In 2001 in a publication in the Archives of Dermatology, an international peer-reviewed journal published by the American Medical Association, Dr. Vu Thuong Nguyen and colleagues first described the Paraneoplastic Autoimmune Multiorgan Syndrome (PAMS), its classification, clinical manifestation, and immunopathological mechanism.
This medical condition was previously reported and described under many different names, “Paraneoplastic Pemphigoid”, “Paraneoplasmic Bullosis”, “Paraneoplastic Skin Syndromes”, “Paraneoplastic Lichen Ruber Pemphigoides”, and Paraneoplastic Pemphigus (PNP), etc. Nine years after the publication, Dr. Vu Thuong Nguyen tells us how he first recognized this cancer associated medical condition as a syndrome, the history, and what he believes to be the significance of science progress in the future diagnosis and management of this syndrome.
Paraneoplastic Autoimmune Multiorgan Syndrome
The History and the Future
Vu Thuong Nguyen
La Belle Cosmeceuticals, Inc. Santa Clara, California USA
Dr Vu Cosmeceuticals, Santa Clara, California USA
Many scientific discoveries start with coincidences or paradoxes. Coincidences play a crucial role in the development of both intuitive and scientific theories. Coincidences are not simply unlikely events. To turn a mere coincidence to a meaningful discovery, the scientist needs to have the ability to recognize the importance of alternative theories in determining what constitutes a coincidence. This ability is strengthened by education, experience and the urge to search for alternatives to solve a contemporary problem.
This is the case for the identification of Paraneoplastic Autoimmune Multiorgan Syndrome (PAMS), a cancer associate pathologic syndrome that involves organs that are covered or layered by epithelial cells, and its pathophysiology. Being self-explained by its name, PAMS refers to the association of several clinically recognizable features, symptoms and signs, that occur in more than one organ, i.e., skin, oral mucosa, lung etc., that are caused by autoimmune disorders that occur in patients who are having tumor or cancer. The neoplasm could be obvious or occult. These phenomena or characteristics often occur together, so that the presence of one feature alerts the physician to the presence of the others including, most importantly, cancer.
Paraneoplastic Autoimmune Multiorgan Syndrome (PAMS)
Clinical Presentation and Classical Diagnosis
PAMS symptoms, reported by the patient, most commonly begin with wax and wane, recurrent painful oral mucosal erosion lesions (therefore the early cases of PAMS could be most commonly first recognized by dentists). Mucosal lesions sometime can also occur in other organs such as eye, pharynx, larynx, gastrointestinal mucosa, and anogenital areas. Lesions can also occur on skin. These lesions do not response to antibiotic treatment. Patients may also present with respiratory problems such as cough and shortness of breath, etc.
The diagnostic tests for PAMS include histopathology and immunopathology to visualize and differentially identify microscopic structures of the lesion and the specific pattern of autoantibody deposition on the epithelial tissue and to identify the specific circulatory autoantibodies in the body fluids of the patient.
Biopsies (samples) of the tissue adjacent to the lesions are obtained, cut to thin slices and placed on glass microscope slides. By the histochemistry technique, these thin slices are stained with chemicals so that cells and tissue structure can be differentially visualized under a light microscope.
In PAMS, there is marked inflammatory infiltration of mononuclear cells at the dermal-epidermal junction (DEJ). There is damage of the DEJ and vacuolar degeneration of the basal layer with suprabasilar clefting (rather than obvious acantholysis seen in classic pemphigus). Dyskeratotic (apoptotic) keratinocytes appear predominantly within the more basal regions of the epithelium. These microscopic damages can occur at different levels of severity leads to different clinical presentation; i.e., blisters develop either within the epidermis in a pemphigus-like pattern, and/or at the DEJ, in a pemphigoid-like or erythema multiforme–like pattern.
Using a technique called direct immunofluorescence, the thin slices are treated with fluorescence labeled antibodies specific to human IgG antibodies and complement factors so that their deposition on the tissue can be differentially visualized under a fluorescence microscope. Examination of perilesional tissue in PAMS reveals the deposition of these immunoreactants in the intercellular regions of the epithelium and at the DEJ.
Circulatory IgG autoantibody in PAMS that can bind to epithelium of skin and other organs is recognized by a technique called indirect immunofluorescence. In this technique, samples skin from healthy individuals, monkey esophagus, and rat bladder are cut to thin slices and placed on glass microscope slides and used as substrates. Serum from patient is obtained and applied on the substrates. Antibody from the serum that reacts to the epithelium in an intercellular pattern and along the DEJ is recognized and visualized with fluorescence labeled animal antibody specific to human IgG antibodies.
The specific targets of the circulatory IgG autoantibody in PAMS are identified by two techniques. In radio-immunoprecipitation assay, the PAMS antigens from the pool of radial-labeled epithelial proteins are precipitated by autoantibody in serum sample from patients affected with PAMS. These labeled antigens are separated by electrophoresis and visualized as separated bands represent the sizes of the antigens. The bands for PAMS antigens are 250, 230, 210, 190, 170, 150, 130, 105, 95, 82, 75, 53, 47, 42 and 40 kDa proteins. Among these bands, the 210 kDa band is the most prominent and frequent while other bands are less or some times undetectable.
Figure 1. PAMS antigens detected by radioimmunoprecipitation assay of serum isolated from PAMS patients and control healthy individual (Contl). Original data from Nguyen et al. Archives of Dermatology. 2001; 137:193-206. Better mathematical method is used to estimate more accurately the molecular weights of the antigens. PAMS antigens have molecular weights of 250, 230, 210, 190, 170, 150, 130, 105, 95, 82, 75, 53, 47, 42 and 40 kDa.
Some of these biomarkers for PAMS are identified as envoplakin (210 kDa), periplakin (190 kDa), desmoplakin (250 kDa), Bullous Pemphigoid Antigen (Dystonin, 230 kDa), desmoglein 4 (130 kDa), desmoglein 3 (130 kDa), and desmoglein 1 (150 kDa). The identities of other protein bands that are precipitated by PAMS autoantibodies are still to be identified. Possible candidates are some genes localize nearby to envoplakin gene in chromosome 17q25.
Other PAMS protein bands co-migrate with many proteins homologous to envoplakin. Their molecular weights include 75kDa, 82kDa, 95kDa, 105kDa, 150kDa, 170 kDa and 190kDa (Genbank accessions BAG63149, BAF83022, BAG59338, BAA25494, BAG65062, BAA86565, and CAP07522; protein sizes are predicted with 10-20 kDa increase by post translational modification). Other smaller protein bands co-migrate with proteins that involve in control of cell replication and growth such as p53 kinase, p47 cytohesin, and P42/p40.
Enzyme-linked immunosorbent assay (ELISA) is used to detect the presence of autoantibodies in PAMS to the above antigens. In this assay recombinant envoplakin, periplakin, desmoplakin, Dystonin, desmoglein 3, and desmoglein 4 are affixed to a surface of a 96-well ELISA plate, and then patient’s serum is washed over the surface so that autoantibodies can bind to the antigens. These antibodies are detected by anti human IgG antibody that is linked to an enzyme, and in the final step a substance is added that the enzyme can convert to some detectable signals that are detected by an ELISA reader.
In patients with history of cancer, the positive histopathological and immunopathological examination confirm the diagnosis of PAMS. In the patient with clinical symptoms of PAMS and without history of cancer, the positive result of the examination above can be the signs for occult neoplasm/cancer that early detection and treatment could be life saving.
Identifying PAMS Started With a Coincidence
The Urge to Search for Alternatives
I have though about finding a way to detect cancer at its early stage.
My father, also my idol passed away in 1994 because of cancer. He had gastric carcinoma - a malignant tumor of the stomach- that was diagnosed in 1992, just shortly after he arrived America to reunite with our family. After the first surgery that removed two third of his stomach, he did well for about a year. However, since his cancer was diagnosed too late and it had metastasized to other organs, it recurred and took my father’s life away.
I had the honor to be with my father the last year of his life observing his great fight against the cancer and learning his virtue. I felt so regret that such a great man like him had to leave so early. He was only 64 years old.
The days close to my father, I observed that he had diffused velvety thickening hyper-pigmentation of the skin in his axillae. It is called acanthosis nigrican. My father told me that he had noticed the discoloration several years before; as early as the time he was an ex-prisoner from a Communist “Re-education camp” in Vietnam – He was an officer in South Vietnam Army before 1975.
Acanthosis nigrican is a significant skin sign because if it is non-inherited and occurs in an appeared healthy individual, it is very commonly associated with an internal malignancy, usually cancer of the gastrointestinal tract.
I thought that my father could have been saved if he had been seen by some knowledgeable doctors who could early recognize this skin sign on skin. It combined with some medical symptoms that he had could be indication for a screening for the cancer that could be removed earlier. Many patients with neoplasm could be saved if their tumors are found and removed early. For many potential lethal cancers, I want to see if the advance of science would provide effective ways to detect the cancers at their early stages and remove or stop them before they could become lethal. The thought keeps pounding in my head since then.
Our study of Paraneoplastic Autoimmune Multiorgan Syndrome (PAMS) began in 1999. At the time, we were studying the pathogenesis of the potential lethal autoimmune skin disease called pemphigus vulgaris. In this disease patients develop antibobodies against many proteins of their own skin cells, most prominent are autoantibodies against desmogleins, and cause the cells to detach from one another, a specific process called acantholysis, which leads to skin blistering.
In the laboratory, we used sera that were isolated from blood samples of patients with pemphigus vulgaris to do experiments because they contained the disease causing antibodies. For the control of the accuracy of the experiments, we used sera from healthy individuals or patients with diseases other than pemphigus.
Our PAMS study started with a serum sample that we used for a control of our experiments in our research laboratory at the University of California at Davis. The serum was isolated from the blood of a patient who had been first diagnosed with lichenoid mucositis, an inflammatory disease of oral tissue that was completely different from pemphigus.
This control serum was used in an experiment called radioimmunoprecipitation assay, a technique that was used to visualize the proteins (antigens) targeted by the antibodies. In the experimental results, the proteins will appear as bands that are separated according to their molecular weights. Each band may represent one protein or many proteins that have the same molecular weight. Sometimes, the pattern of the protein bands of the immunoprecipitation assay can be very characteristic for an autoimmune disease.
When I did the radioimmunoprecipitation experiment with this disease control - lichenoid mucositis- serum and analyzed the pattern of the protein bands from the result, I found that it was indeed contained some bands that could be found in cancer patients who also developed autoimmune skin lesions called paraneoplastic pemphigus (PNP). The presence of a prominent 210kD band envoplakin, a protein component of the desmosome is the most significant one. The gene of this protein has been mapped to the tylosis oesophageal cancer locus on chromosome 17q25. This locus also contains a number of genes that have been found abnormal in other forms of cancer. I suspected that the patient might have a more serious disease than just lichenoid mucositis: NEOPLASM/CANCER.
It appeared that the patient was a 62-year-old man who first had lesions in his oral mucosa that was recognized by a dentist, who referred him to our dermatology clinic. He presented with a 1-year history of marked weight loss, recent respiratory distress, including dyspnea (shortness of breath), recurrent painful oral erosions, hoarseness, and dysphagia (difficult eating/swallowing). Dysphasia due to persistent oral erosion lesions was thought to be the reason to his weight loss. Our routine oral biopsy pathohistology results showed a typical lichenoid lymphocytic infiltrate at the dermal-epidermal junction and slight basal layer clefting that lead to his first diagnosis. He had no history of diagnosed neoplasm.
From the patient’s previous medical record, findings of oral biopsies were said to show focal pemphigus-like acantholysis on one occasion and a pemphigoid-like subepidermal bulla on another occasion.
The patient was suspected to have PNP and was referred to Internal Medicine clinic to look for possible occult neoplasm and the cause of dyspnea. Bronchial washings revealed clusters of bronchial epithelial cells. Computed tomographic (CT) scan of the abdominal region revealed a suprarenal mass, which was excised and histologically identified as a Castleman’s tumor (angiofollicular lymph node hyperplasia).
Further study of the patient’s oral biopsies in our laboratory for the deposition of IgG antibody showed a mixed staining pattern of pemphigus-like (antibodies against cell membrane associate proteins) and diffused basal/dermal-epidermal junction. Correlate to the histopathology study of the same biopsy, there was no pemphigus lesion (acantholysis). Instead, there was profound lymphocyte infiltration, slight epidermal-dermal junction splits, and apoptotic basal cells.
The patient's mucocutaneous lesions were treated with glucocorticosteroid over several months and he was doing well. He occasionally experienced respiratory symptoms, which he controlled using a corticosteroid inhaler.
The Appropriate Name of the Disease
To establish a precise diagnosis for this patient, since the patient had lichenoid mucositis at the time the neoplasm was discovered, his diagnosis should have been “paraneoplastic lichen planus” (“PNLP”). But the discovery of neoplasm in this patient happened by a coincidence. The patient also previously had separate occasions of having pemphigus-like acantholysis lesion and a pemphigoid-like subepidermal bulla. If the coincidence had occurred at either other two episodes, the diagnosis could have been “paraneoplastic pemphigoid” (“PNPd”) or “paraneoplastic pemphigus” (“PNPs”).
However, neither name above could be accurately applied to this patient since his neoplasm has probably been developing long before the development of symptoms of different autoimmune diseases in his body. The coexistence or swing between different clinical dermatologic presentations may be the result of the antigenic spreading and switching of autoimmunity in this paraneoplastic autoimmune syndrome.
In most patients, the autoimmunity feature of the disease is first recognized by lesions caused by autoimmunity of the epithelia of oral mucosa and/or skin. At the end of the disease course, the most common cause of death is autoimmune of the lung epithelial cells leading to small airways obstruction and respiratory failure, and sepsis. The lesions sometimes extend to skin. These symptoms may associate with other symptoms of other organs such as lung (i.e., cough and dyspnea). An early recognition of the existent of neoplasm in a patient would help for a much better out come (i.e., prognosis).
Thus the more precise, accurate and helpful name for this disease should be Paraneoplastic Autoimmune Multiple Epithelial Organ Syndrome or, to make it short, Paraneoplastic Autoimmune Multi-Organ Syndrome (PAMS). The term that reminds health care professionals be aware the possible existence of occult neoplasm when there is a presence of chronic, wax and wane, recurrent or persistent oral blister/erosion lesions that resistant to antibiotics.
The first time we described the pathologic syndrome PAMS was in our publication in the Achieves of Dermatology journal, Volume 137, Issue No. 2, February 2001: Nguyen et al: “Classification, Clinical Manifestations, and Immunopathological Mechanisms of the Epithelial Variant of Paraneoplastic Autoimmune Multiorgan Syndrome. A Reappraisal of Paraneoplastic Pemphigus”.
The survived patient that I mentioned here is the case number 3 in this publication. The patients in other two cases in the publication died. One had the same benign tumor (Castleman’s tumor). The other had suffered from a malignant cancer (non-Hodgkin lymphoma). Their cancer was diagnosed and treated before they presented at the hospital with the manifestation of autoimmune diseases in oral mucosa, skin, and lung that caused their death because of respiratory problem (i.e., fibrosis and shedding of lung epithelial cells that clog the small lung airways).
The determination for naming the syndrome: Paraneoplastic Autoimmune Multi-Organ Syndrome (PAMS) is to present a more accurate description and a better understanding the nature of the syndrome as well as to have the courage to search for a better way to help the patients who have this syndrome; specifically, an early detection of the original cause of the syndrome (i.e., the neoplasm) so we can treat it at its early stage, and to prevent its common cause of dead: chronic toxic autoimmune that cause fibrosis and obstruction of lung airways leading to respiratory failure.
The History of Evolutional Nomenclature and Recognition of
Paraneoplastic Autoimmune Multi-Organ Syndrome (PAMS)
Autoimmune skin blistering diseases such as pemphigus and pemphigoid that occur in patients with neoplasm (i.e., paraneoplastic) have been described since 1950’s, even before the time these skin blistering diseases were known to be autoimmune. There were cases of patients with pemphigus vulgaris, then latter was found to have carcinoma such as one that was described in 1954 by Muller in Germany in his article “Carcinoma development in pemphigus vulgaris” (Zeitschrift für Haut- und Geschlechtskrankheiten1954 Dec 15; 17(12): 373-5). There were other cases of patients with carcinoma, then were found to have pemphigus vulgaris such as those described by Ashmarin et al in Russia in their article “Association of pemphigus vulgaris & cancer of the stomach” (Klinicheskaia Meditsina 1959 Apr; 37(4): 142-4.)
Cases of cancer patients who had pemphigus foliaceus and bullous pemphigoid also have been described since 1960’s such as those described by Goessner and Korting in their article “Metastasizing Islet Cell Carcinoma of the A Cell Type in a Case of Pemphigus Foliaceus with Diabetes Renalis” (Deutsche Medizinische Wochenschrift 1960 Mar 1; 85: 434-7).
Forman in England published his article “Pemphigoid occurring with carcinoma of the rectum” (Proceedings of the Royal Society of Medicine 1960 Jul; 53: 563). Boyd in England published a case in his article “Pemphigoid And Carcinoma Of The Pancreas” (British Medical Journal 1964 Apr 25; 1(5390): 1092). When more cases were reported, the disease had a name. In 1964 Morandi, Panerai, and Bongi in Italy gave the disease a name in their article “Paraneoplastic Syndromes. 3. Clinical Contribution to the Knowledge ofParaneoplastic Pemphigoid” (Rivista Critica di Clinica Medica 1964 Oct; 64: 498-508). Husz et al gave a more general nomenclature for these blistering diseases coexisting in cancer patients “Paraneoplasmic Bullosis” (Dermatologica 1970; 141(6): 421-7).
Since then, there have been several reports of new cases and reviews that emphasized the significance of the clinical cases with the coexistent of neoplasm and autoimmune bullous disease. A review by Jablonska and Chorzelski in 1971 “Autoimmune Skin Manifestations in Malignant Neoplasms of Internal Organs” (Zeitschrift für Haut- und Geschlechtskrankheiten 1971 Oct; 46(19): 548-53) suggested neoplasms as the original cause.
Coexistent of different bullous dermatoses and/or lichen planus in cancer patients has also been described since 1960’s. A report by Szyszymar et al “On the Morphological Variability and Immunology of Various Paraneoplastic Skin Syndromes” (Dermatologische Wochenschrift 1968 Jul 27; 154(30): 706-12) described the coexistent of pemphigoid-like, pemphigus-like, lichen planus-like, and erythema multiforme-like lesions, characterized by both histology and immunofluorescence studies, in a group of patients with neoplasm.
In 1983 Pachinger describe a patient with lesions of both subepidermal vesicles within Lichen planus-papules and acantholytic suprabasal vesicles without pemphigus-autoantibodies. He named the disease “Paraneoplastic Lichen Ruber Pemphigoides” (Zeitschrift für Hautkrankheiten 1983 Jul 15; 58(14):1024-37)
These disease cases were classified as the subtype Paraneoplastic Autoimmune Syndromes of “paraneoplastic syndrome”, a familiar name that was used in 1960’s. It is now very rarely used since it is too general as the nature of cancer now known to be much more complicate and broad.
In 1990, Younus and Ahmed in Boston University, USA published a review of 60 cases of patients who had pemphigus coexisted with neoplasm (Journal of American Academy Dermatology 1990 Sep; 23(3 Pt 1): 498-502). Also in this year, Anhalt et al at Johns Hopkins University, USA, described 5 five patients with underlying neoplasms with oral mucosal and skin lesions, histologically showed vacuolization of epidermal basal cells, keratinocyte necrosis, and acantholysis. Immunofluorescence testing revealed atypical pemphigus-like autoantibodies in perilesional epithelium and serum. He named the disease “Paraneoplastic Pemphigus” (PNP) (New England Journal of Medicine 1990 Dec 20; 323(25): 1729-35).
The most important finding in the study by Anhalt et al in 1990 is the finding of a very characteristic result from the immunoprecipitation assay of to detect the proteins that are targeted by autoantibodies in the patients. The authors found an identical and unique complex of four polypeptides with molecular weights of 250, 230, 210, and 190 that was immunoprecipitated by all serum samples from 5 patients. Some patients also had antibodies against a 170 kD protein. None showed to precipitate a 130 kD protein. There was no precipitated protein with small weights was showed in their result because they used a “low percent” electrophoresis gel (6%) in their experiment. Technically, this low percent gel would mainly separate proteins of high molecular weights while the proteins of low molecular weights would run to the bottom and off the gel.
The authors speculated that the 250 and 230 kD proteins are desmoplakin and bullous pemphigoid antigen 1 (Dystonin) respectively (because these proteins migrated at the same positions of desmoplakin and dystonin when they were analyzed by electrophoresis assay). These proteins are members of the plakin protein family of adhesion junction desmosome and hemidesmosome respectively. The academic guess might stem from the mainstream hypothesis of autoimmune pemphigus at the time: pemphigus is caused by autoantibodies against adhering junction proteins (Stanley JR. Journal of Dermatological Science 1990 Jul; 1(4): 237-43). No other adhering junction proteins known at the time had a molecular weight of 170 kD.
Dr. Anhalt was the first scientist who developed the neonatal mouse model for pemphigus, an in-vivo experimental model that was used to prove that pathogenic antibodies in patients with pemphigus vulgaris alone could cause acantholysis. In his first publication of PNP in 1990, beyond other previous studies on paraneoplastic dermatoses, Anhalt et al used this model to test the pathogenic effect of antibodies isolated from these patients and demonstrated that the antibodies were pathogenic and could induce typical pemphigus-like lesion on the animals; specifically, cutaneous blisters, a positive Nikolsky's sign, and epidermal and esophageal acantholysis. Microscopically these lesions were similar to experimental pemphigus vulgaris.
In 1998, Amagai and Anhalt et al showed in their study of 25 PNP patients that all 25 had antibody against desmoglein 3, a 130 kD protein. They showed that removal of this anti-desmoglein 3 autoantibody could remove the ability to cause disease in the animal model (Journal of Clinical Investigation 1998 Aug 15; 102(4): 775-82)
Thus according to studies by Anhalt et al, paraneoplastic pemphigus (PNP) specifically refers to the disease in which patients have autoimmune mucocutaneous disease that produce autoantibodies against desmoglein 3 that cause lesions characterestic of pemphigus vulgaris; i.e., epidermal acantholysis that is similar to pemphigus vulgaris, and the patients also have neoplasm. In other words, by definition, PNP is a subgroup of pemphigus vulgaris in which the patients also have neoplasm or cancer.
Among biomarkers for PNP, autoantibody to desmoglein 3 (130 kD) is the most important one because it presents in ALL patients with PNP and it is the key factor that causes pemphigus vulgaris like lesions according to the authors’ studies.
In the publication in New England Journal of Medicine 1990 Anhalt et al have established a criteria for diagnosis of PNP. The criteria, however, has been modified by the author in 2004. It is probably because there are an increase number of reports of atypical cases of PNP. In many of these atypical PNP cases the patient presented with signs other than pemphigus vulgaris (i.e., no typical intraepidermal acantholysis) and/or do not have autoantibodies against desmoglein 3.
If one could make more academic guess, the 210kDa and 190kDa antigens found by Anhalt et al in 1990 could be guessed as the two cornified envelope precursors that were first identified by Simon and Green in 1984 (Cell. 1984; 36:827–834). Indeed, years after, the 210kD protein was identified as envoplakin and the 190kD protein was identified as periplakin (Ruhrberg et al, Journal of Cell Biology. 1996; 134(3): 715-29 and 1997; 139(7): 1835-1849). The two proteins turn out to be major antigens of PAMS.
In 1997, Kim et al in Korea used serum from PNP patient to screen expression cDNA library. They found that the antibody in the PNP serum strongly recognized a clone that turned out to be envoplakin (Journal of Investigative Dermatology 1997 Sep; 109(3): 365-9). Their results directly suggested that anti envoplakin antibody is the most prominent one among autoantibodies in the PNP serum that they used.
In 1998, Mahoney et al used 5 recombinant plaque proteins to evaluate the existent of PNP autoantibodies against plaque proteins (J Investigative Dermatology. 1998 Aug; 111(2):308-13). They also found antibody in all PNP sera revealed strongest reaction with envoplakin. They found that the antibody in all PNP sera also recognized periplakin (190 kD). To a lesser extent, antibody in certain PNP sera also recognized desmoplakin and plectin, and weakly, Bullous Pemphigoid Antigen (dystonin). The level of autoantibodies seems to decrease in molecules that have less homology to envoplakin.
Unlike studies by Anhalt et al in 1990, in our study of PAMS in 2000, we used a broader and high percent gel for immunoprecipitation assay that allowed us to identify the antigens of both large and smaller molecular weights. The result was detected by better technology with higher sensitivity. We found additional novel PAMS antigens that appeared to migrate at the same locations of 7 novel proteins that are highly homologous to envoplakin. Thus provided a more complete panel of antigens recognized by PAMS autoantibodies.
The serum of our patients at study had no autoantibodies against desmoglein 1 or desmoglein 3. Our experiment of passive transfer of PAMS antibodies to non-desmoglein 3 neonatal mice model demonstrated that indeed the binding of antibodies to mice epithelial cells could recruit mononucleated lymphocytes to the site of action that caused problems that resemble those in affected patients.
At the experimental mice skin, there is microvesiculation and lichenoid subepidermal mononuclear infiltrate subjacent to the areas of vacuolization of basal keratinocytes and disintegration of the basal cell layer caused by separation of basal keratinocytes from both the basal membrane and the adjacent keratinocytes that produces intraepidermal clefting that is different from classic pemphigus vulgaris lesions.
A loss of epithelial cell adhesion similar to that found in the skin occurs in large airways where the respiratory epithelium cells detach from the lamina propria and neighboring cells and form clusters occluding the airways. Thus demonstrates a cause of bronchiolitis obliterans, the major cause of dead in these cancer associate patients.
Binding of antibodies to epithelial cells that recruit trouble-making mononuclear cells to the site of actions provides a different concept of pathomechanism of the disease. Rather than simply a direct interference of adhesion molecules by antibodies, there is an induction of cytokine secretion that is caused by binding of antibodies to epithelial cells that recruit trouble-making lymphocytes to the sites of action and induce the lesions.
Thus we proposed the name Paraneoplastic Autoimmune Multiorgan Syndrome (PAMS) as a more accurate name for the disease because it describes a syndrome caused by a combination of both humoral and cellular autoimmunity attacking stratified and simple epithelial cell of many organs in patients who have neoplasm/cancer. The mucocutaneous lesions can be lichenoid, pemphigoid-like, pemphigus-like, or a mixture of all. The cell mediated cytotoxic has the primary role in the disease and dominant at early stage. However, the balance could shift to both humoral and cellular at the late stage. Furthermore, completely remove the tumor from the patient could help to improve the symptoms suggests that the tumor is the source inducing toxic autoimmunity (i.e., paraneoplastic).
If one truly respects history, then when diagnose a case with humoral dominant and have sub-basal separation or suprabasal acantholysis, he should call it paraneoplastic pemphigoid (Morandi et al, 1964) or paraneoplastic pemphigus (Anhalt et al, 1990) respectively. But in cases with cellular dominant and that the presence of autoantibody was undetectable, and the histopathology of the lesions is lichenoid with some sub-basal cleft such as those reported by Cummins et al. (Journal of American Academy of Dermatology 2007, 56(1): 153-159), because they do not have characteristics of pemphigus therefore they are not a subgroup of pemphigus, these cases should be better called Paraneoplastic Lichen Ruber Pemphigoides (Pachinger, 1983).
However, naming the disease is more complicate because in reality many patients affected by this paraneoplastic syndrome can present with a mixture of lichen planus and different levels of different bullous dermatoses. In some patients the disease stormy progression could occur rapidly in a few weeks from papulosquamous and lichenoid eruption without autoantibody detected to full blow severe stomatitis and cutaneous erosions with the presence of autoantibodies. In addition, there is the symptomatic involvement of other affected organs.
Thus any atypical case of “Paraneoplastic Pemphigus” in which there is no typical pemphigus vulgaris-like skin lesions or there is a combination of pemphigus-like, pemphigoid-like, or lichenoid lesions and there is infiltration of mononucleated cells such as CD8+ CTL, CD56+ NK cells, and CD68+ monocytes/ macrophages at the lesions in patients with neoplasm, should be appropriately diagnosed as Paraneoplastic Autoimmune Multiorgan Syndrome (PAMS).
Possible Pathogenic Mechanism of Paraneoplastic Autoimmune Multiorgan Syndrome
At epigenetic level, tumor cells wrongly express a group of genes: the source of activators for autoimmune memory T cells
Normal cells in the body follow an orderly path of growth, division, and death. Programmed cell death is called apoptosis. Cell growth and dead is controlled by several genes that locate at different chromosomes. Oncogenes tell cells when to divide and tumor suppressor genes tell cells when not to divide. When there is a damage of DNA that harms normal activity of cell, DNA-repair genes instruct a cell to repair damaged DNA. When something goes so wrong that cell damage cannot be repaired, suicide genes control apoptosis and tell the cell to kill itself.
When these genes are abnormally expressed or suppressed that lead to cells uncontrollably grow and do not die, neoplasm/cancer is the ultimate result.
The genes that control cell growth and dead along with more than 23,000 other genes are encoded in about 1.5% of the DNA of our genome. Our genome wraps around cellular capstans called histones, which are tagged by enzymes such as Histone Acetyl/Methyl Transferase and Histone Deacetylase/Demethylase. Intrinsically, these epigenetic enzymes use molecules such as acetyl and methyl to controls the expression of genes at epigenome level by tagging, wrapping and activating of stretches of DNA that contain these genes.
The arrangement of genes in the genome follows a logic resulted from evolution. The genes that need to activate for cell growth commonly locate in the vicinity of genes that code for structural proteins of the cell. Thus unwrap a stretch of DNA by epigenome control allows activation of many genes in that DNA stretch.
This might possibly be the case of Paraneoplastic Autoimmune Multiorgan Syndrome. There might be an abnormal epigenome activation of a group of genes in chromosome 17q25 that are necessary for initiation of tumorgenesis; i.e., B lymphocyte proliferation in benign Castleman’s tumors and lymphoproliferative malignancies. This activated DNA stretch contains genes for envoplakin and smaller proteins that become targets for low level of autoimmunity, an extrinsic natural surveillance mechanism for gene over expression.
From local low-level autoimmunity, an extrinsic natural surveillance mechanism for gene over expression, to a generalized high-level toxic autoimmunity.
In the case of tumorgenesis, an abnormal over-expression of oncogenes could coexist with over expression of other bystander genes that may signal for initiation of a local low level of natural autoimmunity. The epigenetic control of immunoediting switches on immune genes called MHC in chromosome 6 which helps the immune system to specifically identify and eliminate unwanted excess proteins that otherwise could turn a cell to tumor cells. This is the first line of low level of local natural autoimmunity.
The second line of low level of natural autoimmunity is local activation of natural killer cells to destroy the new tumor cells. It is possible that in Paraneoplastic Autoimmune Multiorgan Syndrome, one of the immunoediting products is the natural autoimmunity that recognizes the imbalance excess of envoplakin in relate to other linked proteins within the adherent structure of these tumor cells (thus explains why the linker domain of envoplakin is the most common epitope for autoimmunity in PAMS).
The important pathogenesis features Paraneoplastic Autoimmune Multiorgan Syndrome is the presence of neoplasm leading to the development of local low level of natural autoimmunity. It is a natural surveillance mechanism that might aid in cleaning up excess proteins and the recognition of neoplastic cells (i.e., by CD8+ T cells); thus reduce the incidence of cancer. However, when the cancer development become more dominant, this low level of natural autoimmunity turns into high-level cytotoxic autoimmunity, cell-mediated with addition of antibody-mediated, against epithelial cells and the basal membrane.
Autoimmune antigenic spreading from plakins to desmogleins
The formation of autoimmunity to additional antigens in PAMS may due to epitope spreading (reviewed by Chan LS. Archives Dermatology. 2000 May; 136(5): 663-4). It is a way that autoimmunity being activated as a consequence of the release of other antigens secondary to the destruction of the primary antigen, that may lead to organ-specific or multiorgan autoimmune disease. The first line of epitope spreading uses the molecular mimicry mechanism that causes autoreactive T cells to be activated de novo by progressively less dominant self epitopes with a homologous immunodominant sequence released secondary to T cell-mediated destruction. Thus in PAMS the next antigens after evoplakin would be the molecules that share the most similarity to the protein.
Sequence analysis of envoplakin using the Basic Local Alignment Search Tool (BLAST, http://blast.ncbi.nlm.nih.gov) showed that the protein is homologous to several member of the plakin family of proteins such as desmoplakin, bullous pemphigoid antigen 1 (dystonin), periplakin, plectin, epiplakin, and other plakin homology proteins including those with molecular weights of about 75kDa, 82kDa, 95kDa, 105kDa, 150kDa, 170 kDa and 190kDa (Genbank accessions BAG63149, BAF83022, BAG59338, BAA25494, BAG65062, BAA86565, and CAP07522, protein sizes are predicted with 10-20 kDa increase by post translational modification). It appears that these proteins have similar sizes with the novel PAMS antigens that we reported in our 2001 publication.
Since proteins of the plakin family express not only in stratified epithelium but also in transitional and pseudostratified epithelium such as those in the lining of urinary bladder and respiratory tract, in striated muscles, and in other organs, the first line of epitope spreading in PAMS explains the multiorgan autoimmunity nature of the syndrome.
Conversely, the second line of epitope spreading could be due to target antigens being physically linked intracellularly as members of a complex to the primary antigen. The result of this is an autoimmune response to antigens that are not homologus to the immuno-dominant antigen. This epitope spreading explains the generation of autoimmunity to other desmosomal associate antigens such as desmogleins 4, desmogleins 3, and desmogleins 1 when PAMS toxic autoimmune manifests in mucocutaneous epithelium.
The concept of antigenic spreading from plakins to desmogleins in PAMS is supported by the fact that autoantibody to envoplakin and other plakins is virtually the markers for PAMS at any stages while many PAMS affected patients do not have anti desmoglein autoantibodies and classic pemphigus-like lesion. The presence of anti desmoglein autoantibodies seems to be more common in affected patients at the late course of PAMS, which is probably the result of chronic, wax and wane manifestation of toxic autoimmune in mucocutaneous tissue.
From an early localized/simple oral lichenoid lesions to a late generalized/severe stomatis/mucositis and skin eruption with necrotic eruptions, basement membrane splitting and acantholysis, respiratory failure, sepsis and multiorgan failure.
Thus the earliest epithelial manifestation of toxic autoimmune in PAMS is the infiltration of cytotoxic mononuclear cells at the dermal-epidermal junction that induce Lichenoid lesions alone or with pemphigoid-like erosion of mucocutaneous tissue. The lesions most commonly localize orally and worsen because of the friction of mastication. Mild pulmonary manifestations include cough and mild shortness of breath may occur at early course of PAMS. Formation of additional autoantibodies to keratinocyte-exclusive antigens such as desmoglein 4, desmoglein 3, and desmoglein 1 due to antigenic spreading leads to additional features including pemphigus-like lesions.
Full attack of toxic autoimmune in PAMS composes of cell-mediated cytotoxic reaction at mucocutaneous tissue that induces apoptosis of keratinocytes, hence pushing the dynamic status and activity of the adherent structures of the cells. This process produces a full opportunity for the attack of autoantibodies to several desmogleins and plakins. Both cell-mediated cytotoxic reaction and humoral attack together disrupt the adhesion function of the keratinocytes of stratified epithelium at all levels, sub-basal, supra-basal, intra-spinosal, and intra-granular, that in mucosa, produces a generalized stomatitis/mucositis, and in the skin, confluent erosive lesions resembling toxic epidermal necrolysis (TEN).
At the most severe level of toxic autoimmune, the trouble of immune system could spread to more serious level that it might not even recognize well the true strangers (microbial infection) that are vulnerable to the patients when their skin and oral mucosa barrier is compromised. This adds up with the complications associated with immunosuppressive therapy applied to the patients at the time the skin and mucosal lesions flare up, could lead to generalized infection, sepsis and multiorgan failure, and death.
In late course of Paraneoplastic Autoimmune Multiorgan Syndrome, manifestation of cytotoxic autoimmunity on respiratory epithelium leads to inflammatory obstruction of airways, which is severe enough to result in death. The pathomechanism of constrictive bronchiolitis obliterans found in many patients affected with PAMS involves sloughing of the overlying epithelium and fibrosis predominantly in respiratory bronchioles.
The latter could be the result of chronic immune inflammation and chronic apoptotic induction, evident by the up-regulation of Fas/Fas-ligand expression in alveolar epithelium and in infiltrating cells. Airways obstruction and fibrosis lead to impaired gas exchange due to ventilation-perfusion inequality that causes hypoxemia and respiratory failure, a major cause of death in many patients affected with PAMS.
Thus the longer time the delay of diagnosis of PAMS, the larger the reservoir of toxic autoimmune memory T cell being established and the more chance for respiratory airway fibrosis to develop. Only a small amount of activators released from the tumor/cancer can activate a full blow of toxic autoimmunity that create a stormy progression of the disease leading to devastated outcome: death.
This explains why in PAMS affected patients who have associated benign or localized tumor such as encapsulated Castleman’s tumor or benign thymoma, complete surgical removal of the tumor will help disease improve substantially or enter a complete remission. However, in affected patients who have associated malignant lymphoma, simply debulking the tumor or reducing tumor burden through chemotherapy or radiation will not halt disease progression. It is because the course of PAMS depends more on the existence than the size of the neoplasm.
Searching For an Early Diagnosis and Treatment for PAMS
Currently, the prognosis of patients who are diagnosed with paraneoplastic Autoimmune Multiorgan Syndrome (PAMS)/paraneoplastic pemphigus (PNP) is very poor. The mortality rate is more than 90 percent. It is probably that to fulfill enough criteria to make the diagnosis of PNP (i.e., to fulfill the pemphigus-like characteristic), the affected patients are already at the late stage of PAMS.
The presence of the typical generalized painful, progressive stomatitis, with preferential involvement of the tongue, and a full blow of autoantibodies against desmoglein 3, two of the diagnostic criteria for PNP could be the signs for a late manifestation of toxic autoimmunity in PAMS. Therefore by the time the diagnosis is made, the disease occurs rapidly, has a short and stormy progression, and leaves doctors with a limited window of opportunity to save the patients. Thus late diagnosis could be responsible for the poor prognosis of the syndrome.
Current treatment of this syndrome when it is diagnosed as PNP is discouraging. Several approaches that are used alone or in combination such as to remove toxic autoantibodies by plasmapheresis, suppress autoantibody production by high-dose intravenous immunoglobulins, inhibit proliferation of B and T cell by immunosuppressant drugs such as glucocorticosteroids, mycophenolate mofetil, cyclophosphamide and cyclosporine, etc., show unsuccessful. Inhibiting the activity and induce apoptosis of CD20+ cells (B lymphocytes) by anti CD20 monoclonal antibody (Rituximab), that could also reduce the burden of cancerous B cells doesn’t seem to give better promise.
Better hope for patients affected with PAMS could be achieved with earlier detection of the syndrome.
New criteria that might help to detect and make the diagnosis of PAMS at its early stage:
(1) Painful stomatitis with lesions that may be lichenoid or erosion. The lesions occur wax and wane and that resistant to antibiotic treatment. Additional helpful-but-not-critical features include polymorphous cutaneous eruption with lesions that may be lichenoid, resemble erythema multiforme, blistering, or resemble toxic epidermal necrolysis.
(2) Histologic findings of lesional/perilesional biopsies reflect a typical lichenoid lymphocytic infiltrate at the dermal-epidermal junction and slight basal layer clefting or interface change. Additional helpful-but-not-critical features include pemphigoid-like or/and pemphigus-like (acantholysis).
(3) Serology using Enzyme-linked immunosorbent assay (ELISA) detects the presence of autoantibodies to envoplakin and at least two other proteins of the plakin family. Additional helpful-but-not-critical features include the presence of autoantibodies against desmogleins.
(4) Demonstrate the presence of underlying neoplasm. Patients who have no history of a diagnosed neoplasm, whose symptoms and signs satisfy criteria 1, 2 and 3, are subjected for work up to detect the occult tumor.
Modern technology such as the microarray ELISA-style assays will accelerate immunodiagnostics of PAMS significantly. PAMS antigen panel is used as substrate for the assay. It is a protein-chip in which recombinant proteins of plakins and desmogleins are affixed to a surface of the chip to screen for the presence of autoantibodies. The signal is amplified and electronically analyzed. The whole process of microarray ELISA-style assays is performed by automated robotic processing system that will make laboratory diagnosis for PAMS become simple and practical and could be sufficient as a single immunopathological laboratory test for the syndrome.
While immunofluorescence assay can be less sensitive and have more false positive results, microarray ELISA-style assay is very sensitive and specific since it can detect a very small amount of the specific autoantibodies even when they are small enough to give a negative result on direct and indirect immunofluorescence assays.
A positive serology result for autoantibodies to envoplakin and other members of plakin family is very important to make the diagnosis of PAMS. It dismisses the necessity to perform the direct immunofluorescence assay on patient’s biopsy sections and radioimmunoprecipitation assay. Neither indirect immunofluorescence assay on monkey esophagus or rodent urinary bladder sections is necessary because these members of the plakin family proteins express in epithelium of many organs, thus declares the multiorgan nature of PAMS.
Recommended recombinant proteins for the PAMS chip for microarray ELISA-style assay:
(1) Envoplakin (GenBank: AAC64662.1), (2) periplakin (GenBank: AAC17738.1), (3) Plectin (GenBank: AAB05427.1), (4) Desmoplakin (GenBank: AAA85135.1), (5) Bullous pemphigoid antigen (GenBank: AAA35606.1), (6) novel protein (GenBank: BAG63149), (7) novel protein (GenBank: BAF83022), (8) novel protein (GenBank: BAG59338), (9) novel protein (GenBank: BAA25494), (10) novel protein (GenBank: BAG65062), (11) novel protein (GenBank: BAA86565), (12) novel protein (GenBank: CAP07522), (13) Desmoglein 1 (GenBank: X56654.1), (14) Desmoglein 3 (GenBank: M76482.1), and (15) Desmoglein 4 (GenBank: AY168788).
Complete removal of tumor is the optimal treatment of PAMS at early stage. This could be done for PAMS affected patients who have encapsulated benign tumor such as thymoma and Castleman’s tumor. Detection of tumor could be done by computerized tomography scan when PAMS is suspected.
It is much more challenging for PAMS affected patients who have malignant tumor because simply debulking the tumor or reducing tumor burden through radiation and chemotherapy will not guarantee to remove completely the tumor. Thus it will not halt disease progression when the toxic autoimmunity in PAMS has already fully developed. However, early detection of PAMS still might help early control of the disease, which might give a better outcome.
New therapy development should aim at the epigenetic level. For example, changes in the behavior of two epigenetic enzymes, Histone Acetyl Transferase and Histone Deacetylase seem to play a role in toxic autoimmunity and cancers by switching on the wrong set of genes. Therefore readjusting their activity might help to control the progression of the disease.
The knowledge of PAMS has progressed substantially since it was recognized more than a half century ago under different names due to different clinical presentations observed by the doctors who first described it. The convergence of different names of the disease to a common name, Paraneoplastic Autoimmune Multiorgan Syndrome (PAMS), reflects more accurately the nature of the clinical presentation as well as the mechanism of the pathogenesis of the disease.
We hope that with increased recognition of PAMS, more timely and accurate diagnosis will be made, more insight to the understanding of pathogenesis mechanisms of toxic autoimmune injury, and better therapy will be developed, allowing doctors to better treat these affected patients.