John D. Reith MD
Metastatic Tumors in Bone
Bone is one of the most common anatomic sites for metastases, along with the lungs and liver, and metastases are the most commonly diagnosed neoplasms encountered in the skeleton. Although most metastases encountered in the adult skeleton are either carcinoma or melanoma, sarcomas may also metastasize to bone, particularly in the pediatric population. The general surgical pathologist is far more likely to encounter a skeletal focus of metastatic cancer than any of the primary malignant bone tumors. Patients frequently present with signs and symptoms related to metastatic bone lesions before a primary tumor has come to clinical attention, and severe pain or a pathologic fracture is commonly the presenting complaint. Despite the often extensive immunohistochemical evaluation of metastatic deposits, a primary source can be elusive and sometimes never located. The incidence and localization, radiologic characteristics, patterns of spread, and histologic and immunohistochemical features of various metastatic lesions in bone are considered in some detail in this chapter. Because fine-needle aspiration cytology plays a major role in diagnosing metastatic lesions affecting the skeleton, we include in this chapter a brief description of cytologic features of metastatic tumors in bone. First, however, it is important to discuss some of the newer information on the biology of tumor metastasis that has been derived from the methods of molecular pathology. This information is important for many reasons, including the understanding of the pharmacologic strategies for treating bone metastases.
Biology of Metastases
The ability of tumor cells to invade within their host organ and their ability to metastasize to distant sites are two biologic hallmarks of malignancy. To successfully metastasize, tumor cells must (1) proliferate within the host organ and gain the ability to invade the surrounding tissue and matrix; (2) invade blood vessels, whether induced by angiogenic properties of the tumor cells or normal vessels at the site; (3) enter the circulatory system and survive both intracellular and extracellular influences attempting to destroy them; (4) adhere to and migrate across sinusoidal walls at distant sites; and (5) form deposits capable of surviving in the new environment10 (Fig. 19-1, A). The basic biology, molecular mechanisms, and genetic and epigenetic changes that drive these steps are extremely complex and probably differ depending on the source of the metastasis and specific type of tumor.
Before metastasizing, tumor cells prepare distant sites to receive metastatic deposits by creating a premetastatic niche.6,9 By modulating hematopoietic cells to increase fibronectin production and by directly producing factors that render the bone microenvironment receptive, tumors are able to prepare the bone for metastases. Such factors include osteopontin,2 matrix metalloproteinases, and parathyroid hormone-related protein (PTHrP).5 Once tumor cells have become disseminated, the local environment within the metastatic site, or metastatic niche, is important in determining whether tumor cells are able to survive.12,16
After induction neovascularization by tumor cells in the primary organ, the tumor cells invade into the vessels and are embolized to distant sites. Those tumor cells that are able to survive in transit become lodged in capillaries within a secondary site such as bone, adhere to the endothelium, and eventually extravasate through the capillary wall and into the stroma, where they proliferate and ultimately form clinically significant masses. Tumor cells home in to bone via the same protein interactions that hematopoietic stem cells use, relying on integrins, chemokines, bone morphogenetic proteins, and osteopontin, among others, to settle in bone.18
An important element of metastasis is the ability of tumor cells to both adhere to and degrade the extracellular matrix, invade blood vessels, and evade cell death, both within their primary organ and within bone.3,17 This is achieved through a complex interaction between integrins; extracellular matrix components such as type I collagen and fibronectin; and proteolytic enzymes such as matrix metalloproteinases (Fig. 19-2). Integrins, a superfamily of transmembrane receptors that modulate cell-to-cell and cell-to-matrix interactions by binding with a variety of ligands, play a key role in skeletal metastases on many levels, matrix and blood vessel invasion, osteoclast signaling, neovascularization, and colonization of bone.4,8,15,20 CD44, a hyaluronic acid receptor that has been studied in a wide variety of tumors, appears to be extremely important in the early development of metastases.19
The tumor cells must invade and destroy the extracellular matrix at several stages of the process to metastasize. Thus the interaction of tumor cells with extracellular matrix seems to be one of the major mechanisms of the metastatic cascade.3,17 The extracellular matrix can be divided into two main components: a basement membrane and interstitial connective tissue. The basement membrane is a sheetlike structure that separates epithelial and endothelial cells from the interstitial matrix. It contains type IV collagen and several distinct noncollagenous proteins, such as laminin, heparan, and sulfated proteoglycan. Laminin, together with collagen IV, is a major component of the basement membrane and serves as an integration unit of the structure. Laminin also plays a major role as a cellular adhesion molecule. This large complex molecule has three short arms containing collagen binding sites and a single long arm with a heparan binding site. The central region of the molecule, where all four arms are connected in a cross-shaped structure, contains the laminin receptor binding site for cells.
Recent advancements have elucidated some additional components of the so-called cell-to-cell and cell-to-matrix adhesion system and its potential role in invasive growth.8,13,15,17,20 A new class of transmembrane cell adhesion receptors (integrins) that has a unique structure and ability to bind fibronectin and laminin has been identified.13,15,17,20 These receptors integrate the extracellular matrix anchorage with an intracytoplasmic cytoskeleton and provide a pathway for signal transduction.
Collagen I and III are the most prominent components of the interstitial extracellular matrix.3,17 Proteoglycans fill the interstitial space among the collagen fibers, and their major role is to retain water and to maintain the shape and volume of the interstitial space. Fibronectin is one of the major noncollagenous components of the interstitial matrix. It is biochemically distinct from laminin but has a similar biologic function. It serves as a major adhesion molecule in the intracellular matrix.
To invade the intercellular matrix and to penetrate the vascular channels, the tumor cells have to follow three general steps:
2. Degradation of the matrix components must proceed through the action of proteolytic enzymes.
3. Tumor cells must continue to migrate into the degraded area of the extracellular matrix.
Major molecular phenomena associated with the development of the metastatic phenotype are summarized in Figure 19-1, B.
Paradoxically, it has been shown that tumor cells exhibiting increased levels of laminin and fibronectin surface receptors have a higher metastatic potential compared with those that have minimal levels of these receptors. Thus to develop metastatic foci, the tumor cells must maintain some degree of adherence to the intercellular matrix elements. In general, some insufficiency of cell adhesion and junction systems can be documented in most cancers.3,13,15,17
For tumor cells to occupy a territory of the stromal matrix, the existing elements of the stromal matrix have to be at least partly destroyed or degraded. Tumor cells that invade vessels have a higher capacity to degrade the stromal matrix than other tumor cells. The enzymatic activity of tumor cells plays an important role in the development of metastases. The involvement of several different proteases, including urokinase, plasminogen activator, cathepsins B and D, and various metalloproteases produced directly by tumor cells, play an important role in the invasive growth and development of metastasis.12 Virtually all proteases appear to be controlled by a cascade of complex-activating and complex-inhibiting factors. Therefore their roles in invasive growth and metastasis depend not only on the level of the enzyme, but also on the presence of adequate amounts of their activators and inhibitors. The major component of the basement membrane (type IV collagen) is degraded by a specific metalloprotease known as type IV collagenase. The activity of this enzyme can be correlated with the metastatic potential of several experimental and human tumors. Overall, an upregulation of multiple proteolytic enzymes of the so-called plasminogen cascade have been documented in malignant tumor cells and have been linked to their invasive and metastatic potential.
The ability to induce vascular growth is another factor that secures the survival of an enlarging tumor mass.13 It appears that the tumors that induce a rich vascular network have a higher metastatic potential. The ability of tumor cells to induce proliferation of vascular cells via a number of growth factors such as endothelial growth factor and fibroblastic growth factor has recently been extensively studied.3 It seems that tumors with higher levels of these factors are more clinically aggressive compared with those that have low levels of these factors. The tumor cells that invade the vessels and circulate in the lymph or blood interact with cellular and humoral components of the environment. The interaction of tumor cells with platelets and other blood clotting factors, both circulating and cell fixed, is an important element in the promotion of tumor cell thrombosis of peripheral sinusoids and growth of metastatic foci.13 In addition, the retention on tumor cells of a unique class of carbohydrate antigens that interact with endothelial surface antigens is a critical factor for settlement in metastatic foci. Moreover, the tumor cells of a metastatic focus must retain their stromal destructive activities and interact with other cells of the new environment to survive and form clinically detectable nodules.
Following the successful invasion and colonization of the bone, metastatic deposits most commonly induce osteolysis and less commonly new bone formation by interacting with complex bone resorption and formation regulatory pathways (Fig. 19-2). Osteolysis is driven by osteoclastic resorption of bone, the control of which comes under a variety of influences. For some metastatic carcinomas, particularly breast cancer, osteoclastogenesis is stimulated by a number of cytokines, one of the most important of which is PTHrP, an osteoclast activator.1,10 As osteoclasts resorb bone, TGFβ is released from bone matrix, resulting in increased expression of PTHrP by tumor cells13; PTHrP may also further increase tumor cell proliferation and act as an angiogenic factor.
PTHrP and other factors stimulate osteoblasts to produce the cytokine RANK-ligand (receptor activator of nuclear factor-κβ ligand), a member of the tumor necrosis factors, which binds to and activates the RANK receptor on osteoclast precursors, inducing both differentiation and activation of these bone resorbing cells. PTHrP also down regulates osteoprotegerin, a decoy receptor for RANKL.7 Blockade of the RANK-RANKL system is the mechanism by which the monoclonal antibody denosumab serves as an effective therapeutic agent in the treatment of metastatic carcinoma.11,14 Bisphosphonates—inhibitors of osteoclastic bone resorption—are also standard pharmacologic agents used for the care of patients with bone metastases, among other conditions.
Not all types of metastatic carcinoma induce osteolysis. Carcinomas of the prostate and breast, as well as neuroendocrine tumors, can induce predominately blastic or sclerotic metastases. The mechanisms that cause blastic metastases are not as well understood as those for osteolytic metastases. Factors such as endothelin-1, platelet-derived growth factor (PDGF), TGFβ2, insulin-like growth factor (IGF), and bone morphogenetic proteins (BMPs) all stimulate bone formation and are produced by various types of carcinomas. These factors induce bone formation by inducing osteoclast apoptosis while stimulating osteoblast differentiation and proliferation.
General Features of Skeletal Metastases
The skeleton is one of the most common sites for the metastasis of virtually all common human malignant neoplasms, and almost every malignant neoplasm has been described as being capable of metastasizing to bone.22,46 In general, metastatic neoplastic cells reach the bones through the complex arterial and venous systems. The blood supply to the skeleton represents a significant proportion of the body’s vasculature. In addition, the vertebral plexus of veins is valveless, and the retrograde venous pressure is often increased in the abdominal and chest regions. This enables a retrograde flow of blood to bypass the caval system and to reach the bones of the vertebral column.24 These two basic anatomic and physiologic features explain why metastases preferentially involve the bones of the axial skeleton.25 From a biologic point of view, it is very unlikely that the abundance of the vascular network within bone is the only factor that predisposes to metastasis because metastases rarely develop in other tissues that have an equally rich vascular network, such as the spleen. Thus the biologic conditions of bone tissue must also be important factors in promoting the growth of tumor cells that reach the marrow via the venous and arterial blood network.
Autopsy reports of large series of patients have shown that, with gross examination and limited sampling, skeletal metastases can be documented in 30% of patients who died of carcinoma, with particular carcinomas such as breast or prostate cancer present in nearly 85% of patients autopsied.21 These findings are supported by bone scintigraphy studies, which suggest that approximately 85% of patients with common carcinomas have skeletal involvement. This is in keeping with observations by Jaffe, who stated that if extensive skeletal sampling were to be performed, metastases could be documented in 70% of patients who died of carcinoma.36
Conventional radiographs are not particularly sensitive in identifying early metastatic lesions in the skeleton.53 Radioisotopic scanning can demonstrate abnormal uptake of bone-seeking isotope approximately 4 months on average before a lesion can be identified on plain radiographs.37 Radioisotopic scans, however, are not specific, and they identify an increased turnover state associated with osteoblastic activity rather than the proliferation of tumor cells. Therefore they may not detect metastatic tumors that are primarily associated with bone destruction and minimal or no osteoblastic activity. Diffusion-weighted magnetic resonance imaging (MRI) and whole-body fluorodeoxyglucose positron emission tomography/computed tomography (FDG-PET/CT) scans have been shown to be very sensitive and accurate in identifying metastatic deposits in bone from a variety of sources.33,51,52
Metastatic tumor in bone typically presents as a destructive focus that has an aggressive pattern of bone destruction (Fig. 19-3). Cortical disruption, extension into soft tissue, and periosteal new bone formation can be present.26,27,39,42,45 Some metastatic tumors can provoke an osteoblastic reaction with new bone formation and appear to be a densely sclerotic lesion32 (Fig. 19-4). Often, focal sclerosis is seen within lytic lesions, and therefore many skeletal metastases produce a mixture of lytic and blastic appearances. In general, a lytic versus blastic radiographic appearance of a metastatic tumor in bone results from a prevalent bone resorptive (destructive) or stimulating (osteoblastic) activity of the tumor.28,30,31,40
Multiple lesions are a radiographic hallmark of metastatic skeletal disease (Fig. 19-4). Although any skeletal site can be affected by metastasis, the majority of lesions involve the axial skeleton and proximal portions of the appendicular skeleton (Fig. 19-5). Consequently, such bones as the vertebrae, pelvis, ribs, skull, sternum, proximal femur, and humerus are most frequently involved. These sites correspond to areas that contain hematopoietic marrow, which has a rich sinusoidal vascular network. This feature and the presence of venous plexus connected with abdominal and thoracic organs may promote metastasis in these regions. Metastases that predominantly involve fatty marrow distal to elbow and knee joints and the mandible are unusual in adults.
Occasionally a single metastatic focus of carcinoma can be present. In some patients, it can be a presenting sign of a clinically silent primary tumor that, most often, is located within the thoracic or abdominal organs.29 Although a solitary skeletal metastasis can be a presenting sign in any type of common carcinoma, it is most frequent in carcinomas of the kidney, lung, breast, pancreas, thyroid, and colon.50 Rarely, bilateral symmetric metastases involving the left and right sides can be present.43,44 Bilateral, symmetric, osteoblastic metastases can simulate osteopoikilosis. The small tubular bones of the hands and feet are rarely affected by metastases34,38,41 (Fig. 19-6). This rare phenomenon more often involves the bones of the feet. Although it can occur in many common cancers, carcinoma of the lung is the most frequent malignancy in which so-called acral metastases occur. In general, acral metastases are most often seen in the small bones of the feet in highly aggressive malignant neoplasms of visceral and thoracic organs. In extremely rare instances, metastatic lesions can have a misleading radiographic appearance; that is, their radiographic appearance may support the diagnosis of a benign condition or a primary bone lesion.
Tenderness, swelling, and pain are typical presenting symptoms of skeletal metastases. The symptoms are insidious in onset and gradually increase in intensity over weeks to months, often preceding changes that are recognizable on conventional radiographs. An abrupt onset of symptoms (typically severe pain) is usually associated with pathologic fracture (Figs. 19-7 and 19-8). In general, the presence of skeletal metastases is a sign of disseminated multisystem disease, and other organs are likely to be involved. However, in a minority of patients, a solitary metastatic focus in the skeleton can be the only identifiable site of the disease. In such cases, a metastasectomy is advocated because it can significantly prolong a patient’s life.23,49 The management of skeletal metastases typically includes internal fixation or joint replacement for fractures or impending fractures, radiation and chemotherapy, and pain control. Pharmacologic therapy with bisphosphonates inhibits bone resorption by blocking osteoclastic activity and has become very important in the management of patients with metastases. These drugs improve bone stock in cases of metastatic carcinoma but do not improve overall patient survival.35
Because a metastasis to the skeleton is often a patient’s initial clinical manifestation of a carcinoma, the surgical pathologist may be asked to assist in identifying the source of the tumor. Although it is not possible to characterize the site of origin in all cases of metastatic carcinoma of unknown primary site (Fig. 19-9), some tumors, such as metastatic renal cell carcinoma and metastatic thyroid carcinomas, have characteristic light microscopic features. In cases of metastatic carcinoma lacking characteristic histologic findings, the use of a battery of commonly employed immunostains can often direct the clinical team to investigate a limited number of potential primary sites, if not identify the specific source for the metastasis. Some tumors, such as squamous cell carcinoma, lack a specific immunohistochemical profile that allows the distinction of one primary site from another with certainty. A focused history and physical examination coupled with a thorough radiographic evaluation of the chest, abdomen, and pelvis are also extremely valuable in identifying the source of metastases to the skeleton.47,48 Figure 19-10 represents an algorithm for the immunohistochemical evaluation of the commonly encountered metastases in bone.
Skeletal Metastases in Selected Common Neoplasms
In this section, some basic clinical, radiographic, and pathologic features of bone involvement in the most common malignant neoplasms are briefly discussed. The entities discussed in this section represent only the malignancies for which skeletal metastases are most frequently a presenting sign or occur during the course of the disease.
Carcinoma of the Lung
Lung cancer is a leading cause of death worldwide and the most frequently occurring human malignancy that has a high propensity to metastasize to a variety of distant sites.76 All major histologic subtypes of lung cancer—including small cell and non–small cell carcinomas—have a high tendency to metastasize to the skeleton, particularly the small cell types.70,77 Carcinoma of the lung typically produces lytic metastases (Fig. 19-11) and is the most frequent human tumor to present as a solitary metastatic lesion in bone; lung cancers are the most frequent tumor that metastasizes to the acral skeleton. Bilateral symmetric metastases that involve unusual sites, such as the patella, can also occur.71 Finally, lung cancer is often centered on the cortex of the appendicular skeleton and may cause unusual radiographic presentations such as entirely intracortical or subperiosteal lesions.
A great deal is known about the molecular alterations in lung cancer, which can impact the use of targeted therapies in the treatment of these patients. Epidermal growth factor receptor (EGFR), the anaplastic lymphoma kinase gene (ALK), and the K-RAS gene should all be evaluated in lung cancers, including metastatic deposits when the primary tumor has not been sampled. ALK gene rearrangements occur in young, nonsmoking patients with adenocarcinoma. Metastatic pulmonary adenocarcinoma may harbor mutations of the EGFR gene, and in such cases patients frequently respond favorably to tyrosine kinase inhibitors.63 Patients with K-RAS mutated adenocarcinoma may be resistant to tyrosine kinase inhibitors.
Aspirates from non–small cell metastatic lung cancer are highly cellular and contain large epithelial cells with pronounced atypia (Fig. 19-12, A). Keratinization and extensive necrosis are frequently seen. Dispersed cells with scanty cytoplasm, occasionally forming small three-dimensional clusters, characterize metastatic small cell carcinoma (Fig. 19-12, B). Nuclei of these cells have coarsely granular chromatin with a few larger chromocenters and no visible nucleoli. Positive immunohistochemical staining for thyroid transcription factor-1 (TTF-1) can be helpful to establish the origin of metastasis from lung. Small cell carcinomas typically show neuroendocrine differentiation and are positive for a wide range of neuroendocrine markers.
Gastrointestinal Carcinoma
Most carcinomas of the gastrointestinal tract are highly aggressive lesions with a high propensity for metastasis.75 Radioisotopic scans reveal abnormal uptake suggestive of metastasis in approximately 45% of patients who have stomach cancer.56 The sites most frequently involved are similar to those affected by other common neoplasms: the spine, ribs, pelvis, femur, and skull. Most metastases occur approximately 1 year after diagnosis in patients with stage III disease.73 Occasionally, skeletal metastasis, either as a solitary lesion or as multiple lesions, can be a presenting sign.67 This is more typical of poorly differentiated gastric carcinoma such as a diffuse type of signet-ringcell carcinoma or a small cell variant (Figs. 19-13 and 19-14).
Colorectal carcinoma is very common in developed countries, but bone metastases from these tumors occur less commonly than from gastric carcinomas.55 The incidence of bone metastasis from colorectal cancer is reported to be approximately 23% in autopsy cases.62 Cancers involving the rectum and cecum are more likely to develop bone metastasis than tumors involving other portions of the colon.69 Poorly differentiated carcinoma and signet-ring cell carcinomas have a higher propensity for metastasizing to bone than other histologic types of colon cancer (i.e., conventional, well to moderately differentiated adenocarcinoma of the colon). Skeletal metastases typically coexist with lung and liver metastases. Solitary skeletal metastasis can be a presenting sign in colorectal cancer. In fact, colon carcinoma is the most frequent gastrointestinal cancer in which solitary skeletal metastasis can appear as the initial clinical event. Some of these lesions may involve unusual sites such as the acral skeleton, skull, or patella or even the temporomandibular joint.59,65,72,74,78 Microscopically, these lesions have typical features of gastrointestinal adenocarcinoma with variable degrees of differentiation, mucin production, and signet-ring cell features.
Aspirates from metastatic colon carcinoma are usually very cellular and show cells organized in three-dimensional clusters, large sheets, small groups, and dispersed singly. Nuclei of cancer cells, arranged in palisade at borders of the large sheet composed of columnar cancer cells, is a characteristic feature of colon carcinoma (Fig. 19-15, A, B). Necrosis is very frequent in cytologic preparations from metastatic colon carcinoma. In many of the cases, cytologic features of metastasis allow recognition of the primary site even without clinical data.
