Pathology Raw Data in Nonclinical Laboratory Studies for the Pharmaceutical Industry: The Pathologists View

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1 Pathology Raw Data in Nonclinical Laboratory Studies for the Pharmaceutical Industry: The Pathologists View Frank J. Geoly* and Roy L. Kerlin Pfizer Global Research and Development, Worldwide Safety Sciences, Groton Laboratories, Eastern Point Road, MS , Groton, CT 06340, USA Summary Post-mortem pathology data from nonclinical laboratory studies contain critical information for characterizing a new drug s safety and are pivotal for gaining an appreciation of potential hazards to humans. An understanding of the historical regulatory and scientific perspective on the nature of pathology data is valuable for assuring appropriate and useful audits of pathology material within nonclinical studies. This discussion reviews the specialty of pathology, the process of medical diagnosis, the definitions of raw data as they pertain to histopathology data, and the rationale behind the consensus decisions made by industry and the Food and Drug Administration (FDA) in interpreting good laboratory practice (GLP) regulations for this type of data. Copyright 2004 John Wiley & Sons, Ltd. Key Words phathology; histopathology; peer review; diagnoses; nonclinical; raw data; GLP; FDA; quality assurance Introduction In the process of pharmaceutical development, nonclinical laboratory animal studies are used to *Correspondence to: Frank J. Geoly, Pfizer Global Research and Development, Worldwide Safety Sciences, Groton Laboratories, Eastern Point Road, MS , Groton, CT 06340, USA. frank_j_geoly@groton.pfizer.com obtain preliminary information about the safety and toxicity of experimental drugs. The results from these studies are used to support applications to regulatory agencies for permission to test new, experimental drugs in people. In order to assure the accuracy of the data generated in these studies, the conduct and reporting of nonclinical studies is regulated in the US by the Good Laboratory Practice Regulations (Code of Federal Regulations (CFR), Title 21, Part 58) of the Food and Drug Administration. The post-mortem pathology data from these animal studies is often the most critical data used to characterize a new drug s safety and any potential risks to humans. A working knowledge of the discipline of pathology and how pathology data are collected allows the quality assurance professional to conduct audits of this data more effectively. The purpose of the current discussion is to review important concepts about pathology and collection of pathology data, and to review the history behind the regulatory definition of microscopic pathology raw data (histopathology data) in the Good Laboratory Practice Regulations. The Specialty of Pathology Many people equate the practice of pathology with the autopsy (more appropriately a term used for human post-mortem examination, but alternatively called a necropsy or post-mortem examination for animal studies). While the post-mortem examination may be a significant part of the pathologists responsibilities, in clinical medicine DOI: /qaj.278

2 162 F. J. Geoly and R. L. Kerlin pathologists also play an important role in the diagnosis of diseases of living patients by evaluating tissue biopsies and analyzing blood and other bodily fluids. Specialists in pathology (pathologists) are generally physicians or veterinarians with specialized training and experience in the identification of the structural changes in organs, tissues, and cells that lead to clinical diseases. In other words, pathologists are specialists in identifying morphologic, molecular or biochemical abnormalities in organs that underlie disease. Their most important tools for performing these evaluations are their eyes, often aided by a microscope, and the sum of their knowledge and experience. Changes or abnormalities in organs may be referred to as lesions and, once identified, these findings are designated a concise technical description or diagnosis. Determining a diagnosis for some lesions may be straightforward. At the other extreme, diagnosis may be a very complex and ultimately subjective process, as will be discussed later on in this review. Once a lesion is diagnosed, pathologists may also suggest the functional consequences of the lesion (i.e. biological effects), a prognosis, and sometimes its cause or mechanism. The research-based pharmaceutical and biotech industries draw on the expertise of pathologists for evaluation and interpretation of nonclinical studies. The majority of pathologists in this setting are veterinary pathologists who have expertise with the diseases of animals, animal models of human disease, and the comparative aspects between animal disease and human disease. The data generated by the pathologist, and the interpretation of that data, is one of the most important parts of characterizing the toxicity and evaluating the safety of a new drug. Raw Data in Nonclinical Studies Raw data in nonclinical studies is defined under the GLP regulations as the recording of the original observations, which are necessary for reconstruction and evaluation of the report of a study [1]. Any changes or modifications made to these original observations must also be made in strict accordance with the GLP regulations, and require the date, time, the identity of the person making the change, and the reasons for making the change, all without obscuring the original data [2]. The typical nonclinical study is conducted by administering the new drug to laboratory animals daily for a specified period of time, ranging from 1 day to 2 years depending upon the purpose of the study. During the in-life phase of the study, the animals are observed for any symptoms, and blood, urine or other samples may be collected for

3 Pathology Raw Data in Nonclinical Laboratory Studies 163 analysis. At the end of the study the animals are euthanatized and a careful post-mortem examination (necropsy) is conducted by the pathologist, which includes gross (visible to the naked eye) and subsequent microscopic examination of the organs. The pathology data that is generated can be broadly subdivided into quantitative data (objective or numerical data) and qualitative data (subjective or categorical data). Table 1 summarizes some of the more common pathology data and documentation collected in a nonclinical study. Examples of the quantitative data and documentation include the numerical output from automated blood analyzers, necropsy sheets (verifying the animals identity and any gross lesions), tissue checklists, body and organ weights, and histological slide preparation documentation. Auditing of this quantitative pathology data is relatively uncomplicated, but is necessary to assure that the pathology examination is conducted according to the study protocol, and to assure that animals and their tissues are identified accurately. Table 1: Pathology data and documentation in a nonclinical dtudy In-life pathology data (blood and urine analysis) Complete blood count (CBC) Blood cell morphology Blood chemistry analysis Urinalysis Other specialized blood or urine measurements Necropsy pathology documentation and data Animal identification Terminal body and organ weights Gross observations/findings Tissues collected (check list) Histological slide preparation documentation Wet tissue identification Paraffin blocks Histological slides Correlation between identification of wet tissues paraffin blocks slides Histopathology data Microscopic diagnoses/histopathology tables Correlation of microscopic diagnoses and gross findings But some issues have historically arisen in auditing the qualitative data, which are principally the microscopic diagnoses (the histopathology data) in the pathology tables. These diagnoses are the basis for many of the conclusions of the study, and they are many times not the result of the pathologist s original observations. Instead, generation of some microscopic diagnoses is an iterative process in which the diagnosis is continually refined and modified until a final diagnosis is reached. Recognizing this complexity in the diagnostic process, microscopic pathology raw data are considered the signed and dated final report of the study pathologist [3]. The issues that arise from a quality assurance perspective are obvious. If changes are being made to the recorded original observations, why should they not be tracked? If the pathologist is continually modifying the data, then how can we know the pathology data is accurate and truly reflects what was happening in the study? Does this invite an opportunity to falsify data? When, if ever, do we begin auditing changes in the data? Why is this practice allowed in the GLP regulations? The remainder of the current discussion will focus on these issues surrounding histopathology data, and how the FDA historically addressed them in formulating the GLPs. Medical Diagnosis An understanding of the practice of diagnostic pathology, and the science and art of making a diagnosis is key to understanding these issues. As in other areas in medicine, a diagnosis represents the pathologist s opinion using all the available data, made in the context of that individual s training and experience. As new information (data) is gathered the diagnosis may be further refined or may change altogether. The pathologist s ultimate goal is to accurately communicate his/her observations so that it is clear to a reader of the report what was observed in the study. The word choice, which becomes the diagnosis, is critical to accomplishing that goal. In Shakespeare s Romeo and Juliet, Juliet recites the familiar line, What s in a name? That

4 164 F. J. Geoly and R. L. Kerlin which we call a rose by any other name would smell as sweet, in reference to Romeo s family name. Albeit less romantic, the process of assigning a microscopic diagnosis to a lesion embraces that same concept. That is, if you mistakenly called the rose a carnation, it would not change the fact that it is a rose. Alternatively, the rose might also be called a flower, which is not incorrect, but is less specific and less informative. The true nature of a lesion (or a rose) is a constant; the diagnosis (or name) we assign to it is subjective but critical to communicating to others the nature of the lesion. Ultimately, the diagnosis of a lesion is based on the pathologist s knowledge of disease and its appropriate nomenclature. The rose as lesion analogy can be advanced further to describe how the pathologist reaches a final diagnosis in a nonclinical study. In reality, most people would be able to identify a rose in a garden without difficulty. Most of the diagnoses a pathologist generates in a given study fall into this category. On occasion, however, there is an unusual flower in the garden (a lesion in an organ) that is difficult to classify, and the best a person can do after his original observation is to call it a flower. After examining the garden for more flowers of similar form (other organs for lesions), other gardens in the area (other animals in the study), reading gardening textbooks, or consulting other gardeners (peer review), the diagnosis may be modified from flower to rose. On the other hand, if insufficient evidence is available to more specifically classify the lesion the diagnosis may remain as flower. This example, in essence, describes the unaudited process of generating the microscopic diagnoses in a preclinical study. While most diagnoses are straightforward, others are more complex. What is really important is documenting the presence of the microscopic finding, and that the final diagnosis accurately defines the characteristics of this finding to the greatest extent possible. The full thought process behind reaching the final diagnosis is not important for describing the microscopic finding. It is only the final signed pathology tables that are important, as these document the pathologist s ultimate professional opinion about the changes observed histologically. Peer Review The questions that should remain are: How do we know the pathologist has diagnosed the lesions acceptably? What if, in the above example, the rose really is a carnation? Has the pathologist unintentionally recorded an inaccurate diagnosis? These questions of data quality are also very important for the pathologists involved in these studies. As such, many organizations have instituted some form of pathologist peer review of studies [4]. This is a process where a second pathologist (the peer review pathologist ) examines a subset of animals from the study to verify that the diagnoses and interpretations of the primary study pathologist are sound according to contemporary practice, and that any toxicologically significant lesions have not been overlooked. Documented peer review is equivalent to a formal second opinion. The ultimate goal of the process should be to assure the development of accurate pathology data [4]. During the process of peer review there may be differences of opinion in the diagnosis of a lesion between the peer-review pathologist and the study pathologist. Often these differences of opinion are easily resolved by discussion between the pathologists resulting in a consensus diagnosis that may (or may not) be different in wording. Rarely, the peer review pathologist will notice a change that results in adding a diagnosis in an organ that was initially described as normal by the study pathologist. Occasionally, the study pathologist and peer review pathologist cannot come to an agreement on a particular diagnosis so mechanisms to resolve the differences must also be in place, including consultation with additional pathologists. Thus, all that transpires during a peer review process is a process whereby two professionals confer and come to an agreement about the histological changes recorded for the study and their interpretation. Any changes that result from this consultation are made before the data are made final. Therefore, the final pathology report (the raw data) reflects a consensus between the study pathologist and the peer-review pathologist, but the specific details of the modifications made to

5 Pathology Raw Data in Nonclinical Laboratory Studies 165 the data are not recorded, so are not subject to audit. The Society of Toxicologic Pathologists published a position paper in 1997 on what information should be included in peer review documentation [5]. The documentation should include information about who conducted the peer review, how it was done, and a statement that the final signed pathology report (the raw data) represents a consensus of the primary and reviewing pathologists. If the pathologists did not agree, a description of the process involved in resolving those differences should also be included. It is the position of the Society that detailed findings of the primary and peer review pathologists need not be retained, which is in accord with the GLP regulations [3], since such information is not necessary for the reconstruction of the study and therefore is not considered to be raw data. GLP Regulations on Histopathology Raw Data The FDA considers histopathology raw data as the signed and dated report of the diagnostic pathologist, respecting the sometimes complex process of reaching a microscopic diagnosis. Some have considered this definition of raw data to be at odds with GLP regulations [6], but close analysis of the language, reveals that this does conform to the regulations. Recall that the official definition of raw data has two components: (1) the recording of original observations, and (2) that these observations are necessary to reconstruct the report of the study. The origins of the debate came in the FDA s management briefings in 1978 prior to the effective date of the GLPs [6]. There was concern expressed by industry managers and pathologists that the interim notes and diagnoses made by pathologists would have to be considered raw data under the language of the regulations. However, the agency wisely recognized the complexity of the diagnostic process and was in agreement with the concerns that were expressed. The policy set at the time was to consider raw data as the signed and dated report of the pathologist [3]. Over the next 9 years, uneasiness spread inside the FDA. Many thought that this policy was an invitation to falsify data [6]. In order to insure that the correct decision was made, the case was re-opened and the issue re-examined in order to clarify the position. The FDA s final rule on the GLPs in 1987 used the following reasoning in reference to pathology raw data: Although the notes taken by the pathologist during histopathologic examination of slides are indeed the result of original observations, these notes are not necessary for the reconstruction and evaluation of the final report. The final report is evaluated by an analysis of the pathology syndrome as described in the pathologist s report, which is required under Section (a) (12). Further, because Section (a) requires histopathological blocks, tissues, and slides to be retained as specimens, the final report can be reconstructed by verification of the pathology findings by a second pathologist or by a team of pathologists. The pathologists interim notes, therefore, which are subject to frequent changes as the pathologist refines the diagnosis are not raw data because they do not contribute to study reconstruction. Accordingly, only the signed and dated report of the pathologist comprises the raw data respecting the histopathological evaluation of tissue specimens [3]. A key element of this reasoning that allowed the FDA a degree of comfort is the GLP requirement of retention of pathology specimens. Specimen retention allows all microscopic pathology findings to be confirmed independently at any time. The pathology data is permanent. Pathology data is unlike observations of clinical signs or symptoms during the in-life phase of the study, which are ephemeral, and can never be verified at a later time. The second key element is the acknowledgement that the original observations of the pathologist are not important for the reconstruction of the final report. In other words, even if the interim notes were to be audited, there would not be a valid purpose or use for such information. The pathologist would

6 166 F. J. Geoly and R. L. Kerlin not defend anything that differed from the final signed tables, because these have been crafted to best describe the tissue changes defined in the final diagnoses. 21 CFR Part 11 (electronic records/signatures) and Future Considerations The previous discussion applied to standard types of pathology raw data with results recorded either on paper or more commonly in electronic databases. In the latter case, results in electronic format are also subject to the regulations pertaining to electronic records (21 CFR Part 11). 21 CFR Part 11, which only pertains to the reliability of electronic records, is meant to be applied subordinate to the predicate rule of 21 CFR Part 58 (the GLP regulations). In other words, since the GLP regulations do not require audit of interim pathologist notes and diagnoses, changes made to interim diagnoses in an electronic database are not subject to an electronic audit trail until the pathology data is recorded as final in the database. Understanding this concept is also important with regard to the capture, storage, and data derived from microscopic images, such as morphometric measurements. If such measurements are used to develop new data, then the images and measurements should be considered raw data, and 21 CFR Part 58 as well as Part 11 regulations should apply. If, on the other hand, an image is simply a photographic example of a diagnosis that is already in the pathology tables, then it should not be considered raw data, since it is not necessary for the reconstruction of the study. Development of new computerized and electronic technologies in pathology is constantly ongoing, including the computerized analysis of histologic sections of organs. These computerized systems are being developed in order to screen all histologic tissue sections for a given study, and determine which tissues are normal, and which ones are abnormal and need to be evaluated by the pathologist. Validating this technology, and other technologies, will present new quality assurance and regulatory challenges for both pathologists and quality assurance professionals for years to come. Summary Unlike other raw data, the final microscopic diagnoses in a pathology report are generally not the result of the pathologist s original observations. Instead, they are the result of repeated microscopic examination and may include the opinion of a second pathologist in the sometimes complex process of reaching a final diagnosis. Recognizing this process, the FDA considers pathology raw data from nonclinical studies as the signed and dated report of the pathologist (in accordance with the definition of raw data), and requires retention of pathology specimens so that the findings can, if necessary, be independently verified. The FDA ruled in 1987 that the interim diagnoses and notes of the pathologist need not be retained for audit because they were considered unimportant for reconstructing the study report. References 1. United States Code of Federal Regulations. Title 21. Part 58, Section 58.3(k). Revised July gov/scripts/cdrh/cfdocs/cfcfr/cfrsearch.cfm. 2. United States Code of Federal Regulations. Title 21, Part 58, Section (e). Revised July fda.gov/scripts/cdrh/cfdocs/cfcfr/cfrsearch.cfm. 3. United States Federal Register. Preamble to the Good Laboratory Practice Regulations 1987; 52(172) September 4, Society of Toxicologic Pathologists. Peer review in toxicologic pathology: some recommendations. Toxicol Pathol 1991; 19(3): Society of Toxicologic Pathologists. Documentation of pathology peer review. Toxicol Pathol 1997; 25(6): Lepore PD. Pathology raw data. Toxicol Pathol 1996; 24(1):147.