Double trouble: an unusual combo of tumors with (un)usual molecular events

 

 

78-year-old male was admitted to hematological department with worsening anemia (despite iron supplementation) and persistent leukocytosis. Recently the patient was diagnosed with bronchitis and received two courses of oral antibiotics; however, due to the remaining leukocytosis and elevation of C reactive protein (CRP) leukemia was suspected. At the time of admission, the patient complained of periodic left upper quadrant abdominal pain, night sweats and headaches. He also had a history of myocardial infarction 11 years ago and was now suffering from congestive heart failure and gout.

Complete blood count showed leukocytosis (WBC 12,16 x10⁹/L), neutrophilia (9,30 x10⁹/L) and anemia (HgB 105 g/l). CRP was elevated up to 82,4 mg/l. Also elevated were LDH (339 U/L) and creatinine (149 µmol/l). Moreover, monoclonal protein was detected. Immunofixation revealed IgA kappa biclone and urine analysis showed monoclonal free light kappa chains.

Bone marrow aspirate flow cytometry revealed 1,39% clonal kappa+ plasma cells with bright CD56 and heterogenous CD117 expression.

Trephine biopsy showed hypercellular marrow for age with an eye-catching focus of confluent trabeculae and nests with some lumen-like formations, surrounded by desmoplastic stroma (Fig.1-2). Tumor cells were large, with abundant amphophilic cytoplasm and large, oval or round, vesicular nuclei with distinct nucleoli (Fig.3). Perineoplastic lymphoid infiltration and eosinophilia was also noted (Fig.4). Tumor cells showed diffuse expression of keratin 7 and partial expression of high molecular weight keratin. Partial expression of Cadherin-17(Fig.5), diffuse expression of Islet-1(Fig.6) and, intriguingly, alfa-fetoprotein (Fig.7) was also seen. Moreover, strong diffuse expression of p53 indicated TP53 gene mutation (Fig.8). No mucine (MUC2/MUC5A/MUC6) expression was noted. Full immunophenotype of the metastatic tumor:

• CK7+; CKHMW+/-; CK5-/+; CK20-;
• Cadherin17+; AFP+; Islet1+;
• p53+ (likely mutated);
• Glypican-3/Pax8/RCC/CD10/TTF1/NKX3.1/CDX2/CD117/CD5/SATB2/GATA3/p504s(-);
• Muc2/Muc5AC/Muc6(-);
• BRAF(-);
• Her2(-);
• Ki67 prolif. index up to 30%.

Hematopoietic tissue architecture was distorted, with attenuated granulopoiesis and “punched out“ nodules of NASDE negative infiltrates (Fig.9-11). The infiltrate was composed of intermediate and large CD138 positive plasma cells (Fig.12). Intriguingly, although CD138 positive infiltrate was quite abundant (about 20% of the bone marrow hematopoietic cells, Fig.13), it was barely perceptible on H&E, except for large cells, that demonstrated Russell bodies – eosinophilic, homogenous immunoglobulin inclusions (Fig.14-15).  Plasma cell infiltrate showed IgA/kappa restriction (Fig.16-17) and also had an aberrant expression of CD56 (Fig.18) and focal expression of CD117. About 60% of the infiltrate also demonstrated cytoplasmic staining with anti-BRAF V600E (Fig.19-20). Surprisingly, strong cytoplasmic expression of Glypican-3 was also seen in the majority of neoplastic plasma cells (Fig.21). Full immunophenotype of the infiltrate: CD138+; CD56+; CD117+/-; IgA/kappa+; BRAF+; Glypican-3+; CD20/Pax5/CyclinD1(-); Ki67 prolif. index 20%.

The final proposed diagnose:

a)       Metastatic (ductal?) adenocarcinoma involving bone marrow. The most probable site of the primary tumor is upper gastrointestinal tract (gastric vs pancreatic/bile ducts).

b)      Multiple myeloma (nodular and interstitial pattern), 20% of bone marrow hematopoietic cells.

Simultaneous abdominal ultrasound examination revealed two irregular hypoechoic liver nodules (involving segments II-IV), the larger of the two measuring 100x70 mm. Considering pathological finding, most probable primary tumor was intrahepatic cholangiocarcinoma.

Skeletal radiographic examination revealed no osteolytic lesions.

Plasma cell myeloma clinically was rather smoldering: there was no definite myeloma related symptoms (keeping in mind the difficulty distinguishing from the symptoms, caused by the metastatic solid tumor) and bone lesions. Hence, no urgent myeloma directed treatment was indicated. The patient was then referred to an oncologist for the comprehensive examination, staging and treatment of the solid tumor.

 

 

This case has lots of interesting points. First of all – two synchronous neoplasms within the same bone marrow trephine biopsy. Multiple myeloma (MM) association with solid tumors as well as hematological neoplasms is a known phenomenon. In a study of Hasskarl et al., 10% of MM patients had a different neoplasm with 78% of these cases being solid tumors¹. Most of the solid tumors (53%) were diagnosed prior to MM, 20% were diagnosed subsequent to MM and only 5% were diagnosed synchronously¹. Most commonly reported cases of MM with synchronous solid tumors include renal clear cell carcinoma, lung adenocarcinoma and small cell carcinoma, prostate adenocarcinoma, colorectal adenocarcinoma, breast adenocarcinoma and gastric adenocarcinoma. There are only sparse published case reports (4) of synchronously via trephine biopsy diagnosed MM and solid tumor, three of them being MM with prostate adenocarcinoma^2-4 and one being lung adenocarcinoma⁵. Hence, this is the first case report of synchronously diagnosed MM and metastatic cholangiocarcinoma on the trephine biopsy.

Other interesting aspect of the case is aberrant expression of alfa-fetoprotein (AFP) in intrahepatic cholangiocarcinoma (ICC). Elevation of serum AFP and AFP immunohistochemical (IHC) expression is typically seen in hepatocellular carcinoma (HCC), as well as yolk sack tumor and tumors of hepatoid or enteroblastic differentiation. Although AFP is usually negative in ICC and serve as a useful marker differentiating between ICC and HCC, Zhou et al. reported, that 24,4% of ICC patients had elevate serum AFP levels⁶. AFP seropositive ICC patient group was also significantly associated with HBsAg and liver cirrhosis, hence clinically similar to HCC⁶. A recent study by Zhang et al. found a corresponding ratio (17,7%) of AFP seropositive ICC cases and worse overall survival, in comparison to ICC patient with normal AFP serum levels⁷. AFP seropositive patients also had a higher proportion of HBV infection, a more advanced T stage and TNM stage⁷. Although elevated serum AFP level is not that uncommon, we found only four case reports with immunochemical expression of AFP in ICC^8-11. This paradox most likely is due to the absence of larger studies on ICC immunophenotype, including anti-AFP staining.

Another interesting finding of the present case is the partial BRAF V600E expression in the neoplastic plasma cells. Although BRAF is a well-established cancer driver gene, it is often found to be only subclonal in MM patients. Hence, it appears that BRAF can function both as initiator of MM and as well as potentiator of the tumor progression^12-13. Rustad et al. found that BRAF V600E clone size in MM, evaluated by whole exome sequencing was comparable with that found by IHC, when the clone was dominant¹⁴. The current case demonstrates subclonal diversity of the MM infiltrate not only by partial BRAF V600E IHC expression but also by detection of IgA kappa biclone by immunofixation. Interestingly, MAPK pathway mutations (KRAS, NRAS or BRAF), found mutually exclusive in other tumors, are commonly present together in MM¹⁵. Moreover, in some patients this phenomenon is not due to subclonal diversity but are observed at the same clone^16-17.

After reviewing the studies of BRAF mutated plasma cell neoplasms, we can conclude, that BRAF mutations are only detected in MM and plasma cell leukemia and is not present in MGUS and smoldering MM^18-19. Studies investigating BRAF mutated MM prognosis demonstrate contradicting results. One of the first study of BRAF mutated MM by Andrulis et al. found patients with BRAF mutated MM to have significantly shorter median and overall survival (OS) and high prevalence of extramedullary disease (EMD)¹⁹. This study also demonstrated a case with impressive results of mutation-specific BRAF inhibitor vemurafenib, in a patient with BRAF V600E mutated and relapsed myeloma with extensive EMD, achieving partial response after one course of vemurafenib and continuing to be in a stable remission after eight courses of this treatment¹⁹. On the contrary, Rustad et al. found no significant difference between BRAF V600E mutation status in MM and the progression free survival, OS and EMD¹⁴. The authors stressed that differences of the results could be explained by different selection of patients and concluded that the presence of BRAF V600E mutation at diagnosis or early in the disease is not correlated with aggressiveness of MM, regardless of clone size¹⁴. Other study from China found significant correlation of BRAF V600E mutation with shorter OS and PFS only in younger MM patient group (<65 years)²⁰.

Due to the genetic heterogeneity of the disease, question of the treatment choice in BRAF mutated MM is more difficult than it seems. Although BRAF inhibition in vitro downregulates MAPK pathway in BRAF mutated subclones, it causes paradoxical upregulation in BRAF-wildtype MM cells²¹. Therefore, only patients with predominant BRAF mutated MM clone could benefit from the targeted BRAF therapy. Moreover, adding broad acting drugs in combination with BRAF inhibitors, might be the best solution¹⁴.

Finally, the last topic for discussion is the unexpected glypican-3 (GPC3) expression in a subset of MM cells. GPC3 is a cell-surface glycoprotein, playing an important role in cell growth, differentiation and motility²². GPC3 modulate WNT signaling pathway, that is activated in several types of tumors and metabolic disorders²². Although GPC3 is most commonly enrolled in development and progression of HCC, it is also associated with other tumors²³. Nonetheless, we couldn’t find any case of GCP3 expressing MM. While various GPC3 targeting therapies are under investigation, with the main focus being HCC treatment, it might also be beneficial for the patient of the present case.