Supplemental Data For:

Mutter GL, Ince TA, Baak JPA, Kust G, Zhou X, Eng C. Molecular identification of latent precancers in histologically normal endometrium. Cancer Res 2001; 61(8).

 Supplemental Figure 1: Sequence confirmed PTEN mutation in proliferative endometrium. Single-plex PCR and denaturing gradient gel electrophoresis of the exon 5 of PTEN gene. DNA from PTEN-null glands (NULL) have aberrantly migrating species (arrowheads) compared to PTEN expressing (POS) glands. Bidirectional (for, forward; rev, reverse) direct sequence confirmation of mutations is shown in panels below. PTEN immunohistochemistry of these two proliferative endometria is shown in Figure 1, left of the published paper.

Supplemental Figure 2: Model of endometrial cancer pathways. Endometrioid endometrial adenocarcinoma (excluding papillary serous and non-endometrioid cancers) can arise through a PTEN dependent (left) or independent (right) pathway(7). Three stages of EIN-mediated endometrial carcinogenesis are shown, top to bottom, as tissue compartments proportionately shaded to show that fraction (% PTEN defect) of endometrial glands at each stage  which have PTEN mutation and/or deletion (black) or normal PTEN (white). Normal proliferative endometria contains a small fraction (42% of women have 2-3% PTEN mutated/deleted glands, for overall PTEN null rate estimated here as 1%) of PTEN-defective cells. EIN and cancer (CA) are usually homogenous regarding PTEN expression, so defective and non-defective fractions are reported as published PTEN mutation/deletion rates for these diagnoses in cases with a documented premalignant phase {4636}. Latent PTEN mutant clones within proliferative endometria are, individually, at least 50 times more susceptible to EIN conversion than glands with normal PTEN genotype.

Supplemental Table 1. PTEN mutation, deletion, and expression status in Normal Proliferative Endometria.
Laser capture microdissection was directed by PTEN immunohistochemistry of an adjacent serial section to enable DNA isolation from PTEN expressing (+) and non-expressing (null, -) endometrial glands.  All mutations listed are based upon sequence confirmed changes screened by Denaturaing Gradient Gel Electrophoresis.  Examples of Sequence Confirmation appear in Supplemental Figure 1, above.

*          PTEN protein Null (-) and Positive (+) glands from individual biopsies.
†          LOH, Loss of Heterozygosity (NI=not informative, ROH=retention of heterozygosity;
            LOH=loss of heterozygosity). 

Supplemental Table 2: Morphometric Characteristics of PTEN-null Glands, by Diagnosis within the Morphometry Window.  This is a tabular format of the same data which appears in published Figure 2.
*          two-tailed ANOVA, significance