It's 2 a.m. the night after your mastectomy; that afternoon, the surgeon had told you that the surgery went well, that the cancer is out, and that the sentinel node looks negative on the preliminary report. But in the quiet darkness, the doubts and anxiety mount. How do they really know? When will I know for sure?

The sentinel node identified by the surgeon is immediately taken to the Pathology lab, where it is examined by a physician specializing in pathology. This pathologist measures the lymph node, and then slices it into thin slices. Each slice is carefully examined for visible evidence of metastatic tumor, which will look like white or tan spots in the usually pinkish-tan lymph node. If suspicious areas are found, that slice can be immediately frozen hard enough to make it possible to shave very thin slices (sections) which can be stained and examined with the microscope to determine whether or not tumor is producing that spot (i.e., a frozen section examination).

While this procedure can readily identify visible spots of metastatic tumor, deposits of microscopic size can be much more difficult to identify quickly enough to provide the necessary information for the surgeon in the operating room. Since finding metastatic tumor in the sentinel lymph node is a critical factor in whether or not an axillary lymph node dissection is done, it is important to find out if tumor is there in the most efficient way.

If all of the lymph node slices appear normal, the pathologist is reluctant to select one (or even two) slices at random for frozen section examination, because a very small tumor deposit may very well be in the slices not chosen. Even if all of the slices are frozen, the process of shaving the frozen block to produce a perfectly flat surface from which to cut very thin sections results in some of the lymph node tissue being irretrievably lost, with the risk of forever losing that small deposit of metastatic tumor.

The touch imprint evaluation has been developed to allow a rapid evaluation of the lymph node without loss of tissue. The lymph node slices are laid out next to each other, and several imprints from the array of slices are "stamped" onto the surface of a clean glass slide. Both normal cells and cancer cells (if present) will stick to this slide and can be distinguished under a microscope after staining. Within a few minutes of receiving the sentinel node from the operating room, the pathologist telephones a report to the surgeon, who can proceed with an axillary lymph node dissection if it is needed.

While infrequent, the sentinel lymph node touch imprint diagnosis can miss tiny metastatic tumor deposits. This is not too surprising, since the slices are about 1.5 millimeters thick and a group of tumor cells that is only 0.5 millimeters across could be "hiding" in the middle of the slice, where it will not touch the glass slide. Published false negative rates for not identifying metastatic tumor in a sentinel lymph node by frozen section or touch imprint range from 5-10% (i.e., 5-10% of the time, small tumor foci are not identified at this stage).

That is why the sentinel lymph node slices are subjected to a more thorough examination that requires overnight processing and special staining techniques. The slices, now embedded in paraffin, will have thin sections cut for microscopic examination from different levels within the slice, thus increasing the likelihood that a very small tumor cell group will end up on one of the slides. In addition, one or two slides will be stained with a special stain that reacts only with the cancer cells present (an immunohistochemical or IHC stain for keratin, present in the cancer cells, but not in the normal lymph node cells). This stain makes it much easier to identify very small tumor cell groups, but requires additional time to perform.

The final report from the pathologist will summarize not only the features of the tumor itself, but also the number of lymph nodes examined, how many contained metastatic tumor, and the size of the largest group of metastatic tumor cells. All of this information is used to determine the stage of the tumor (ranked I through IV), using the TNM (Tumor, Nodes from the region, Metastases to distant locations) system of the American Joint Committee on Cancer (AJCC). Low stage tumors are small with no positive lymph nodes or distant metastases, while higher stage tumors are larger with positive lymph nodes and/or spread to distant locations such as to bone. The stage is related to the probability of the tumor recurring or metastasizing, with higher stages having a greater probability of recurrence or spread than lower stages.

In the nodal part of this staging system for breast cancer, both the number of axillary nodes involved and the size of the metastatic tumor deposits are used in determining the stage. For example, if there are four positive axillary lymph nodes, this would be at least stage III, no matter what the size of the tumor in the breast. Metastatic tumor deposits larger than 2 millimeters have always been considered significant, while the behavior of the smallest metastases is now a subject of focus due to the adoption of sentinel lymph node examination.

The lymph node staging process has been updated to include a new category as a result of these sentinel lymph node studies. When immunohistochemical stains for keratin are performed as part of the sentinel lymph node examination, tumor deposits as small as a single cell or a group of a few cells can be found (isolated tumor cells or ITC). ITC have been defined as groups of tumor cells less than 0.2 millimeters in diameter (distinguishing them from micrometastases, which fall into the 0.2 to 2 millimeter range). The clinical significance of these ITC is still a matter of debate, because while they have travelled away from the main tumor location, the tumor cell(s) do not appear to have proliferated in the lymph node. In contrast, a 1 millimeter micrometastasis is estimated to contain about 500,000 cells - clearly, these cells have been multiplying since their arrival in the lymph node.

The separation of the smallest metastatic tumor deposits into micrometastasis and ITC is reflected in a recent modification to the TNM staging criteria. ITC are classified as pN0 (specifically, pN0(i+) reflecting the role of the immunohistochemical stain in detecting this tiny metastatic group), while a micrometastasis is classified as pN1 (specifically, pN1mi). This means that a woman with a 1.5 cm tumor having a lymph node which is pN0 would be classified as stage I, while a woman with the same size tumor with a lymph node which is pN1 would be classified as stage II.

Clinical studies continue since there are still questions about the real significance of the ITC, and we may see further refinements in the criteria for staging breast cancer.

Sharon Bannister, M.D.
Department of Pathology