1. Cancer Unknown Primary
Sarah W. Gordon, DO
Assistant Professor
Virginia Commonwealth University

2. What is cancer of unknown primary?
Clinical syndrome representing many different types of cancer
Heterogeneous group: many primary sites with varying biology
Goal remains: improved diagnosis so that site-specific treatments can be applied

3. Frequency
Pavlidis, N, et al. Eur J Cancer

4. Pathologic Evaluation
Histology examination remains gold standard for initial evaluation
Five major light microscopy histologic diagnoses:
Poorly differentiated neoplasm
Poorly differentiated carcinoma (± feature of adenoca)
Well/moderately well-differentiated adenoca
Squamous cell carcinoma
Neuroendocrine carcinoma

5. Other histologies? Sarcoma Melanoma
Treat as per established guidelines

6. Poorly differentiated neoplasm
35-65% found to be lymphoma
Primarily Non-Hodgkin’s
Remaining tend to be carcinomas
Includes poorly differentiated neuroendocrine
Melanoma & sarcoma < 15%

7. Poorly differentiated neoplasm
To determine differential diagnosis: IHC, electron microscopy, genetic analysis
Most common cause of nonspecific light microscopy diagnosis is inadequate or poorly handled biopsy specimen
Avoid FNA: histologic pattern is not preserved and limited ability to perform special studies

8. Poorly differentiated carcinoma
~ 30% of carcinomas of unknown primary
~ 1/3 have features of poorly differentiated adenocarcinomas
Need IHC staining
Electron microscopy can be useful to identify by subcellular features
Lymphoma
Occasionally: sarcoma, melanoma, mesothelioma, neuroendocrine tumors

9. Poorly differentiated carcinoma
Consider histologically atypical germ cell tumors
Cytogenetics: abnormalities of chromosome 12 may be diagnostic of germ cell tumors
At autopsy, primary site only found in 40% of patients

10. Adenocarcinoma Most common tumors identified by light microscopy
~ 60% of CUP diagnoses in US
Typically elderly patients with multiple sites of metastatic tumors
Certain histologic features associated with particular tumor type, but still not specific
Papillary features in ovarian ca
Signet cells with gastric cancer

11. Adenocarcinoma
Autopsy series of patients with adenocarcinoma of unknown primary site (884 patients, 1944 – 2000), most common sites:
Lung
Pancreas
Hepatobiliary tree
Kidney
2/3 of cases
Pentheroudakis G, et al. Eur J Cancer

12. Squamous Carcinoma ~ 5% of patients with CUP
Diagnosis usually made by histology
Can consider IHC or molecular studies if poorly differentiated squamous carcinoma, especially if clinical presentation atypical

13. Squamous Carcinoma Upper & midcervical lymphadenopathy: H&N primary
Consider ENT referral, DL, possible tonsillectomy
Inguinal lymphadenopathy: often detectable primary site in genital or anorectal area
Adenopathy other sites (including lower cervical + supraclav): usually primary lung cancer

14. Neuroendocrine Carcinoma
~ 3% of call CUP
Three histologic subgroups:
Well-differentiated/low-grade neuroendocrine: histology similar to carcinoids/islet cell tumors, frequently secrete bioactive substances
Small-cell carcinoma/atypical carcinoid/poorly differentiated neuroendocrine carcinoma: typical neuroendocrine features and aggressive histology
Poorly differentiated neoplasm/poorly differentiated carcinoma

15. Extragonadal Germ Cell Tumors
Poorly differentiated carcinoma or adenocarcinoma of unknown primary site with some, but not all characteristics:
Young age
Male gender
Predominant tumor location in mediastinum or retroperitoneum
Elevated of hCG or AFP
Presence of 12p chromosomal gain (isochromosome 12p)
IHC staining for octamer binding transcription factor 4 (aka POU domain class 5 transcription factor 1)
Excellent response to chemotherapy & some are cured with cisplatin-based regimens
Treat like poor prognosis germ cell tumors

16. Immunohistochemistry
Conner, JR, et al. Adv Anat Pathol

17. Immunohistochemistry
Oien, K. Semin Oncol

18. Immunohistochemistry
Oien, K. Semin Oncol

19. Immunohistochemistry
Oien, K. Semin Oncol

20. Immunohistochemistry
Fizazi, K, etc al. Ann Oncol

21. Figure 1. Basic immunohistochemical work-up of carcinomas of unknown primary. Reproduced with permission: Varadhachary GR. Carcinoma of unknown primary origin, Gastrointest Cancer Res 2007; 1: 229–235.
From: Cancers of unknown primary site: ESMO Clinical Practice Guidelines for diagnosis, treatment and follow-up
Ann Oncol. 2011;22(suppl_6):vi64-vi68. doi: /annonc/mdr389
Ann Oncol | © The Author Published by Oxford University Press on behalf of the European Society for Medical Oncology. All rights reserved. For permissions, please 21

22. Immunohistochemistry
Problems:
Technical expertise of performing tests accurately and reproducibly
Proper interpretation by experienced pathologist
False-positives, false-negatives
Classic staining patterns often overlap among primary sites

23. Electron Microscopy Reliable in undifferentiated sarcoma
Ultrastructural features:
Neurosecretory granules: neuroendocrine tumors
Premelanosomes: melanoma
Absence of cannot be used to rule out specific diagnosis
Should not be used for adenoca of unknown primary site

24. Karyotypic or Cytogenetic Analysis
Specific chromosomal abnormalities are well characterized in several hematopoietic neoplasms
Limit to patients with histologic diagnosis of poorly differentiated neoplasm or poorly differentiated carcinoma
No aid in evaluation of adenocarcinomas

25. Gene Expression Profiling
Cancer cells retain some normal cell-type specific functional characteristics in their gene expression profile
Usually origin can be predicted regardless of neoplastic differentiation
Molecular profiling assays designed to determine type of cancer do not look at tumor-specific markers, but rather gene expression dynamics in relation to cell lineage

26. DNA array technology.
DNA array technology. Tumor cDNA and normal cDNA are labeled with different fluorescent dyes. The labeled cDNAs are hybridized to DNA arrays containing oligonucleotides representing known genes. The fluorescence intensity is analyzed using statistical algorithms, and color intensities are assigned to the ratios of gene expression.
David M. Mintzer et al. The Oncologist 2004;9:
©2004 by AlphaMed Press

27. Histologic Dx: poorly differentiated carcinoma with pleomorphic features.
Comment: Immunohistochemical staining performed on the needle core biopsy (1A) shows that the malignant cells are positive for CK7, GATA3, Uroplakin 2/3 (patchy), CDX2, p63 (focal, LungSquamous 2 multiplex), CD56 (patchy), while negative for CK20, TTF1, synaptophysin, CK5 (LungSquamous 2 Multiplex), and OCT 3/4, . Control sections stain appropriately.
The malignant cells are poorly differentiated and have ambiguous features. The immunophenotype, with positivity of CK7, GATA3, Uroplakin 2/3 (patchy) and p63 (focal), suggests a urothelial origin. CDX2 is also positive, which can be seen in a subset of poorly differentiated urothelial cell carcinoma. However, a lung primary squamous cell carcinoma cannot be definitively excluded. Clinical and radiologic correlation is necessary.

30. Fizazi, K, etc al. Ann Oncol. 2015.

31. Focused Diagnostic Eval
Intial Evaluation
Additional Evaluation
Women with features of breast cancer (bone, lung, liver mets; CK7+)
Breast MRI
ER, GCDFP-15, HER2 stains
Women with features of ovarian cancer (pelvic/peritoneal mets; CK7+)
Pelvic/vaginal US
WT-1 stain
Mediastinal/retroperitoneal mass
Testicular US
Serum HCG, AFP
PLAP, OCT4 stains; FISH for i(12p)
Features of lung cancer (hilar/mediastinal adenopathy; TTF-1 +)
Bronchoscopy
Features of colon cancer (liver/peritoneal metastases; CK20+/CK7–, CDX2+)
Colonoscopy
Poorly differentiated carcinoma, with or without clear cell features
Stains for chromogranin, synaptophysin, RCC, Hepar-1, HMB-45
(If Hepar-1+, obtain serum AFP; if neuroendocrine stains +, obtain octreotide scan)
Adapted from DaVita, et al. Cancer

34. Clinical Management Algorithm
Fizazi, K, etc al. Ann Oncol

35. Fizazi, K, etc al. Ann Oncol. 2015.

36. Study Site Contact: Alison Ryan (SIIT Team)
SWOG – 1609
Local PI: Sarah Gordon
Study Site Contact: Alison Ryan (SIIT Team)

39. NCI-Molecular Analysis for Therapy Choice (NCI-MATCH / EAY131)
A phase II precision medicine cancer trial
Co-developed by the ECOG-ACRIN Cancer Research Group and the National Cancer Institute
Version Date: 11/27/2016

40. What is NCI-MATCH? Now 6000 **Screening about 500 pts/month!
11/27/2016

41. NCI-MATCH Tumor Gene Testing
aMOI: actionable mutation of interest
11/27/2016

42. NCI-MATCH Testing Expectations
One in every four to five (23%)
11/27/2016

43. NCI-MATCH Objective
To determine whether matching certain drugs or drug combinations in adults whose tumors have specific gene abnormalities will effectively treat their cancer, regardless of the cancer type
This is a signal-finding trial—treatments that show promise can advance to larger, more definitive trials
11/27/2016

44. NCI-MATCH Updates! NCI will no longer be doing screening
No more on-trial biopsies – NCI site reimbursement will end
Deadline to consent patients has passed
6 new arms to open (June 2017)
May be more treatment arms to come…
If patients go on treatment, biopsy at time of progression is strongly encouraged.
Funding for this expected to continue

45. NCI-MATCH Updates!
Outside assays will now be accepted for screening from designated labs
Foundation Medicine (live now)
CARIS
MDACC institutional assay
MSKCC institutional assay
Any samples sent to these labs from a CIRB approved trial site will be screened for eligibility to some arms of MATCH
Start with rare gene variants (< 1.5%)
19 arms
Delay before opening other arms (2-3 months)

46. Patients do NOT need fresh biopsy! Archived tissue allowed
Other Changes…
Patients do NOT need fresh biopsy!
Archived tissue allowed
Treatment after biopsy is allowed
No more off treatment waiting period between biopsy & genetic results

47. Other Changes…
If patient has a MATCH by the outside lab, tissue will be requested to be sent to NCI-MATCH for verification
Study team will need to coordinate sending archived specimens
If MATCH  patient gets treated
If no confirmation by NCI-MATCH of outside assay results, then patient will be assessed for toxicity only, not response
New version of genetic assay being introduced
Increasing # of gene fusions from 271 to 762
Also adding new genes

48. Cases?