Phylogenetic analysis of classical swine fever virus (CSFV) field isolates from outbreaks in South and Central America.

To date, there is little information concerning the epidemiological situation of classical swine fever (CSF) in the Americas. Besides summarizing the available data, genotyping of isolates from outbreaks in domestic pigs in several countries of South and Central America was performed. For this, a 190 base fragment of the E2 envelope glycoprotein gene was used. European strains and isolates, and historical isolates from the United States (US) were included for comparison. In contrast to the situation in most parts of Europe, where group 2 isolates predominate, it was found that all the isolates from the American continent analyzed belonged to group 1 and were further resolved into three subgroups. The Cuban isolates clustered in subgroup 1.2, whereas the isolates from Honduras and Guatemala clustered in subgroup 1.3. The remaining isolates from Argentina, Brazil, Colombia and Mexico generated four poorly resolved clusters in subgroup 1.1, together with the vaccine strains, with historical European and US isolates, and with a recent Russian isolate. While the vaccine strains and the historical European isolates formed a relatively distinct cluster, one of the US isolates clustered together with the Mexican, and another one with Colombian isolates. Historically, CSF (hog cholera) was observed almost simultaneously in the US and in Europe in the first half of the 19th century, and its origin remains a matter of discussion. Our results showed that the US isolates are closely related to isolates from South America, while appearance of isolates in Cuba on one hand and in Honduras and Guatemala on the other hand, seems to have been due to unrelated events. This allows to speculate that at least in the American continent, CSF virus may have appeared independently in several regions, and spreading may have been a secondary effect.

Preclinical modeling of EGFR-specific antibody resistance: oncogenic and immune-associated escape mechanisms.

To define the molecular basis of secondary resistance to epidermal growth factor receptor (EGFR)-specific antibodies is crucial to increase clinical benefit in patients. The limited access to posttreatment tumor samples constitutes the major barrier to conduct these studies, representing preclinical experimentation as a useful alternative. Anti-EGFR antibody-based therapy has been reported to mediate tumor regression by interrupting oncogenic signals and, more recently, by inducing antitumor immunological responses. However, resistance models have been focused only on tumor escape associated with EGFR blockade, whereas studies describing immune-associated escape mechanisms have not been reported thus far. To address this idea, we modeled resistance induction in D122 metastasis-bearing C57BL/6 mice treated with 7A7 (an anti-murine EGFR antibody). Similarly to patients receiving EGFR-specific antibodies, 7A7 resistance promotion represents an important drawback to successful therapy. Characterization of primary cultures derived from metastasis in 7A7-treated mice revealed a high frequency of tumor variants resistant to in vivo and in vitro antibody treatment. We showed, for the first time, the convergence of alterations in oncogenic and immunological pathways in 7A7-resistant variants. To identify key molecules behind resistance, seven 7A7-resistant variants were screened. HER3 overexpression and PTEN deficiency leading to hyperactivation of protumoral downstream signaling were found in these variants as a consequence of 7A7-mediated EGFR inhibition. Concomitantly, we found a high percentage of resistant variants carrying abnormalities in the constitutive and/or interferon gamma (IFN-γ)-inducible major histocompatibility complex I (MHC-I) expression. A significant decrease in mRNA levels for MHC-I heavy chains, ß2-microglogulin and antigen processing machinery genes as well as transcriptional alterations in IFN-γ pathway components were identified as the main mechanisms underlying MHC-I expression defects in 7A7-resistant variants. Notably, these defects have not been previously associated with EGFR-specific antibody resistance, providing novel immunological escape mechanisms. This study has strong implications for the development of new combination strategies to overcome anti-EGFR antibodies refractoriness.