Triaging Times

As a clinical instructor and lead cytologist at my institution, I like to remind our newer cytologists and cytology students of the importance of being prepared for FNA biopsies so they develop good habits or best practices as they become more experienced. This level of preparation helps to create a culture of ongoing learning and improvement, which is necessary for the laboratory. In my experience, I’ve met some cytologists who prefer to go into a case blind, with the mindset that knowing the patient’s clinical history in advance muddies their knowledge, skills, and abilities, limiting their mindset by excluding the possibility of other diagnoses. While diving into the unknown might seem exciting, it is also a hindrance and could result in errors, especially when the clinical history helps us triage the patient’s sample. For example, knowing that the patient has a history of lymphoma or that the presentation state includes bulky lymphadenopathy prompts us to collect additional needle passes to send for flow cytometry analysis. Another concern is not knowing whether the patient has a history of breast, gastric, or esophageal cancer, and consequently processing the specimen routinely, which may result in an extended cold ischemic time. This delay in fixation along with insufficient formalin fixation can yield false negatives on ER/PR IHC in breast cancers and HER2 FISH in breast, gastric, and esophageal cancers, which could restrict the use of hormone therapies, such as tamoxifen and aromatase inhibitors for hormone receptor-positive (HR+) cancers, or trastuzumab for HER2+ cancers. I cannot overemphasize the importance of familiarizing yourself with clinical history and communicating case specifics while you act as a mediator between clinician and pathologist.

Whether the clinical history impacts the pre-analytical phase, such as specimen collection (limiting cold ischemic time or collecting additional needle passes for ancillary studies) or the analytical phase, as such processing (formalin fixation) and diagnosis (selecting an appropriate immunoprofile), we must remain vigilant and proactive in laboratory medicine. In this case, knowing the patient’s clinical history was of the utmost significance as it helped to reduce the number of immunostains and ancillary studies necessary to make the diagnosis. Using morphologic criteria in tandem with the patient’s clinical history narrowed the differential diagnoses to just two possible types of cancer, presented below.

A 59 year old male patient presented to the emergency room after an automobile accident. On imaging, the X-ray and CT scan identified a left humerus mass and fracture, and bloodwork was performed. His medical record was sparse and uneventful with no recent visits or encounters. To build a more comprehensive wellness profile and prepare for surgery, he was also offered a one-time screening for Hepatitis C, as an adult who was born between 1945 and 1965.

The left humerus mass was biopsied via CT-scan guidance and two passes were obtained. The Diff-Quik stained smears demonstrate large polygonal cells, some with abundant, granular cytoplasm and some isolated cells with naked nuclei. Vessels also appear to traverse some of the cell groups.

Images 1-2: Bone, Humerus, Left, CT-guided FNA. Diff-Quik-stained smears.

The Pap-stained smears also demonstrate polygonal cells with granular cytoplasm, nuclei with coarse chromatin, and prominent nucleoli. An interesting feature frequently identified in this case is the intranuclear inclusions, and in hindsight, a focus on these may have further reduced the number of immunostains performed.

Images 3-5: Bone, Humerus, Left, CT-guided FNA. Pap-stained smears.

The H&E-stained cell block sections show trabeculae with endothelial wrapping around the cell cords. While renal cell carcinoma was listed as a differential diagnosis due to its telltale oncocytic cytoplasm and vascularity, hepatocellular carcinoma was favored.

Images 6-7: Bone, Humerus, Left, CT-guided FNA. H&E sections (6: 100x, 7: 400x).

Immunostains were performed using proper positive and negative controls on the cell block sections, and the tumor cells show positive staining for Arginase, cam5.2, and Hepar1, while negative staining for CK7 and PAX8 (not shown).

Images 8-10: Bone, Humerus, Left, CT-guided FNA. Cell block section immunohistochemistry. 8: Arginase-positive; 9: cam5.2-positive; 10: Hepar1-positive.

Fortunately, before ordering immunostains, both our cytologist and pathologist working on the case peered into the patient’s medical record and noticed that he had recent bloodwork which demonstrated a positive Hepatitis C screening. This diagnosis was as recent as the identification of his humerus mass. Had it not been for his car accident, I can’t imagine how long he would have gone undiagnosed for both hepatitis and metastatic hepatocellular carcinoma. Incidental findings save lives, folks.

Granted, in settings of unknown primaries with widespread metastatic disease, such as carcinomatosis, an extensive workup is almost always inevitable. Narrowing down possible etiology based on information such as gender, age, and environmental or occupational exposure can help, but that doesn’t always yield a definitive answer as time- or cost-effectively as possible. In this case, that one clue of untreated Hepatitis C was all the cytopathology team needed. A rarity, sure, but as we are asked to do more personalized tests with less material, think of the patient’s specimen as a puzzle and keep your eye out for a clue both under the microscope and behind the computer. You never know what you might find that reduces errors and unnecessary testing while efficiently leading to a definitive diagnosis.

-Taryn Waraksa-Deutsch, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

BOGO: Biopsy One, Get One Free

I’ve mentioned before how important it is to know clinical history before attending a biopsy, and I cannot stress this point enough. As the first line of screening, the intermediary between clinician and pathologist, the role of the cytologist is to prepare, assess, and convey. In a cancer center, we have three main populations: the patients with the unknown primary, the patients with the suspected primary, and the patients with the suspected metastasis. In the event of a suspected metastasis, we’ll review previous relevant pathology material if we have it onsite. Unless the clinician is requesting additional prognostic markers, the review process helps us eliminate the unnecessary repetition of immunostains (IHC) by confirming that the current material is morphologically consistent with the prior material. Sometimes we still perform old-school cytology without a plethora of ancillary studies. HA!

Most of the endobronchial ultrasound (EBUS) procedures performed at our institution are for lung cancer staging or differentiation between a lung cancer metastasis and an extra-pulmonary metastasis. Not that we don’t see the occasional sarcoid- or anthracosis-related process from time to time, but our most common indication is cancer. For an 88-year-old male patient with multiple lung nodules and both mediastinal and hilar lymphadenopathy, confirmation of metastasis was the main objective of the EBUS procedure. The patient’s pertinent medical history includes former tobacco use, squamous cell carcinoma of the lung (diagnosed percutaneously in 2022), clear cell renal cell carcinoma (s/p partial nephrectomy in 2020), prostate cancer (radiated in 2007), melanoma (excised in 2001), and cutaneous squamous cell and basal cell carcinoma (also previously excised in 2002 and 2008). With an extensive cancer history, the lung nodules and thoracic nodes could be any of them, although metastatic squamous cell carcinoma of the lung was clinically favored. My awesome cytologist colleague, Kelly, attended the EBUS procedure. The Rapid Onsite Evaluation (ROSE) was a clear-cut “adequate for diagnostic material,” and the attending pathologist added “tumor cells present.” The following morning, Kelly stopped by my desk to ask my opinion of the 12R (right hilar) lymph node she was screening. She said, “look at my dots. Do these look like the same cells to you? Or are they different? Because I feel like they’re different.” Before putting the slide on my scope, I asked, “so… like a combined adenosquamous? Or a small cell component?” She replied, “not small cell. Something… I don’t know, but they look different. The patient was recently diagnosed with lung cancer and has a history of renal cell.” I fixated on the H&E cell block slides (Images 1-3) before perusing the Diff-Quik and Papanicolaou-stained slides (Images 4-5). “Uhm… Why are there two different types of tumor cells here?! The cytoplasm here is so… vacuolated, but it’s not quite like lung adeno, and the other group… even the n/c (nuclear-to-cytoplasmic) ratio is different. What is this?” Kelly replied, “okay, so there are definitely two different types of tumor here.” I looked up, “It has to be. Absolutely, yes.”

Images 1-4. Lymph node, 12R, EBUS-guided FNA. 1-3: H&E cell block sections 1, 100x; 2, 400x; 3, 100x. 4: Diff-Quik stained smear.
Image 5. Lymph Node, 12R, EBUS-guided FNA. Pap-stained smear.

Kelly entered her diagnosis into our laboratory information system and brought the case over to the pathologist on cytology service for the day. She explained her thought process, and the pathologist also questioned if it was a combined process, such as a lung adenosquamous and maybe the original lung biopsy only sampled the squamous component. With the most recent clinical history of both lung squamous cell carcinoma and clear cell renal cell carcinoma, an IHC panel was appropriately selected. Later that afternoon, the pathologist exclaimed, “IT’S BOTH! IT’S SQUAMOUS AND RCC!” The clusters of squamous cell carcinoma did not stain for PAX8 (a renal cell carcinoma marker) (Image 6), and the same cluster stained positive for p40 (a squamous cell carcinoma marker) (Image 7). Within the same level of the cell block, the cluster of cells that appeared morphologically different than squamous cluster stained positive for PAX8 (Image 8) and negative for p40 (Image 9), confirming a renal cell carcinoma component. A small focus of p40-positive cells was present next to the p40-negative renal cell carcinoma (Image 9), further demonstrating mixed histology. This finding was shared with other pathologists, and the results were immediately called to the pulmonologist as this was a critical finding. Sometimes we encounter a partially involved node where the tumor cells are intermixed with lymphocytes, sometimes the lymph node yields more tumor than the primary site, and sometimes, albeit rarely, we encounter a lymph node infiltrated by two different carcinomas.

Images 6-9. Lymph Node, 12R, EBUS-guided FNA. Cell block section immunocytochemistry. Squamous cell carcinoma cluster – 6: PAX8-negative; 7: p40-positive. Renal cell carcinoma cluster – 8: PAX8-positive, 9: p40-negative (with small focus of p40-positive squamous cell carcinoma).

Due to the patient’s bulky disease and PD-L1 expression of 30%, the medical oncologists primary aim was to treat the squamous cell carcinoma first and follow up renal cell carcinoma therapy second. After the first few cycles of treatment, the lung nodules have decreased in size, but the thoracic nodes remain unchanged. Once the squamous cell carcinoma is controlled or demonstrates a more significant response, immunotherapy may be added to target both, with a tyrosine kinase inhibitor directed at renal cell carcinoma metastases in the event of progression.

-Taryn Waraksa-Deutsch, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

While We Are Young

Preface: I wish we lived in a world where cancer did not exist. While I do not foresee that being a reality in my lifetime, I hope I can witness a milestone in oncology when the development of safe and effective therapies surpasses the rate of metastasis. I hope to see the day when instead of being five steps behind cancer, we gain our footing and come out at least one step ahead. We can’t be that far away, right? When I see a patient who is younger than me or around my age, I can’t help but ask myself, “why they have to go through this? Why do they have to be diagnosed with cancer?” Now, they have to choose whether or not to undergo fertility sparing treatment in addition to making a shared decision regarding cytotoxic therapy and radiation. They’re just beginning their life, their career, their family; they’re finally building their story, and it’s interrupted by cancer. Rather than a partner or friend, I see many younger patients present to our clinic with a parent. Imagine how that feels to be a parent – knowing your child was just diagnosed with a terminal disease. How does one cope with that? No parent should have to endure the pain and emotional suffering of losing their child to cancer, let alone anything. We’re meant to outlive our parents, we’re meant to enjoy a full and healthy life, and this case tore right through me.

The patient, a 28 year old male, first presented to an out-of-state health system after managing intermittent right hip discomfort with NSAIDs and ice for three years. When the pain worsened, he went to an urgent care toward the end of 2018, but his X-rays were negative. After his pain became unbearable six months later, he had an MRI which demonstrated a proximal femur lesion with hip and sacral involvement. The femoral lesion biopsied via FNA, and the patient underwent 6 cycles of induction chemotherapy prior to resection. The patient’s PET scan showed improvement, and the 10.5 cm hip mass was resected demonstrating fibrosis and no residual disease. Additional chemotherapy was prescribed for consolidation followed by radiation to the sacral lesions. Seven months after no evidence of disease, a surveillance PET scan demonstrated abundant bone lesions in the arms, ribs, and pelvis. A new relapse chemotherapy regimen was prescribed for refractory disease, but within cycle 2, new calvarial lesions were identified. By the beginning of 2021, the patient underwent stem cell harvesting followed by myeloablative/salvage chemotherapy and an autologous stem cell transplant. Soon after he transferred his care to our institution a few months later, the patient underwent a successful stem cell transplant and remained stable for three months post-transplant. The patient began experiencing severe pain which prompted earlier surveillance imaging, and both PET scan and MRI demonstrated extensive disease progression. To improve comfort and confirm etiology, a right thoracentesis was performed, and 1,200 mL’s of bloody fluid was sent to our cytology lab for analysis. The findings are presented below.

At our institution, we use one or two drops of fresh fluid to make a cytospin for triage purposes. We examine the air-dried, Diff-Quik-stained cytospin to determine whether or not the cells are likely to contaminate our routine preparations, and if the cells are overtly malignant, the remaining cytopreparations need to be stained in our known-positive fluid set-up. In this case, the specimen was deemed routine despite the few clusters of cells that do not resemble the predominating mesothelial cells. The tightly clustered group in Image 1 has small, round nuclei; fine chromatin; scant, blue cytoplasm; and atypical mitoses.

Image 1. Pleural fluid, right. DQ-stained cytospin.

The pap-stained smears and liquid-based SurePath preparation were then screened and the cells of interest highlighted below.

Images 2-4. Pleural fluid, right. 2-3, Pap-stained smears; 4, SurePath liquid-based prep.

Similar to the Diff-Quik preparation, the small, round cells of interest are forming tight clusters with each cell approximately twice the size of a normal lymphocyte (Image 3). The nuclei are fairly uniform with fine chromatin, relatively small, inconspicuous nucleoli, and scant cytoplasm (Image 2 & 3). The clustering, while not characteristic of this type of tumor, is most likely artifact due to reactivity of being suspended in fluid. What is not pictured in this specimen is the classic Homer-Wright pseudorosettes.

The cell block sections are consistent with the previous cytopreparations, demonstrating a two-cell population. Small, round blue cells (some forming distorted rosettes) in a background of benign-appearing mesothelial cells.

Images 5-6: pleural fluid, right, H&E cell block sections. 5, 100x; 6, 400x.

This pleural fluid was diagnosed as positive for malignant cells: consistent with the patient’s known Ewing sarcoma. While immunostains were not necessary for this stage IV diagnosis, the cell block sections are expected to stain positive for CD99 (membranous), vimentin, and FLI1 (as an EWSR1 [22q12] break apart rearrangement (83%) was detected in this patient). A few days after this fluid was signed out, the patient passed away, and his family is now undergoing genetic testing.

-Taryn Waraksa, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

Peritoneal Problems

A 74 year old male patient with an extensive cardiac history initially presented to the ER with black stool, warranting a CT scan, upper endoscopy, and colonoscopy, identifying a large, obstructive mass in the colon, smaller, yet unresectable polyps, and subcentimeter liver lesions and lung nodules. The colonic mass was biopsied, consistent with adenocarcinoma; however, the liver lesions were too small to characterize. One month after the onset of symptoms, a right hemicolectomy was performed, and the pathology was signed out as moderately differentiated adenocarcinoma, microsatellite stable, with evidence of lymphovascular and perineural invasion, placing the patient’s stage at IIA (pT3, pN0, cM0). Through shared decision-making, the medical oncologist and patient elected for surveillance due to multiple comorbidities. Forgoing adjuvant therapy, the patient was discharged to physical therapy/rehabilitation. The patient returned for imaging 4 months after his hemicolectomy, demonstrating an enlargement in one of the liver lesions, but then, the patient was lost to follow-up for 20 months.

The patient reestablished care and surveillance imaging, which demonstrated a hypodense liver lesion (in a background of poorly visualized subcentimeter liver lesions), a nonocclusive thrombus in the right portal vein, a heterogenous enhancement of the left portal vein (suggestive of an underlying tumor thrombus), and an 8 cm heterogenous right adrenal mass. Based on the most recent CT scan, the differential diagnoses of the adrenal mass include metastatic disease or a primary adrenal lesion including adrenal cortical carcinoma or pheochromocytoma (for which biochemical analysis should be performed before attempting a biopsy). Extensive peritoneal lymphadenopathy was visualized as well. The area of the right hemicolectomy, however, did not show evidence of recurrence. After biochemical evaluation for metanephrines ruled out a pheochromocytoma, the patient underwent a CT scan-guided adrenal FNA and core biopsy.

The Diff-Quik smear assessed at the time of biopsy revealed a highly cellular specimen, some cells with bare nuclei, enlarged nuclei, and some pseudoglandular structures.

Images 1-2: Adrenal Gland, Right, Fine Needle Aspiration. 1-2: DQ-stained smears

Telepathology confirmed an adequate sample of tumor cells present, and core biopsies were obtained.

The following morning, the pap-stained smears and H&E cell block sections were screened. The cells appeared polygonal with a high N/C ratio and prominent macronucleoli. Cell arrangements formed thickened trabeculae. However, the cytoplasm is more granular than the lipid-rich cytoplasm seen in an adrenal cortical carcinoma. The H&E cell block sections depicted a beautiful trabecular pattern with endothelial cells wrapping the periphery.

Images 3-6: Adrenal Gland, Right, Fine Needle Aspiration. 3-4: Pap-stained smear; 5-6: H&E Cell Block sections.

The preliminary morphology was interpreted as carcinoma, and both cytotechnologist (or cytologist, as we now prefer to be called) and pathologist suggesting features of adrenal cortical carcinoma; however, the IHC markers proved otherwise!

Images 7-9: Adrenal Gland, Right, Fine Needle Aspiration, IHC Cell Block Sections. 7:HepPar1+; 8: Arginase+; 9: pCEA (canalicular pattern)+.

Other differential diagnoses considered renal cell carcinoma and pheochromocytoma (to be safe). The IHC profile ruled out adrenal cortical carcinoma as the cells of interest were negative for inhibin, calretinin, and Melan A. Negative PAX-8, EMA, AE1/AE3, and vimentin staining ruled out renal cell carcinoma, and negative chromogranin, synaptophysin, GATA-3, vimentin, and S100 staining enabled us to safely say that a pheochromocytoma was out of the equation as well. Positive staining for HepPar1, arginase, pCEA (canalicular pattern), and CAM5.2 supported the unlikely diagnosis of metastatic hepatocellular carcinoma (HCC).

This diagnosis placed the patient at Stage IV HCC. It came to light that the patient has a remote history of hepatitis and a high-risk history of drinking, contributing to a poor prognosis. Due to the patient’s condition, they held off on HCV antiviral therapy and decided to observing viral load through regular blood work. The patient and clinician discussed the risks and benefits along with alternatives of systemic therapy, as his multiple comorbidities still pose a significant risk. Immunotherapy was determined to be the best option to delay the progression of his cancer and maintain quality of life.

-Taryn Waraksa, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

E(cto)pic Metastasis

A 72 year old female originally presented with lung carcinoid and bilateral renal masses. The patient’s left kidney biopsy demonstrated ectopic thyroid parenchyma by an outside institution. Her thyroid function tests were unremarkable, she had no known previous head and neck radiation, and to the best of her knowledge, there was no family history of thyroid cancer. She underwent FDG PET imaging, which showed increased bilateral uptake in the neck (thyroid and lymph nodes), and an avid right posterior renal mass. Otherwise, her scan was relatively clear. Her left renal mass was resected and demonstrated thyroid parenchyma, but the differential diagnoses included thyroid heterotopia and metastatic well-differentiated thyroid carcinoma.

FNA and core biopsy were then obtained from the right upper quadrant of the kidney. The findings are depicted below.

Images 1-6: Kidney, Right, Fine Needle Aspiration. 1: Pap-stained smear; 2: DQ-stained smear; 3: H&E Cell Block section; 4: TTF-1+; 5: Thyroglobulin +; 6: CK7+.

The FNA was signed out as “Atypical thyroid tissue present.” Immunohistochemical stains demonstrated positive staining for CK7, vimentin (partial), TTF-1, thyroglobulin, and PAX-8 (partial), and negative staining for RCC, Napsin A, synaptophysin, and chromogranin. While these immunostains suggest thyroid-type tissue, morphology was most worrisome for metastatic thyroid carcinoma. The chromatin presented as hypochromatic and powdery, nuclear grooves and pseudoinclusions were present, and the nuclei were enlarged with irregular membranes. However, the scant material present precluded a definitive diagnosis.

Images 7-8: Kidney, Right, Core Biopsy. 7, H&E section 100X; 8, H&E section 400X.

The core biopsy suggested benign-appearing thyroid tissue similar to that seen in the left kidney, however, the surgical pathologist diagnosed the material as metastatic thyroid carcinoma.

A thyroid FNA was obtained from one of the patient’s multiple right-lobed thyroid nodules consistent with TI-RADS category 5 the next day. This was diagnosed as atypia of underdetermined significance due to scant cellularity.

Images 9-10: Thyroid, Right Lobe, Fine Needle Aspiration. 9: DQ-stained smear; 10: Pap-stained smear.

The right renal mass was resected after molecular profiling was performed on the left renal mass tissue. Mutation Detection by Next Generation Sequencing demonstrated a tumor mutation burden of 3.6Muts/Mb and identified mutations in the PRKDC, PTEN, and KRAS genes. Two kidney tumors were identified in the right kidney (one measuring 8.0 cm and the other 4.5 cm), both diagnosed as metastatic thyroid carcinoma with papillary features.

Images 11-12: Kidney, Right, Resection. 11, H&E section 40X; 12, H&E section 400X.

The thyroid was then resected, and pathologic findings were consistent with invasive follicular carcinoma with extensive angioinvasion to 4 or more vessels. While renal metastases are rare, the high affinity for angioinvasion makes the kidney a prime metastasis site due to its vascular-rich tissue. The patient was prescribed a low iodine diet and Thyrogen-stimulated radioiodine ablation to remove any remaining thyroid tissue or micrometastases and enhance the sensitivity of thyroglobulin as a tumor marker for surveillance purposes. While thyroid cancer (papillary and follicular types) is typically considered “the best cancer to have” due its slow growth and low-risk of widespread malignancy, it doesn’t mean that it won’t metastasize, even to a distant organ that you normally wouldn’t suspect. Great caution must be taken to ensure that lumps, bumps, and swallowing issues are addressed at annual physicals to catch a low-risk cancer before it has the opportunity to become an epic metastasis.

-Taryn Waraksa, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

By the Book

One of my favorite parts of being a cytotechnologist is the delight of having cytology students rotate through our institution as a practicum site. The pandemic caused a clinical rotation hiatus for the safety of both our staff and students, but thanks to widespread healthcare vaccination, we were able to bring in some fresh minds to experience the variety of interesting cases we enjoy every day. I think what I love most about having students here is reminiscing of when I was in their shoes seven years ago. I remember going into my rotations using nothing but morphologic criteria I memorized from lecture and labs. My clinicals served as a rude awakening that we rarely see any textbook perfect cases. Cancer is like a shape-shifter – one melanoma looks entirely different than another. Two lung squamous cell carcinomas from the right upper lobes from two different patients could look entirely different. The unique variation within and between cancer types is what makes this field so beautifully fascinating. The first time a cytotechnology student shows me a case, tells me their thoughts, works through the criteria, and lists the differentials, I look up and say, “nothing is quite by the book.” How often we fall into a routine of relying on criteria, closing our minds to certain diagnoses because it doesn’t quite look like the clinical impression. When the pathologic and clinical impressions divide, more diagnostic tests are performed, CPT codes fill our billing tab, and we start to panic. “It’s supposed to be adenocarcinoma, so why doesn’t it look like adenocarcinoma?!?

A few weeks ago, the lab received a left pleural fluid from a patient who presented with a history of small cell cervical cancer. I remember learning about this in my first semester of grad school – how rare a finding of small cell carcinoma is, accounting for less than 5% of cervical cancers. It essentially mimics small cell carcinoma of the lung and other neuroendocrine carcinomas, where you should be able to identify the telltale salt-and-pepper chromatin, nuclear molding, scant cytoplasm, loosely cohesive or isolated, necrosis, usually an absence of nucleoli, a high proliferation index with mitotic figures, etc. It’s an aggressive disease to say the least, just like its lung counterpart. When this cancer metastasizes, it takes its same characteristics with it, spreading rapidly without care.

The first step in processing a fluid is to prepare a fresh, air-dried, Diff-Quik-stained cytospin to triage the specimen and decide whether the specimen should be processed routinely or hand-prepped and stained with overtly positive fluids to prevent cross-contamination. There was one cluster identified on the Diff-Quik preparation, but compared to the background of mesothelial and inflammatory cells, the tumor content was insufficient to push it up to hand-processing. The bluish cytoplasm caught my attention as a feature of neuroendocrine tumors AND lymphomas, but the nuclear molding had me favoring neuroendocrine.

Image 1. Pleural fluid, left. DQ-stained cytospin.

That afternoon, I examined the pap-stained smears and SurePath liquid-based preparation, identifying similar cells of interest. However, despite the presence of nuclear molding and scant cytoplasm, the nuclei presented with prominent nucleoli. An interesting feature, to say the least.

Images 2-5. Pleural fluid, left. 2-3, Pap-stained smears (2, lightened to highlight nucleoli); 4-5, Pap-stained SurePath liquid-based preparation.

The following morning, I screened the cell block slides and came across molded groups of cells (appearing as a garden aerial view). Still the prominent nucleoli baffled me, and I thought, “Why doesn’t this look like a classic small cell carcinoma? They clinical history even included known lung mets from the patient’s small cell cervical cancer!”

Images 6 and 7: pleural fluid, left. 6, H&E cell block section 100X; 7, H&E cell block section 400X.

When I sent the case for review by the pathologist, I wrote up a diagnosis of Positive for Malignant Cells; Carcinoma, small cell? Recommend correlation with IHC.” My attending was just as intrigued. She ordered a thorough panel of immunohistochemistry stains based on the morphologic findings.

Images 8-11. Pleural fluid, left. 8, synaptophysin+; 9, CD56+; 10, TTF-1+; 11, BerEP4+.

The tumor cells are positive for synaptophysin, CD56, TTF-1, and BerEP4, focally positive for CK7 and chromogranin (not shown), and negative for calretinin, PAX-8, and p40 (also not shown). The findings support the diagnosis of metastatic high grade carcinoma with neuroendocrine differentiation.

While the stains support a diagnosis of small cell carcinoma, the morphologic diagnosis was mildly questionable. I went back to the patient’s record to see what we may have missed in the clinical history. It turns out the patient initially presented with Stage IB2 HPV+, moderately-differentiated cervical adenocarcinoma in 2020. After completing brachytherapy and one cycle of chemotherapy, but could not tolerate additional treatments due to leukopenia and elevated LFTs. Shortly thereafter the patient complained of abdominal pain and a liver mass and bulky lymphadenopathy were identified on imaging. An FNA of a supraclavicular lymph node confirmed not only metastasis of the patient’s cervical cancer, but discovered a small cell/neuroendocrine transformation. And this is why proper documentation of clinical history is so important to pathologists and laboratory professionals. In one of my earlier posts, I preached that cancer doesn’t discriminate; so why should we? Keeping an open mind is paramount to both succeeding in and enjoying the field of cytopathology. If it looks like a duck, and it walks like a duck, it might actually have transformed into a goose.

-Taryn Waraksa, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

The Great Mimicker

I’ve witnessed that radiologists are often hesitant to perform a core biopsy on a spleen due to its vascularity, so when I attend spleen FNAs, I rarely push. Even when the oncology team requests both a core biopsy and FNA for a hematology workup, I will acquire as many passes as possible from an FNA to work up cytomorphology and flow cytometry before risking a core-induced hemorrhagic complication. When I was called to attend an ultrasound-guided spleen biopsy, I went in knowing two things: the patient has both splenic and brain lesions, and I was going to make the most of what I was given. When I arrived in ultrasound, the radiologist informed me that the patient had polycythemia vera (PV), which would explain the splenomegaly, but not the brain lesions. The patient, a 65 year old male, received the diagnosis in 2009 and was managed with phlebotomies for six years until a rising platelet and white blood cell count required an intervention of hydroxyurea. Within 18 months, the patient developed a PE and dizziness and began therapeutic anticoagulation. At the same time, the patient’s “metastatic lesions” were identified on imaging. The first state of business is finding out if his PV had progressed into myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML). There was something… off… about this case though. It didn’t feel like a heme case (not that I prefer non-heme cases… well, okay, I’m a cytotech, so I kind of do). The brain lesions concerned me, and I didn’t know what to infer.

Let’s progress to the FNA. Here are the air-dried, Diff-Quik smears that changed everything.

Images 1-2: Splenic mass, fine needle asperation, DQ-stained smears.

What non-heme surprise is THIS?! Are these histiocytes? There’s no way, those nucleoli are aggressive! Look at the chromocenters! The variation in nuclear size! What epithelial beauty is this? Is it epithelial?! Is that a fibrovascular core? It can’t be sarcomatoid, could it? Twenty inner monologue questions later, “okay, it’s adequate,” I said to the radiologist. But wait… should I do it? Should I press for a core biopsy? The radiologist asked me if I needed anything else for the diagnosis. Perhaps she saw my puzzled expression, compelling her to tell me that she felt the imaging looked weird – she thought it wasn’t a heme case either. At least I’m not alone here. I took a deep breath, expecting the worst and hoping for the best, and I asked the radiologist to collect a core biopsy. She checked the ultrasound Doppler for excessive vascularity, and much to my surprise, she agreed to perform the core. After collecting more FNAs for my cell block and dropping the core biopsies off in surgical pathology, I showed the case to my attending pathologist. He also agreed that we don’t need to triage it for heme. He asked if the patient had any significant history other than PV, splenic lesions, and brain mets, and I told him nothing was reported in his chart. I checked the chart again for good measure while he was running through his differential diagnoses of lung, GI, prostate, etc., and saw the patient was scheduled to see dermatology later that week. I mentioned to that to my attending, and he suggested it could be a melanoma.

The following morning, I examined the pap-stained slide and began to second guess myself. Was I wrong? Could it be a heme case? Hodgkin’s wasn’t in the differential, and these cells look so much more aggressive than Reed-Sternberg cells. At least we have a core biopsy either way. We could run immunos on the cell block and save the cores for molecular. I screened the pap-stained slides a little longer, focusing on the macronucleoli, the owl-eye and eccentric nuclei, the poorly-differentiated nature of this… MELANOMA. YES! So, the cells might not contain any melanin pigment, but my attending’s inkling was exactly right. Waiting for my cell block to arrive, I listed melanoma as my primary diagnosis with a differential of lung or prostate cancer.

Images 3-4: Splenic mass, fine needle aspiration, Pap-stained smear.

The cell block confirmed my non-heme diagnosis and kept my differentials at bay. The attending pathologist ordered an immunohistochemistry profile of S100, HMB-45, and Melan A, as well as AE1/AE3. The first three immunostains (prior to our adoption of SOX-10) confirmed a diagnosis of metastatic malignant melanoma. Soon thereafter, the patient’s primary lesion was identified on his back, and he was treated with radiation and immunotherapy. Unfortunately, the metastases were not responding to the immunotherapy, and a few days after a clinical trial was offered, the patient passed away.

Images 5-8: Splenic mass, fine needle aspiration. 5 and 6, cell block, H&E; 7 and 8, Melan A+.

Melanoma is known as the great mimicker, especially in amelanotic form, and it should always be in the back of your mind as a differential diagnosis. Lack of melanin pigment and a large cherry red macronucleoli leads us to favor lung, prostate, or serous adenocarcinoma), renal cell carcinoma, hepatocellular carcinoma, Hodgkin’s lymphoma, or even an epithelioid sarcoma. This case highlights the need to remember that metastatic melanoma is always a possibility, even when you do not have a primary site or previous clinical history of the disease.

-Taryn Waraksa, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

Cytology Case Study: Little Gland, Big Disease

A 59 year old female initially presented with DCIS, treated by mastectomy and 5 years of adjuvant tamoxifen at another institution. 4 years later, she presented to another hospital with an adrenal mass, uterine fibroids, and an ovarian cyst, where a biopsy and right-sided adrenalectomy confirmed a 10.5 centimeter adrenocortical neoplasm. Margins were close, but negative at <0.1 cm. Microscopically there were areas of necrosis, high nuclear grade, a diffuse growth pattern, and clear cells representing less than 25% of the tumor. A malignancy was favored, but lack of metastasis could not confirm the diagnosis. She presented to the cancer center with stage II adrenal cancer, T2N0M0 and mitotane-induced adrenal insufficiency. Multiple hepatic and pulmonary metastases were subsequently identified and treated with extensive surgery, including a VATS wedge resection, right nephrectomy, hepatic mobilization, lysis of adhesions, dissection of the adrenal vein and vena cava with repair, and resection of an ileal mass.

Palliative radiation therapy targeted the remaining lung nodules, and six cycles of chemotherapy were administered. A CT scan-guided fine need aspiration biopsy was obtained of a 4 centimeter retroperitoneal mass that was suspicious for recurrence on imaging, which cytopathology then confirmed. Taking into consideration the history, my additional differentials included renal cell carcinoma, hepatocellular carcinoma, and plasmacytoma, with metastatic breast cancer as the least likely differential.

Images 1-2. Retroperitoneal mass, right side, FNA-DQ stained smears.
Images 3-4. Retroperitoneal mass, right side, FNA-Pap stained smears.
Images 5-6, Retroperitoneal mass, right side, FNA-H&E cell block sections.

Palliative radiation therapy was then administered to the retroperitoneal and psoas masses, and microwave ablation targeted the segment 3 and segment 6 liver lesions, reducing pain and stabilizing growth, respectively. However, disease continued to slowly progress, so the oncology team sent the retroperitoneal metastasis tissue for molecular testing to assess for potential next lines of therapy. Testing revealed a variant of undetermined significance in MSH6, indeterminate tumor mutational burden, stable MSI, and negative for PDL1. When the case was brought to tumor board, the team recommended ongoing surveillance and palliative therapy (when needed) given the patient’s slowly progressing disease. Often thought of as a rare disease, I’ve examined a fair share of primary and metastatic adrenocortical carcinomas through working at a cancer center. These tiny triangular glands that sit on top of the kidneys have SO much power. Producing and regulating cortisol, aldosterone, and androgenic hormones, the adrenal cortex is an active outer layer. Whether hormonal or neoplastic, it truly is fascinating how a tiny gland could wreak so much large-scale havoc on the human body.

-Taryn Waraksa, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

Mye-Lo-Mye: Cytology Case Study

I quickly transitioned from from learning at Jefferson to teaching at Jefferson less than 6 months after I graduated from the Cytotechnology program. Assuming this new role of teaching future cytotechnologists was a refreshing twist after continuously learning at work. I stood at the podium as an alumna in my previous classroom with a breadth of fresh material to work with, appreciate, and share. At the beginning of each lecture, I would check in on my students, ask them how their other classes were going, ask if they had any questions, and then, I would share an interesting case from my work week thus far. Aside from the weekly lecture on ancillary techniques, such as molecular and immuno-diagnostics, I assigned multiple activities such as journal club/conferences, discussions, and my personal favorite – a mock tumor board. Each student would take turns playing the role of the physician, radiologist, cytotechnologist, pathologist, and oncologist (surgical, medical, or radiation). For my first year of instruction, I had assigned the group their “tumor” of interest, but I quickly encouraged their creativity run rampant during subsequent years.

Despite working in a cancer center and being able to recognize some fairly obscure tumors with little experience, I did not realize how much I still had to learn, even as a part-time lecturer and full-time cytotechnologist. These tumor boards taught me so much more than I expected, and I am forever thankful for the experience of having such wonderfully bright students teach ME! For one very memorable tumor board, the students elected to present the diagnosis and treatment of a male patient with multiple myeloma. Yes, plasma cells! Plasma cell neoplasm, plasmacytoma, multiple myeloma. Awesome, let’s see what this group can do! The “physician” said the patient complained of widespread bone pain, malaise, and recurrent fevers and infections. The “radiologist” presented the images of osteolytic lesions throughout the skull and vertebrae, the latter of which core biopsies and FNAs were obtained. The “cytotechnologist” described a mix of plasmablastic cells, as well as mature and immature plasma cells, some with clock face chromatin and a perinuclear hof (which is my telltale feature that I now emphatically describe to everyone else). The “pathologist” bypassed flow cytometry and performed Kappa/Lambda light chain immunohistochemistry on the core biopsy, diagnosing the patient with multiple myeloma. Unfortunately, due to the extent of the patient’s disease, the “oncologist” and her team could not increase the life expectancy, and the student’s patient expired.

Now, whenever I have a plasma cell neoplasm or multiple myeloma case, I think back to my students and their mock tumor board and everything they taught me. I just recently attended an FNA on a 79-year-old male with a history of multiple myeloma who presented with a PET positive right facial mass and right cervical lymph nodes. The radiologist performed an ultrasound-guided FNA of a right peri-mandibular soft tissue mass, and it took everything in me to not tell the radiologist anything more than “adequate.” But when I saw those perinuclear hofs, I was elated to have a definitive diagnosis!

Images 1-2. Perimandibular Mass, Right, FNA-DQ-stained smears.

Later that afternoon, I couldn’t wait to screen my pap-stained slides. The clock face chromatin was so beautiful! My cell block the following morning highlighted the textbook perfect features diagnostic of a plasma cell neoplasm.

Images 3-4. Perimandibular Mass, right, FNA. Pap-stained smears.
Image 5. Perimandibular Mass, Right, FNA. H&# cell block section.

The attending pathologist ordered a routine myeloma immunocytochemistry panel, including CD138, kappa light chain, lambda light chain, CD20, CD45, and MUM1.

Immunocytochemical stains performed on the unstained paraffin sections showed the tumor to be positive for CD138, kappa light chain, and MUM1, focal equivocal staining for CD20, and negative staining for lambda and CD45. The case was signed out as a plasma cell neoplasm.

Images 6-8. Perimandibular Mass, right, FNA. Cell block section immunocytochemistry. 6, CD138; 7, MUM1; 8, kappa light chain.

The bone marrow core biopsy was sent for Cytogenetic Microarray Analysis and Next Generation Sequencing. The CMA results revealed gains of chromosomes 3, 5, 9, 11, 15, 19, and 21 and losses of chromosome segments 1p and 2p and 7 p in mixed states. Loss of 1p is associated with a poorer prognosis for multiple myeloma. Next gen sequencing identified a tumor mutation burden of 8.4Muts/Mb with mutations detected in the following genes: FAM46C, BRAF, KAT6A, TSC1, KRAS, FLT3, and NFKBIA.

-Taryn Waraksa, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

Eye Spy 2.0

Now that you’ve seen some malignant ocular entities (both primary and metastatic), I’d like to share some (in my eyes) curveballs. Working in a cancer center, I see more cancer than I do benign processes. You can say I’m fine tuned to identifying malignant cells, and I find it difficult to rest assured that what I’m diagnosing truly is benign. When you find one malignant cell in a pleural fluid, changing the patient’s diagnosis from Stage II to Stage IV, that primed search for atypia is always on. With that said, not every specimen screened at a cancer center is malignant. We rule out malignancy and confirm benign processes as well. As for benign ocular FNAs, well… you have to be as certain calling benign as you do malignant because the process of acquiring an additional sample to confirm your diagnosis is just as involved and unpleasant as the first biopsy.

Here, I present six non-malignant eye FNA cytopreparations that had me searching for atypia longer than I expected.

Case 1. A 72 year old female with a history of breast carcinoma; She presented with a choroidal nevus, OD in 2014. The oncologist noted a slight increase in base and thickness six years after the initial evaluation. The clinical diagnosis is a choroidal nevus, rule out low-grade melanoma.

Images 1-2: Eye, Right, Choroid, Fine Needle Aspiration. Pap-stain.

Final Diagnosis: No malignant cells identified. A few benign-appearing melanocytes, consistent with nevus.

Note: Material is scant cellular and consists mainly of scattered retinal pigment epithelial cells and sensory cells. There are a few benign-appearing spindle-shaped cells with bland nuclei, consistent with nevus. However, the paucity of the sample precludes a definitive diagnosis. Recommend clinical correlation to exclude sampling error.

Case 2. A 21 month old male with a history of an iris stromal cyst, OS. Lesion is status post-FNA (twice), had regrowth, and aspirated for the third time. Previous FNAs showed no malignant cells.

Images 3-4: Eye, Left, Choroid, Fine Needle Aspiration. Pap-stain.

Final Diagnosis: No malignant cells identified. Rare pigment-laden macrophages, a few lymphocytes, and benign surface epithelial cells present. (Consistent with stromal cyst).

Case 3. A 65 year old female with a history of nevus, OD. Ocular oncology followed up and noted an increase in thickness from 2.76 to 3.00 mm. In light of family history of cutaneous melanoma, treatment is suggested. Clinical diagnosis: ciliary body melanoma.

Images 5-6: Eye, Right, Choroid, Fine Needle Aspiration. Pap-stain.

Final Diagnosis: Scant mildly atypical amelanotic spindle cells, consistent with nevus.

Note: Diagnostic material is very scant. There are a few benign-appearing single spindle cells and one cluster of cohesive spindle cells with no pigment. Prominent nucleoli are not seen. While the cytologic features in the context of clinical presentation is consistent with nevus, the paucity of the sample precludes a definitive diagnosis. There is no diagnostic evidence of malignant melanoma in this sample. Recommend clinical correlation to exclude sampling error.

Case 4. A 9 year old female with a history of retinoblastoma, OD; status post IAC (intra-arterial chemotherapy) x5, IVit melphalan x 4, and plaque radiotherapy. She recently developed vitreous hemorrhage (OD). Since the hemorrhage is obscuring the retinal view, vitrectomy is planned.

Image 7: Eye, Right, Vitreous, Fine Needle Aspiration. Pap-stain.

Final Diagnosis: No malignant cells identified. Lymphocytes and histiocytes present.

Case 5. An 11 year old female with vitreal retinoblastomas, OU; status post CRD therapy x6; plaque I-125, EBRT, proton beam; IVit Metphalan x2, and PPV (pars plana vitrectomy), OS. She now presents with dense vitreous hemorrhage blocking view since 2019.

Images 8-9: Eye, Left, Vitreous, Fine Needle Aspiration. Pap-stain.

Final Diagnosis: No malignant cells identified. Proteinaceous material and scattered small lymphocytes present.

Case 6. A 52 year old female with no cancer history. In 2019, she presented with pain, floaters, decreased visual acuity, and photophobia, OS. She was treated with antibiotic drops and steroids. In 2020, she was noted to have panuveitis with yellow iris nodules. FNA showed necrotic cells inadequate for diagnosis, as well as a negative culture and PCR for HSV, toxoplasmosis, CMV, and VZV. Clinical diagnosis: suspicious for iris lymphoma, OS.

Images 10-11: Eye, Left, Anterior Chamber, Fine Needle Aspiration. Pap-stain.

Final Diagnosis: Mixed inflammatory cells, favor an inflammatory process.

Note: There are neutrophils, lymphocytes, and scattered histiocytes in a background of numerous fragments of an amorphous substance. The cellularity is inadequate for flow cytometric analysis. We performed immunocytochemical stains on cytospin preparations. The lymphocytes are predominantly T-cells showing positive staining for CD45 and CD3 and negative for CD20. There is no diagnostic evidence of malignant lymphoma in this sample. The amorphous substance may represent lens fragments, in which case the possibility of lens-induced uveitis should be considered in the differential diagnosis. Recommend clinical correlation.

As you can tell, benign processes are complicated and require thorough explanations in our pathologic diagnoses, especially when they differ from the clinical impression. The majority of benign ocular FNAs are paucicellular, and we make the most of what we have through optimal preservation and preparation of cellular material. And don’t forget-not everything is cancer. Cyst contents, proteinaceous debris, and inflammatory cells make up a share of cases we see, and it’s okay to diagnosis them as benign. Just keep an eye out.

-Taryn Waraksa, MS, SCT(ASCP)CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.