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Validating whole slide imaging for diagnostic purposes in pathology

Validating whole slide imaging for diagnostic purposes in pathology

Background A tumor marker is a substance such as a protein, antigen or hormone in the body that may indicate the presence of cancer. Generally, these markers are specific to certain types of cancer and can be detected in blood, urine and tissue samples. The body may produce the marker in response to cancer or the tumor itself may produce the marker.

The detection of tumor markers may be used to determine a diagnosis or as an indicator of disease cancer progression. It can also be used to document clinical response to treatment. Tumor markers are normally produced in low quantities by cells in the body. Detection of a higher-than-normal serum level by radioimmunoassay or immunohistochemical techniques usually indicates the presence of a certain type of cancer.

Currently, the main use of tumor mfarkers is to assess a cancer's response to treatment and to check for recurrence. In some types of cancer, tumor marker levels may reflect the extent or stage of the disease and can be useful in predicting how well the disease will respond to treatment. A decrease or return to normal in the level of a tumor marker may indicate that the cancer has responded favorably to therapy.

If the tumor marker level rises, it may indicate that the cancer is spreading. Finally, measurements of tumor marker levels may be used after treatment has ended as a part of follow-up care to check for recurrence.

However, in many cases the literature states that measurements of tumor marker levels alone are insufficient to diagnose cancer for the following reasons: Examples of tumor markers include: High levels of B2M are an indicator of certain kinds of cancer, including chronic lymphocytic leukemia, non-Hodgkin's lymphoma and multiple myeloma or kidney disease; Beta Human Chorionic Gonadotropin beta HCG — A type of tumor marker that may be found in higher than normal amounts in individuals with some types of cancer; Calcitonin — Hormone secreted by the thyroid that lowers blood calcium; Calretinin — A calcium-binding protein that is used as a marker in the evaluation of lung cancer and other diseases.

Chromogranin A — A protein found inside neuroendocrine cells, which releases chromogranin A and other hormones into the blood. Chromogranin A may be found in higher than normal amounts in individuals with certain neuroendocrine tumors, small cell lung cancer, prostate cancer and other conditions Guanylyl cyclase c GCC — An enzyme that may be expressed only in the cells that line the intestine from the duodenum to the rectum.

Inhibin — One of two hormones designated inhibin-A and inhibin-B secreted by the gonads by Sertoli cells in the male and the granulosa cells in the female and inhibits the production of follicle-stimulating hormone FSH by the pituitary gland; Lactate Dehydrogenase LDH — Marker used to monitor treatment of testicular cancer; Mucin-1 MUC-1 — Carbohydrate antigen elevated in individuals with tumors of the breast, ovary, lung and prostate as well as other disorders; Napsin A — Protein used as a marker in the evaluation of lung cancer; Prealbumin — Marker of nutritional status and a sensitive indicator of protein synthesis.

Levels of PSA in the blood often increase in men with prostate cancer. Thyroglobulin — Protein found in the thyroid gland. Some thyroglobulin can be found in the blood and this amount may be measured after thyroid surgery to determine whether thyroid cancer has recurred; Thyroid Transcription Factor-1 TTF-1 — A protein that is used as a tumor marker in the evaluation of lung cancer; Transferrin — A protein in blood plasma that carries iron derived from food intake to the liver, spleen and bone marrow.

Tumors may be evaluated with histology, which involves examination of the structure, especially the microscopic structure, of organic tissues. Methods of detecting tumor markers include, but are not limited to: Immunohistochemical IHC Analysis — Laboratory process of detecting an organism in tissues with antibodies.

Gene mutation testing can purportedly be used to find somatic mutations in cancerous cells that are not inherited. Some examples of genes that may have somatic mutations include: Individualized molecular tumor profiling is a laboratory method of testing a panel of tumor markers, which may include genetic as well as biochemical markers, to establish a personalized molecular profile of a tumor to recommend treatment options.

Next-generation sequence NGS tests use select genes to purportedly identify molecular growth drivers for improved risk stratification and targeted therapies. Liquid biopsy refers to serum testing for DNA fragments that are shed by cancer cells and released into the bloodstream. The laboratory performs an algorithmic analysis using the results of the assays and sometimes other information, such as sex and age and converts the information into a numeric score, which is conveyed on a laboratory report.

Generally, MAAAs are exclusive to a single laboratory which owns the algorithm. MAAAs have been proposed for the evaluation of pelvic masses, including assisting in the determination of referral for surgery to a gynecologic oncologist or to a general surgeon. Topographic genotyping eg, PathFinderTG is a test that examines a panel of 15 to 20 genetic markers in tissue biopsy or other tissue specimens to purportedly aid in the determination of indeterminate or equivocal cancer diagnoses.

Elevated levels of Prostate-Specific Antigen PSA may also be found in the blood of men with benign prostate conditions, such as prostatitis and benign prostatic hyperplasia BPH. While PSA does not allow distinction between benign prostate conditions and cancer, an elevated PSA level may indicate that other tests are necessary to determine whether cancer is present. PSA levels have been shown to be useful in monitoring the effectiveness of prostate cancer treatment, and in checking for recurrence after treatment has ended.

Use of PSA for screening remains very controversial. Although researchers are in the process of studying the value of PSA along with digital rectal exams for routine screening of men ages 55 to 74 for prostate cancer; and the literature does not show at this time whether using PSA to screen for prostate cancer actually does reduce the number of deaths caused by this cancer.

The American Cancer Society recommends clinicians and patients consider screening with PSA and digital rectal exam for African American men and men with familial tendency age 40 or older and all men age 50 or older. The guidelines state that "[c]urrent evidence shows that PSA kinetics does not reliably predict disease stability or reclassification to higher risk state.

There was conflicting evidence whether PSA is a good predictor of disease progression or reclassification. PSA monitoring is considered a necessary component of an AS protocol, but a rising PSA may be best viewed as a trigger for reappraisal e. This gene has been investigated as a potential diagnostic marker for prostate cancer. However, there are no published clinical outcome studies of the effectiveness of the PCA3 gene in screening, diagnosis or management of prostate cancer.

The main target population of this non-invasive test is men with raised PSA but a negative prostate biopsy. Other target groups include men with a slightly raised PSA, as well as men with signs and symptoms suggestive of prostate cancer. These investigators performed a multi-center study to validate the diagnostic performance of the PCA3 urine test established in an earlier single-institution study. These researchers determined the PCA3 score in these samples and correlated the results with the results of the prostate biopsies.

The area under the receiver-operating characteristic curve, a measure of the diagnostic accuracy of a test, was 0. The authors concluded that the findings of this multi-center study validated the diagnostic performance of the PCA3 urine test in the largest group studied thus far using a PCA3 gene-based test. Marks and associates examined the potential utility of the investigational PCA3 urine assay to predict the repeat biopsy outcome.

The RNA yield was adequate for analysis in the urine samples from of men i. Receiver operating characteristic curve analysis yielded an area under the curve of 0.

In contrast, the area under the curve for serum PSA was 0. The authors concluded that in men undergoing repeat prostate biopsy to rule out cancer, the urinary PCA3 score was superior to serum PSA determination for predicting the biopsy outcome.

The high specificity and informative rate suggest that the PCA3 assay could have an important role in prostate cancer diagnosis. Groskopf et al reported that the PCA3 score is independent of prostate volume and was highly correlated with the risk of positive biopsy. The PCA3 test was performed on men scheduled for prostate biopsy.

Haese et al presented preliminary results from a European multicenter study of PCA3. Enrolled patients had a PSA level of less than or equal to 2. In a review on biomarkers for prostate cancer detection, Parekh, et al. Tosoian et al evaluated the relationship between PCA3 and prostate biopsy results in men in a surveillance program.

Urine specimens were obtained from men with prostate cancer enrolled in the Johns Hopkins surveillance program. Patients with progression on biopsy The authors concluded that in men with low risk prostate cancer who were carefully selected for surveillance the PCA3 score was not significantly associated with short-term biopsy progression.

They stated that further analysis is necessary to assess the usefulness of PCA3 in combination with other biomarkers or in selected subsets of patients undergoing surveillance. While there are studies examining the positive and negative predictive values of the PCA3 urine assay, there is currently a lack of evidence of the effect of this test on management of individuals with or suspected of prostate cancer. The PCA3 urine assay shows promise as a prostate cancer diagnostic tool, however, more research is needed to ascertain the clinical value of this assay for screening and diagnostic purposes.

For all other settings, comparators, and outcomes, there was insufficient evidence. The EGAPP Working Group found insufficient evidence to recommend PCA3 testing in men with cancer-positive biopsies to determine if the disease is indolent or aggressive in order to develop an optimal treatment plan.

However, further study data will be needed before such markers can be used in standard clinical practice. In a Lancet review of prostate cancer, Attard, et al. To be clinically effective, it must be shown that thymosin B15 is released by the tumor into body fluids in detectable concentrations.

These researchers developed a quantitative assay that can measure clinically relevant levels of thymosin B15 in human urine. One antibody, having stable characteristics over the wide range of pH and salt concentrations found in urine and minimal cross-reactivity with other beta thymosins, was used to develop a competitive enzyme-linked immunosorbent assay ELISA.

No cross-reactivity with other urine proteins was observed. A stable thymosin B15 signal was recovered from urine specimens stored at degrees C for up to 1 year. The authors concluded that an ELISA that is able to detect thymosin B15 at clinically relevant concentrations in urine from patients with prostate cancer has been established. They noted that the assay will provide a tool for future clinical studies to validate urinary thymosin B15 as a predictive marker for recurrent prostate cancer.

CEA Carcinoembryonic antigen CEA is a normal cell product that is over-expressed by adenocarcinomas, primarily of the colon, rectum, breast, and lung. It is normally found in small amounts in the blood of most healthy people, but may become elevated in people who have cancer or some benign conditions.

CEA is an oncofetal glycoprotein present in the gastrointestinal tract and body fluids of the embryo and fetus Chin, et al.

It is also present in certain adult gastrointestinal cells, including the mucosal cells of the colorectum, and small amounts are present in blood. Blood levels are often elevated in patients with disseminated cancers and in some patients with nonmalignant disease.

According to the available literature, the primary use of CEA is in monitoring colorectal cancer, especially when the disease has metastasized. CEA is also used after treatment to check for recurrence of colorectal cancer. However, the literature indicates a wide variety of other cancers can produce elevated levels of this tumor marker, including melanoma; lymphoma; and cancers of the breast, lung, pancreas, stomach, cervix, bladder, kidney, thyroid, liver, and ovary.

Elevated CEA levels can also occur in patients with non-cancerous conditions, including inflammatory bowel disease, pancreatitis, and liver disease. CA is a protein that is found more in ovarian cancer cells than in other cells. Approximately half of women with metastatic ovarian cancer have an elevated CA level. The Gynecologic Cancer Foundation, the Society of Gynecologic Oncologists, and the American Cancer Society have issued a consensus statement to promote early detection of ovarian cancer, which recommends that women who have symptoms, including bloating, pelvic or abdominal pain, difficulty eating or feeling full quickly, and urinary frequency and urgency, are urged to see a gynecologist if symptoms are new and persist for more than three weeks ACS, ; SGO, Ovarian cancer is among the deadliest types of cancer because diagnosis usually comes very late, after the cancer has spread.

It is estimated that 22, new cases and 15, deaths will be reported in ACS, The consensus statement recommendations are based on studies that show the above symptoms appeared in women with ovarian cancer more than in other women Goff, et al. The recommendations acknowledge that there is not consensus on what physicians should do when patients present with these symptoms. According to a consensus statement issued by the Gynecologic Cancer Foundation, pelvic and rectal examination in women with the symptoms is one first step.

If there is a suspicion of cancer, the next step may be a transvaginal ultrasound to check the ovaries for abnormal growths, enlargement, or telltale pockets of fluid that can indicate cancer.

Testing for CA levels should also be considered. There is no evidence available that measurement of CA can be effectively used for widespread screening to reduce mortality from ovarian cancer, nor that the use of this test would result in decreased rather than increased morbidity and mortality.

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Validating whole slide imaging for diagnostic purposes in pathology

Background A tumor marker is a substance such as a protein, antigen or hormone in the body that may indicate the presence of cancer. Generally, these markers are specific to certain types of cancer and can be detected in blood, urine and tissue samples. The body may produce the marker in response to cancer or the tumor itself may produce the marker. The detection of tumor markers may be used to determine a diagnosis or as an indicator of disease cancer progression.

It can also be used to document clinical response to treatment. Tumor markers are normally produced in low quantities by cells in the body. Detection of a higher-than-normal serum level by radioimmunoassay or immunohistochemical techniques usually indicates the presence of a certain type of cancer.

Currently, the main use of tumor mfarkers is to assess a cancer's response to treatment and to check for recurrence. In some types of cancer, tumor marker levels may reflect the extent or stage of the disease and can be useful in predicting how well the disease will respond to treatment.

A decrease or return to normal in the level of a tumor marker may indicate that the cancer has responded favorably to therapy. If the tumor marker level rises, it may indicate that the cancer is spreading. Finally, measurements of tumor marker levels may be used after treatment has ended as a part of follow-up care to check for recurrence. However, in many cases the literature states that measurements of tumor marker levels alone are insufficient to diagnose cancer for the following reasons: Examples of tumor markers include: High levels of B2M are an indicator of certain kinds of cancer, including chronic lymphocytic leukemia, non-Hodgkin's lymphoma and multiple myeloma or kidney disease; Beta Human Chorionic Gonadotropin beta HCG — A type of tumor marker that may be found in higher than normal amounts in individuals with some types of cancer; Calcitonin — Hormone secreted by the thyroid that lowers blood calcium; Calretinin — A calcium-binding protein that is used as a marker in the evaluation of lung cancer and other diseases.

Chromogranin A — A protein found inside neuroendocrine cells, which releases chromogranin A and other hormones into the blood. Chromogranin A may be found in higher than normal amounts in individuals with certain neuroendocrine tumors, small cell lung cancer, prostate cancer and other conditions Guanylyl cyclase c GCC — An enzyme that may be expressed only in the cells that line the intestine from the duodenum to the rectum.

Inhibin — One of two hormones designated inhibin-A and inhibin-B secreted by the gonads by Sertoli cells in the male and the granulosa cells in the female and inhibits the production of follicle-stimulating hormone FSH by the pituitary gland; Lactate Dehydrogenase LDH — Marker used to monitor treatment of testicular cancer; Mucin-1 MUC-1 — Carbohydrate antigen elevated in individuals with tumors of the breast, ovary, lung and prostate as well as other disorders; Napsin A — Protein used as a marker in the evaluation of lung cancer; Prealbumin — Marker of nutritional status and a sensitive indicator of protein synthesis.

Levels of PSA in the blood often increase in men with prostate cancer. Thyroglobulin — Protein found in the thyroid gland. Some thyroglobulin can be found in the blood and this amount may be measured after thyroid surgery to determine whether thyroid cancer has recurred; Thyroid Transcription Factor-1 TTF-1 — A protein that is used as a tumor marker in the evaluation of lung cancer; Transferrin — A protein in blood plasma that carries iron derived from food intake to the liver, spleen and bone marrow.

Tumors may be evaluated with histology, which involves examination of the structure, especially the microscopic structure, of organic tissues. Methods of detecting tumor markers include, but are not limited to: Immunohistochemical IHC Analysis — Laboratory process of detecting an organism in tissues with antibodies. Gene mutation testing can purportedly be used to find somatic mutations in cancerous cells that are not inherited. Some examples of genes that may have somatic mutations include: Individualized molecular tumor profiling is a laboratory method of testing a panel of tumor markers, which may include genetic as well as biochemical markers, to establish a personalized molecular profile of a tumor to recommend treatment options.

Next-generation sequence NGS tests use select genes to purportedly identify molecular growth drivers for improved risk stratification and targeted therapies. Liquid biopsy refers to serum testing for DNA fragments that are shed by cancer cells and released into the bloodstream. The laboratory performs an algorithmic analysis using the results of the assays and sometimes other information, such as sex and age and converts the information into a numeric score, which is conveyed on a laboratory report.

Generally, MAAAs are exclusive to a single laboratory which owns the algorithm. MAAAs have been proposed for the evaluation of pelvic masses, including assisting in the determination of referral for surgery to a gynecologic oncologist or to a general surgeon. Topographic genotyping eg, PathFinderTG is a test that examines a panel of 15 to 20 genetic markers in tissue biopsy or other tissue specimens to purportedly aid in the determination of indeterminate or equivocal cancer diagnoses.

Elevated levels of Prostate-Specific Antigen PSA may also be found in the blood of men with benign prostate conditions, such as prostatitis and benign prostatic hyperplasia BPH.

While PSA does not allow distinction between benign prostate conditions and cancer, an elevated PSA level may indicate that other tests are necessary to determine whether cancer is present. PSA levels have been shown to be useful in monitoring the effectiveness of prostate cancer treatment, and in checking for recurrence after treatment has ended.

Use of PSA for screening remains very controversial. Although researchers are in the process of studying the value of PSA along with digital rectal exams for routine screening of men ages 55 to 74 for prostate cancer; and the literature does not show at this time whether using PSA to screen for prostate cancer actually does reduce the number of deaths caused by this cancer.

The American Cancer Society recommends clinicians and patients consider screening with PSA and digital rectal exam for African American men and men with familial tendency age 40 or older and all men age 50 or older. The guidelines state that "[c]urrent evidence shows that PSA kinetics does not reliably predict disease stability or reclassification to higher risk state.

There was conflicting evidence whether PSA is a good predictor of disease progression or reclassification. PSA monitoring is considered a necessary component of an AS protocol, but a rising PSA may be best viewed as a trigger for reappraisal e. This gene has been investigated as a potential diagnostic marker for prostate cancer. However, there are no published clinical outcome studies of the effectiveness of the PCA3 gene in screening, diagnosis or management of prostate cancer.

The main target population of this non-invasive test is men with raised PSA but a negative prostate biopsy. Other target groups include men with a slightly raised PSA, as well as men with signs and symptoms suggestive of prostate cancer. These investigators performed a multi-center study to validate the diagnostic performance of the PCA3 urine test established in an earlier single-institution study. These researchers determined the PCA3 score in these samples and correlated the results with the results of the prostate biopsies.

The area under the receiver-operating characteristic curve, a measure of the diagnostic accuracy of a test, was 0. The authors concluded that the findings of this multi-center study validated the diagnostic performance of the PCA3 urine test in the largest group studied thus far using a PCA3 gene-based test. Marks and associates examined the potential utility of the investigational PCA3 urine assay to predict the repeat biopsy outcome. The RNA yield was adequate for analysis in the urine samples from of men i.

Receiver operating characteristic curve analysis yielded an area under the curve of 0. In contrast, the area under the curve for serum PSA was 0. The authors concluded that in men undergoing repeat prostate biopsy to rule out cancer, the urinary PCA3 score was superior to serum PSA determination for predicting the biopsy outcome.

The high specificity and informative rate suggest that the PCA3 assay could have an important role in prostate cancer diagnosis. Groskopf et al reported that the PCA3 score is independent of prostate volume and was highly correlated with the risk of positive biopsy. The PCA3 test was performed on men scheduled for prostate biopsy. Haese et al presented preliminary results from a European multicenter study of PCA3. Enrolled patients had a PSA level of less than or equal to 2. In a review on biomarkers for prostate cancer detection, Parekh, et al.

Tosoian et al evaluated the relationship between PCA3 and prostate biopsy results in men in a surveillance program. Urine specimens were obtained from men with prostate cancer enrolled in the Johns Hopkins surveillance program. Patients with progression on biopsy The authors concluded that in men with low risk prostate cancer who were carefully selected for surveillance the PCA3 score was not significantly associated with short-term biopsy progression.

They stated that further analysis is necessary to assess the usefulness of PCA3 in combination with other biomarkers or in selected subsets of patients undergoing surveillance.

While there are studies examining the positive and negative predictive values of the PCA3 urine assay, there is currently a lack of evidence of the effect of this test on management of individuals with or suspected of prostate cancer. The PCA3 urine assay shows promise as a prostate cancer diagnostic tool, however, more research is needed to ascertain the clinical value of this assay for screening and diagnostic purposes.

For all other settings, comparators, and outcomes, there was insufficient evidence. The EGAPP Working Group found insufficient evidence to recommend PCA3 testing in men with cancer-positive biopsies to determine if the disease is indolent or aggressive in order to develop an optimal treatment plan. However, further study data will be needed before such markers can be used in standard clinical practice. In a Lancet review of prostate cancer, Attard, et al.

To be clinically effective, it must be shown that thymosin B15 is released by the tumor into body fluids in detectable concentrations. These researchers developed a quantitative assay that can measure clinically relevant levels of thymosin B15 in human urine.

One antibody, having stable characteristics over the wide range of pH and salt concentrations found in urine and minimal cross-reactivity with other beta thymosins, was used to develop a competitive enzyme-linked immunosorbent assay ELISA. No cross-reactivity with other urine proteins was observed. A stable thymosin B15 signal was recovered from urine specimens stored at degrees C for up to 1 year.

The authors concluded that an ELISA that is able to detect thymosin B15 at clinically relevant concentrations in urine from patients with prostate cancer has been established. They noted that the assay will provide a tool for future clinical studies to validate urinary thymosin B15 as a predictive marker for recurrent prostate cancer. CEA Carcinoembryonic antigen CEA is a normal cell product that is over-expressed by adenocarcinomas, primarily of the colon, rectum, breast, and lung.

It is normally found in small amounts in the blood of most healthy people, but may become elevated in people who have cancer or some benign conditions. CEA is an oncofetal glycoprotein present in the gastrointestinal tract and body fluids of the embryo and fetus Chin, et al. It is also present in certain adult gastrointestinal cells, including the mucosal cells of the colorectum, and small amounts are present in blood. Blood levels are often elevated in patients with disseminated cancers and in some patients with nonmalignant disease.

According to the available literature, the primary use of CEA is in monitoring colorectal cancer, especially when the disease has metastasized.

CEA is also used after treatment to check for recurrence of colorectal cancer. However, the literature indicates a wide variety of other cancers can produce elevated levels of this tumor marker, including melanoma; lymphoma; and cancers of the breast, lung, pancreas, stomach, cervix, bladder, kidney, thyroid, liver, and ovary.

Elevated CEA levels can also occur in patients with non-cancerous conditions, including inflammatory bowel disease, pancreatitis, and liver disease. CA is a protein that is found more in ovarian cancer cells than in other cells. Approximately half of women with metastatic ovarian cancer have an elevated CA level. The Gynecologic Cancer Foundation, the Society of Gynecologic Oncologists, and the American Cancer Society have issued a consensus statement to promote early detection of ovarian cancer, which recommends that women who have symptoms, including bloating, pelvic or abdominal pain, difficulty eating or feeling full quickly, and urinary frequency and urgency, are urged to see a gynecologist if symptoms are new and persist for more than three weeks ACS, ; SGO, Ovarian cancer is among the deadliest types of cancer because diagnosis usually comes very late, after the cancer has spread.

It is estimated that 22, new cases and 15, deaths will be reported in ACS, The consensus statement recommendations are based on studies that show the above symptoms appeared in women with ovarian cancer more than in other women Goff, et al. The recommendations acknowledge that there is not consensus on what physicians should do when patients present with these symptoms. According to a consensus statement issued by the Gynecologic Cancer Foundation, pelvic and rectal examination in women with the symptoms is one first step.

If there is a suspicion of cancer, the next step may be a transvaginal ultrasound to check the ovaries for abnormal growths, enlargement, or telltale pockets of fluid that can indicate cancer. Testing for CA levels should also be considered.

There is no evidence available that measurement of CA can be effectively used for widespread screening to reduce mortality from ovarian cancer, nor that the use of this test would result in decreased rather than increased morbidity and mortality.

Validating whole slide imaging for diagnostic purposes in pathology

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