No abstract

Detection and sequencing of mutations from clinical specimens is often complicated by the presence of an excess of nonmutated cells. To facilitate the detection and sequencing of minority mutations from clinical specimens, we developed wild-type blocking polymerase chain reaction (WTB-PCR). This technique allows sensitive detection of minority mutations in a tissue sample containing excess wild-type DNA. In WTB-PCR, a nonextendable locked nucleic acid (LNA) oligonucleotide binds tightly to a region of wild-type DNA known to develop point mutations. This LNA sequence blocks amplification of wild-type DNA during PCR while permitting amplification of mutant exon 15. Our results show that the LNA blocking oligonucleotide inhibits amplification of wildtype DNA in a dose-dependent manner. WTB-PCR was able to detect mutant DNA in clinical samples of melanoma tissue containing an excess of nonmelanoma cells. This method was also able to detect small amounts of point mutated or tandem mutated DNA diluted with a much larger concentration of wild-type DNA. This rapid and simple assay overcomes the limitations of current methods to detect minority mutations. The potential applications of WTB-PCR include early diagnosis and prognosis of various cancers.

Psoriatic arthritis (PsA) is an inflammatory arthritis associated with irreversible joint damage in a subset of individuals. There is a need to screen early for this condition to prevent damage. To meet this need, we have developed the psoriatic arthritis screening and evaluation (PASE) questionnaire. The 15-item PASE questionnaire was administered to 190 individuals with either psoriasis or PsA. The PASE questionnaire was readministered to a subset of individuals with PsA in order to assess test–retest reliability and sensitivity-to-change. Receiver operator curves were constructed to optimize sensitivity and specificity for the diagnosis of PsA. Of the 190 participating in the study, 19.5% (37/191) participants were diagnosed with PsA. PASE total scores ranged from 15 to 74 (possible range, 15–75). The PsA group had a median Total score of 51 (25th and 75th percentile 44 and 57), and non-PsA group had a median total score of 34 (25th and 75th percentile 21 and 49) (p < 0.001). A PASE total score of 44 was able to distinguish PsA from non-PsA participants with 76% sensitivity and 76% specificity. Furthermore, 13 of the 15 items demonstrated significant test–retest reliability as assessed by Pearson correlation coefficient (r ≥ 0.5). PASE was sensitive-to-change with therapy; PASE scores were significantly lower for PsA individuals after systemic therapy (p < 0.034). The PASE questionnaire is a valid and reliable tool to screen for active PsA among individuals with psoriasis. PASE scores may be used as a marker of therapeutic response.