilation and mortality [21, 22]. Although antiviral drugs would be predicted to reduce viral shedding, and several studies have correlated viral load (VL) reduction with changes in chemokines and cytokines as well as clinical symptoms, the regulatory agencies do not permit virology to be a primary endpoint. Virological measurements include detection and quantification of shed virus by viral culture or quantitative reverse transcription-polymerase chain reaction (RT-qPCR). Virologic endpoints that are typically assessed, as secondary endpoints, in clinical studies of novel antivirals include baseline viral RNA copy number or infectious viral titer during the baseline visit and change in viral RNA copy number or titer over the course of illness [16–18, 21]. Quantitation of viral RNA by molecular techniques is a standard and sensitive method for VL determination. Cell-based assays are more labor intensive but have the advantage of detecting the infectious virus particles. Regulators preclude virology from being a primary endpoint because they feel that there is insufficient correlation between viral titers and change in titers with clinical symptoms. Furthermore, there is significant variability in viral shedding from patient to patient. Lastly, there is a lack of standardization in the collection of samples and location (eg upper vs lower respiratory tract) and assays for virologic measurements [21]. In this study, we investigated the relationship between influenza A RNA copy numbers and infectious viral titer in laboratory-confirmed influenza A patients during hospitalization with the hope that this virological data will inform future study endpoints.