**1. Measles Virus Epidemiology**

Measles virus (MeV) is the etiologic agent responsible for measles disease. Humans are the only known reservoir for MeV. Despite the availability of a very efficient vaccine [1], measles remains one of the most contagious diseases with a R0 ranking from 12 to 18 [2] meaning that (in a fully susceptible population) an infected patient will on average transmit the infection to 12 to 18 individuals. This propagation rate may even increase among people with low or compromised immunity [3]. Viral transmission generally occurs from person to person through aerosols [3] and precedes onset of skin rash, making the disease even more difficult to contain. After decades of emergences mainly restricted to the poorest countries, measles has made a strong comeback and re-emerged in industrialized countries [4] where access to the vaccine was supposed to be easier. Measles killed more than 100,000 people every year [5] since 2010. In 2017, 110,000 people died from measles, mostly children under five years old [3]. Indeed, in the absence of vaccination, children are the main targets of MeV [6], although adults can be infected as well [3]. Last year, WHO documented 268,038 confirmed cases. Nevertheless, according to other estimations, there are 7 to 20 million people getting infected by measles each year [7,8].

In most developed countries measles was considered eliminated, in recent years. However the rate of vaccination decreased due to a vaccination hesitancy, and as consequence the decreased herd immunity led to large outbreaks and today measles is considered re-emerged [4,9]. This year, in many developed countries including USA and France, there is a 300% increase in reported MeV cases compared to last year [10]. Notably, 1250 cases have been reported in the USA in 2019 (from January to October) [11]. Those outbreaks confirm the re-emergence of measles, already announced by the NIAID following MeV epidemics in 2014 (CDC).

#### **2. Virus**

MeV belongs to the Morbillivirus genus within the *Paramyxoviridae* family and *Mononegavirales* order. This enveloped virus produces pleiomorphic viral particles with an average size ranging from 150 to 300 nm and up to 900 nm [12]. Its genome is a negative-sense, single stranded RNA of 15,894 nucleotides that encodes six structural proteins: The nucleocapsid (N) protein, the phosphoprotein (P), the matrix (M) protein, the fusion (F) protein, the haemagglutinin (H) protein, and the polymerase (large, L) protein. Two non-structural proteins, V and C are produced from the P gene [13] and mainly alter the innate immune sensing and response [14–17].

Wild type MeV strains use signaling lymphocytic activation molecule 1 (SLAMF1, also called SLAM or CD150) and nectin-4 receptors to infect target cells [18–20]. MeV vaccine strains use the ubiquitously expressed CD46 molecule as an additional entry receptor in vitro [21,22]. MeV entry is pH-independent and occurs directly at the cell surface [23]. However, MeV entry may also occur by endocytosis mediated by SLAM in B-lymphoblastoid cells or A549-SLAM cells [24], and through a nectin-4-mediated macropinocytosis pathway, in breast and colon cancer cell lines (MCF7, HTB-20, and DLD-1) [25]. It was also suggested that MeV Edmonston or Hallé strains could use a macropinocytosis-like pathway in non-lymphoid and lymphoid cells when SLAM and CD46 are engaged but this remains poorly documented [26,27].

To initiate the infection of the main target cells, the MeV H protein binds to entry receptor on the surface. This attachment triggers the F protein and leads to exposure of its hydrophobic fusion peptide that then inserts into the host cell membrane. The F protein undergoes serial conformational changes allowing the merge of the host and viral membranes creating a fusion pore allowing the ribonucleocapsid (RNP) delivery in the cytoplasm (Figure 1A,B) [28,29]. Infection also spreads efficiently via cell-to-cell contact [30,31].

Transcription by the RNA-dependent RNA polymerase (RdRp) starts from a single promoter resulting in a transcriptional gradient from the most abundant mRNA for N to the least abundant mRNA for L in order to allow efficient viral cycles. These mRNAs are then translated into viral proteins. The accumulation of N and P leads to viral genome replication into positive stranded RNA anti-genome that will allow further synthesis of negative sense RNA strands that will be encapsidated by newly synthesized N, P, and L proteins [32]. Viral RNA synthesis and assembly are regulated through the interaction between M and N [33]. Viral proteins assemble to the plasma membrane and the budding of new virions can occur (Figure 1A). Alternatively, the surface glycoproteins are transported to the plasma membrane and allow cell-to-cell dissemination.

The viral RNA is encapsidated by the protein N and forms the helical nucleocapsid [34]. Each N protein covers six nucleotides, hence the genome length has to follow the "rule of 6" for being fully protected [35,36]. Together, the proteins L and P form the viral RdRp. That polymerase interacts with the nucleocapsid to progress on the viral RNA: Altogether they form the RNP.

The M protein generally ensures the viral particle integrity. The M protein also orchestrates the viral assembly at the plasma membrane and the budding of the new infectious viral particles [23].

The H and F proteins constitute the viral fusion complex that is responsible for the viral entry into the host cell. The H protein is a tetramer organized as a dimer of dimers responsible for the binding to the entry receptor. The F protein mediates the fusion between the virus and the host plasma membranes. The F is a trimer first produced as a precursor F0 that is cleaved in the trans-Golgi by a furin protease in F1 plus F2 subunits linked by a disulfide bond. The extracellular domain is constituted by the F1 and F2 subunits containing the fusion peptide at the N terminus followed by two complementary heptad repeat domains, respectively at the N terminus (HRN) and the C terminus (HRC). While the crystal structure of the prefusion form of the F protein has been described [37], the exact delimitations of the F sub-domains are still not completely defined [38–40].

**Figure 1.** Measles Virus (MeV) replication cycle. (**A**) In order to infect a susceptible and permissive cell, MeV binds to its entry receptors on the cell surface (1) and initiates the virus-cell membrane fusion (2), as described in detail in (**B**). Virus and cell membranes fusion leads to genome delivery into the cytoplasm (3). Viral RNA is transcribed in mRNA (4) that is further translated into viral proteins (5). Viral glycoproteins maturate during their transport to the cell surface (6). The replication of positive stranded anti-genomic RNA starts in the cytoplasm (7) and serves as a template for synthesis of new negative stranded genomic RNA (8). Viral proteins assemble at the cell surface, leading either to budding of new virions (9) or cell-to-cell fusion (10). (**B**) The haemagglutinin (H) protein binds to the MeV receptor at the cell surface, allowing the triggering of fusion (F) which reaches a metastable conformation. Then, F protein anchors its fusion peptide in the target cell membrane, F undergoes serial conformational changes bringing the two membranes close enough to merge and form a pore throughout which the viral ribonucleocapsid (RNP) is delivered to the cytoplasm.

Based on bioinformatic tools the HRN domain encompasses residues 116/138 to residue 190 and the HRC domain is included between residues 438 and 488/489. The current crystal structure however shows the region between 438 and 458 as disorganized while a canonical heptad repeat is shown after residue 458 [41,42].

#### **3. Vaccines**

A highly efficient live-attenuated virus vaccine is available to prevent measles outbreaks. MeV transmissibility is very high and 95% of the population needs to possess anti-measles immunity for disease eradication [43]. In 1997, during a meeting co-sponsored by the World Health Organization (WHO), the Pan American Health Organization (PAHO), and the Centers for Disease Control (CDC), the experts agreed that measles eradication was technically feasible by 2005–2010. Nevertheless, vaccination coverage decreased and led to a re-emergence of measles infection. Nowadays, measles global eradication is one of the top priorities of the expanded program on immunization (EPI) supported by the WHO. The Global Vaccine Action Plan aims to eliminate measles in five WHO Regions by 2020. Based on confirmed cases reported by the WHO, the countries with the most measles cases in 2018 were India, Ukraine, Philippines, Brazil, and Yemen. Recently, measles strongly re-emerged in industrialized countries due to the significant decrease in vaccination coverage [4,44].

Different MeV strains have been used for vaccine purpose starting with the Edmonston strain isolated in 1954 that was very reactogenic. Five vaccines were derived from Edmonston: Edmonston-Zagreb, AIK-C, Moraten, Schwarz, and Edmonston-B [45]. Some of them such as Edmonston-B remained too reactogenic. The Edmonston vaccine was replaced by the more attenuated Schwarz vaccine strain in early 60s and Moraten vaccine strains in 1968. Years later, studies have shown that Schwarz and Moraten were in fact the same virus [45]. Other vaccines derived from other strains have also been developed. Leningrad strain (isolated in 1957) attenuation successively led to Leningrad 4 and more recently to the Chinese vaccine Changchung-47. Shanghai isolate (1960) attenuation allowed production of shanghai-191 vaccine while Cam-70 which was currently produced and used in Indonesia and Japan, derived from the Tanabe (Japan, 1968) strain. All vaccines strains belong to the measles virus genotype A [45]. Measles vaccine is usually combined with mumps and rubella vaccines, known as MMR (Measles, Mumps, and Rubella) vaccine, or with mumps, rubella, and varicella (chickenpox) vaccines, called MMRV (Measles, Mumps, Rubella, and Varicella) vaccine. MMR is a live-attenuated measles virus [46]. MMR vaccination is given in a two-dose schedule, with a first dose generally administered to 12–15 months old children, and a second one three to five years later [4]. While MMR vaccine cannot be used in immunocompromised patients (with low CD4+ cell count, or severely immunedepressed), the WHO strongly recommends the vaccination of human immunodeficiency virus (HIV) positive patients without severe immunosuppression [47].

Generally, vaccinated people develop a strong humoral and cellular immunity. Only 2–10% of people who received the two vaccine doses do not produce protective measles antibodies. However, most of them remain protected by their T cell immunity [48,49].

Taken together, the too low vaccination coverage combined with the increasing proportion of immunocompromised and other non-vaccinable people call for the development of an efficient, preventive, and/or curative treatment.

#### **4. Disease**/**Generalities**

#### *4.1. Symptoms and Complications*

During the acute phase of MeV infection, the patients develop several symptoms, including fever, cough, nasal congestion, characteristic erythematous maculopapular rash, conjunctivitis, and pathognomonic Koplik spots on oral mucosa. Diarrhea and vomiting are often observed in infected children during the disease [50,51] or appear as a complication following the disease [5,52]. Additionally, MeV infection leads to a strong immunosuppression that can last for several months and lead to severe secondary infections [53,54]. Moreover, MeV seems to impact FoxP3 T regulatory cells homeostasis by increasing their frequency and attenuating the hypersensitivity cellular response [55]. A more recent study suggests a MeV-induced immune amnesia relying on the depletion of pre-existing memory lymphocytes [50].

MeV infection can lead to several complications such as pneumonia, which is the main cause of measles mortality [56] or to central nervous system (CNS) complications, and to a lower extent to thrombocytopenia, blindness, or hearing loss [57]. Briefly, interstitial pneumonitis associated with mucosal inflammation due to large syncytia formations in the lungs are mainly observed in immunocompromised patients (Hecht's pneumonia) [56,58]. This cytopathic effect leads to bronchio-epithelial destruction generally resolved within few days of hospitalization (Figure 2A).

**Figure 2.** Course of MeV infection leading to measles encephalitis. (**A**) Initially, MeV infects myeloid cells in the respiratory tract. Then, MeV-infected lymphocytes disseminate the infection via the lymphatic and vascular systems. As a consequence of transient immunosuppression or autoimmunity, patients can develop acute post-infectious measles encephalitis (APME) shortly after exposure without systematic central nervous system (CNS) infection. However, measles inclusion-body encephalitis (MIBE) and subacute sclerosing panencephalitis (SSPE) are associated with MeV infection of the CNS. (**B**) The occurrence of MeV encephalitis may range from one day to 15 years following initial infection.
