AQUA Analysis of Thymidylate Synthase Reveals Localization to be a Key Prognostic Biomarker in 2 Lar - PDF by ProQuest


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									           AQUA Analysis of Thymidylate Synthase Reveals
          Localization to be a Key Prognostic Biomarker in
             2 Large Cohorts of Colorectal Carcinoma
Mark D. Gustavson, PhD; Annette M. Molinaro, PhD; Greg Tedeschi, BS; Robert L. Camp, MD, PhD; David L. Rimm, MD, PhD

● Context.—Increased thymidylate synthase expression is                         58%] for the top 54% of cytoplasmic-expressing tumors
a marker for decreased survival in colorectal cancer.                           [P     .02]) was observed for the training set. A higher nu-
  Objective.—Thymidylate synthase localizes to both the                         clear to cytoplasmic ratio also correlated significantly with
nucleus and cytoplasm, but how the relationship of these                        decreased survival (15% decreased survival [66% to 51%]
expression levels affects colon cancer outcome has yet to                       for the top 25% of tumors [P         .001]). Applying these
be determined.                                                                  findings to the validation set, as a function of time to re-
  Design.—Using AQUA, we assessed prognosis of thymi-                           currence, only the ratio (P     .03 [expression ratio]; P
dylate synthase expression as a function of subcellular lo-                     .18 [nuclear]; P    .71 [cytoplasmic]) showed a significant
calization in 2 retrospective cohorts of colorectal carci-                      association with decreased time to recurrence. Addition-
noma. We used the first cohort (n 599) as a training set,                        ally, the expression ratio significantly added to the prog-
subsequently validating optimal expression cut points in                        nostic value given by the primary tumor pathologic clas-
the second cohort (n     447).                                                  sification and nodal status.
  Results.—A significant association between decreased                              Conclusions.—These data suggest the relationship of nu-
5-year disease-specific survival and increased nuclear ex-                       clear to cytoplasmic thymidylate synthase expression, giv-
pression (16% decreased survival [72% to 56%] for the                           en as a ratio of continuous AQUA scores, to be a strong
top 60% of nuclear-expressing tumors [P .001]) and cy-                          predictor of colon cancer survival.
toplasmic expression (12% decreased survival [70% to                               (Arch Pathol Lab Med. 2008;132:1746–1752)

T  hymidylate synthase (TS) catalyzes the reductive meth-
     ylation of deoxyuridylate in the pathway for the pro-
duction of deoxythymidine triphosphate, which is critical
                                                                                However, considerable heterogeneity exists in the percent-
                                                                                age of positivity within the population as well as vari-
                                                                                ability in the literature about the overall prognostic value
for DNA synthesis.1 Thymidylate synthase expression, as                         of TS expression.9 This variability is most likely due to
a determinant of sensitivity to fluoropyrimidines, has                           differences in methodologies, including differential defi-
been demonstrated in vitro,2,3 and TS expression in vivo                        nitions of TS positivity determined by subjective assess-
may play an important role in determining tumor re-                             ments of expression levels by traditional immunohisto-
sponse to 5-fluorouracil (5-FU).4,5 Thymidylate synthase                         chemical techniques. Studies measuring mRNA have re-
has been suggested to be both prognostic6,7 and predictive                      moved considerable subjectivity but have still failed to be-
of response to therapy (see Aschele et al8 for review).                         come part of the routine practice for management of colon
   Accepted for publication May 6, 2008.                                           Recently, it has been demonstrated that TS may have
   From the Departments of Pathology (Drs Gustavson, Camp, and                  other cellular functions, including translational regulation
Rimm, and Mr Tedeschi) and Epidemiology and Public Health (Dr Mol-              (see Liu et al12 for review). Thus the subcellular localiza-
inaro), Yale University, New Haven, Conn. Dr Gustavson and Mr Te-               tion of expression may be an important determinant of
deschi are now with HistoRx, Inc, New Haven, Conn.
   Dr Gustavson and Mr Tedeschi, although not at the time of the re-            the functional role of TS. Because of the potential impor-
search, are currently employed by HistoRx, Inc, a private company to            tance and functional consequence of TS subcellular local-
which Yale University has given exclusive rights to the AQUA tech-              ization, we wished to examine the role of TS localization
nology. Drs Camp and Rimm are stockholders, scientific founders, and             as a function of survival. We recently developed a method
consultants to HistoRx, Inc. Dr Molinaro has no relevant financial in-           of automated quantitative analysis (AQUA) that allows
terest in the products or companies described in this article.
   Sections of this article previously appeared in ‘‘Methods for Making         rapid analysis of immunofluorescence on tissue samples.13
Cancer Prognoses Based on the Subcellular Localization of Biomark-              This method yields quantitative results comparable to en-
ers,’’ p
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