Mutant targets

A general mechanism of resistance is reducing the affinity of the antiretroviral compound for its mutant target protein. Resistance mutations associated with reduced affinity are observed during treatment failure with a fusion inhibitor, nonnucleoside reverse transcriptase inhibitors (NNRTl), integrase inhibitor, and protease inhibitors as reviewed in Chaps. 3,4, 6, and 7 (Hazuda et al. 2007 Hsiou et al. 2001 King et al. 2002 Mink et al. 2005). 

As we optimized Tethering we used a variety of mass spectrometers. In our experience, the sensitivity and high resolution of TOP analyzers has provided the most rapid and accurate analyses of intact proteins. An example of an ESI-TOF data set from a standard experiment is illustrated in Fig. 9.2. Figure 9.2A is the deconvoluted mass spectrum of a Cys-mutant target protein after equilibration... 

Genetic disposition, either by interaction of the drug with a mutant target or by the lack of certain (or mutant) metabolic enzyme (e.g., the inability of about 1—3% of the Caucasian population to metabolize S-warfarin, due to a CYP2C9 deficiency). 

AdA24 is an adenoviral mutant targeting pRb-dysftinction [107]. It contains a 24-bp deletion in the pRb-binding domain of ElA and therefore depends on a deregulated pRb pathway for replication. This mutant has been shown to replicate in proliferating or Gi-arrested glioma and osteosarcoma cell lines (in which the pRb/pl6 " pathway is disrupted), but not in normal Gi-arrested fibroblasts and to inhibit tumor growth after intratumoral injection [107]. 

Random or specific mutants Mutants targeting a known interaction... 

Wolfel T, Hauer M, Schneider J, Serrano M, Wolfel C, Kle-hmann-Hieb E, De Plaen E, Hankein T, Meyer zum Buschenfelde KH, Beach D (1995) A pl6JNK4a-insensi-tive CDK4 mutant targeted by cytolytic T lymphocytes in a human melanoma. Science 269 1281-1284... 

Fig. 5. Generation of mutants using single-stranded DNA. After cloning the target gene into M13, the phage is propagated in the E. coll dut, ung strain of E.

Although the NNRTIs target HIV-1 RT, they are clearly different from the nucleoside RT inhibitors (NRTIs). They are highly selective for HlV-1 and do not inhibit HlV-2 or any other retrovirus. Moreover, the resistance spectrum of NNRTIs is different from that of NRTI, and, as a rule, NRTl-resistant mutant virus strains keep full sensitivity to the inhibitory effects of NNRTIs, and NNRTI-resistant mutant virus strains keep full sensitivity to the inhibitory effects of NRTIs. However, some influence of NRTI mutations on NNRTl susceptibility has been observed (Shuhnan etal. 2004). 

Trono D, Feinberg MB, Baltimore D (1989) HlV-1 Gag mutants can dominantly interfere with the replication of the wild-type virus. Cell 59 113-120 Tung FY, Bowen SW (1998) Targeted inhibition of hepatitis B virus gene expression a gene therapy approach. Front Biosci 3 all-al5... 

The Rieske protein in mitochondrial bci complexes is assembled when the protein is incorporated into the complex. The Rieske protein is encoded in the nucleus and synthesized in the cytosol with a mitochondrial targeting presequence, which is required to direct the apoprotein to the mitochondrial matrix. The C-terminus is then targeted back to the outside of the inner mitochondrial membrane where the Rieske cluster is assembled. In addition, the presequence is removed and the protein is processed to its mature size after the protein is inserted into the bci complex. In mammals, the presequence is cleaved in a single step by the core proteins 1 and 2, which are related to the general mitochondrial matrix processing protease (MPP) a and (3 subunits the bovine heart presequence is retained as a 8.0 kDa subunit of the complex (42, 107). In Saccharomyces cerevis-iae, processing occurs in two steps Initially, the yeast MPP removes 22 amino acid residues to convert the precursor to the intermediate form, and then the mitochondrial intermediate protease (MIP) removes 8 residues after the intermediate form is in the bci complex (47). Cleavage by MIP is independent of the assembly of the Rieske cluster Conversion of the intermediate to the mature form was observed in a yeast mutant that did not assemble any Rieske cluster (35). However, in most mutants where the assembly of the Rieske cluster is prevented, the amount of Rieske protein is drastically reduced, most likely because of instability (35, 44). 

Resistance to DDT has been developed in many insect species. Although there are some cases of metabolic resistance (e.g., strains high in DDT dehydrochlorinase activity), particular interest has been focused on kdr and super kdr mechanisms based upon aberrant forms of the sodium channel—the principal target for DDT. There are many examples of insects developing resistance to dieldrin. The best-known mechanism is the production of mutant forms of the target site (GABA receptor), which are insensitive to the insecticide. 

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