Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards

ASCO University
Sep 14, 2016 8:48 AM

User Instructions: Welcome to the Molecular Oncology Tumor Board Series! This educational initiative is a collaboration between the American Society of Clinical Oncology (ASCO), College of American Pathologists (CAP), and Association for Molecular Pathology (AMP).

A new case will be presented each month with discussions led by an expert pathologist and medical oncologist. Have an interesting case in mind? Submit your hypothetical patient cases for consideration in an upcoming Molecular Oncology Tumor Board discussion forum.

This month’s topic is led by Drs. Dung Le (Johns Hopkins) and Chanjuan Shi (Vanderbilt University).

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Please see the attached for supplemental resources related to the case.

Comments

13321

ASCO University
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 14, 2016 8:52 AM

Patient Case

Age/Sex: 47-year-old female

Medical History: Resected stage II microsatellite instability high cecal cancer (T3N0M0, poorly differentiated, 15 lymph nodes evaluated) at the age of 27.  Resected endometrial cancer at the age of 42.

Co-morbidities: Neuropathy

Family History: Brother with colon cancer at the age of 40.  Mother with endometrial cancer at the age of 42 and colon cancer at the age of 50.

History of Present Illness:  Presented with 1 week of rectal bleeding.  Given family and personal history of cancer was seen immediately for a colonoscopy which revealed a sigmoid mass.   Biopsies were obtained which revealed poorly differentiated adenocarcinoma.  CT scan revealed multiple enlarged retroperitoneal lymph nodes and a single large liver metastasis.  Biopsy of the liver metastasis also revealed a poorly differentiated adenocarcinoma.  CEA was 120 ng/dl.

Pathology specimen from multiple biopsies of liver metastases:
Source of specimen:  Biopsy of liver metastases
Gross description:  3 cores of white-pink tissue measuring 1.0 to 1.2 cm in length and 0.1 cm in diameter
Diagnosis: INVASIVE POORLY DIFFERENTIATED ADENOCARCINOMA. No non-neoplastic liver tissue included in specimen.
Molecular testing order: RAS and BRAF mutation, mismatch repair IHC for MLH1, MSH2, MSH6, and PMS2
Pre-analytics:  Review of histopathology; marking of slide for high-cellularity tumor; recuts of block for unstained sections for microdissection for tumor enrichment and immunohistochemistry; DNA extraction and quality control checks
Analysis: Molecular diagnostics laboratory for RAS mutations and for BRAF (V600E). Immunohistochemistry for MSH2, MLH1, MSH6, and PMS2 mismatch repair gene products.

Type of Tumor: Poorly differentiated adenocarcinoma. BRAF wild-type.  KRAS mutation (G12D). Mismatch repair proteins are intact.

Relevant Markers:  CEA 120 ng/ml

13326

ASCO University
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 14, 2016 8:53 AM

Discussion Questions

1.    What pathologic features are frequently associated with MSI-H colorectal cancers?

2.    What are the NCCN guidelines for Lynch syndrome screening in patients with colorectal cancer or endometrial cancer? Was immunohistochemistry for MLH1, MSH2, MSH6 and PMS2 sufficient for screening for Lynch syndrome based on this patient’s medical history?

3.    What adjuvant therapy would you have recommended at her diagnosis of stage II disease and why?

13331

Gary Spitzer, MD
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 14, 2016 6:20 PM

<p>These tumors are frequently associated with an extensive lymphoid infiltration and are more ofter right sided&nbsp;</p>

<p>given a colon cancer before age 50 a lynch associated&nbsp;cancer ( uterine) and the family history this is compatible with lynch more than the other hereditary colon cancer syndromes . This at the time of diagnosis could have included formal molecular&nbsp;MSI&nbsp;testing. But now can include genetic panel testing directed to hereditary colon cancer</p>

<p>given the probability&nbsp;that MSI tumors are not sensitive to FU, stage&nbsp;2 with adequate LN and no adverse features no adjuvant therapy would be recommended&nbsp;</p>

<p>&nbsp;</p>

<p>a small percentage of Lynch can be negative with immunochemistry and should reflex with this age of onset and family history to&nbsp;</p>

<p>&nbsp;</p>

<p>&nbsp;</p>

13341

Victor Manuel Villalobos, MD, PhD
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 16, 2016 10:04 AM

Do the authors think immunotherapy in MSI-H patients overtake chemotherapy in the neoadjuvant/adjuvant setting? 

13366

Dung T. Le, MD
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 18, 2016 12:56 PM

At this time, outside of melanoma, there is not a lot of evidence for immunotherapy in the adjuvant setting.  However, the idea is very appealing that we could potentially avoid chemotherapy altogether in a subset of our patients.  There are studies currently in development in the adjuvant setting.  Neoadjuvant in a purely resectable setting would be interesting but a study could be challenging because of the cure rate and we would have to get to them before the surgeons do!

13371

Chanjuan Shi
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 19, 2016 8:17 AM

Course Faculty Response

MSI-H colorectal cancers (CRCs) as a group show some distinct pathologic features compared to microsatellite stable (MSS) tumors. They are always associated with increased tumor-infiltrating lymphocytes and prominent Crohn’s-like lymphoid reaction. Medullary, mucinous and signet ring cell histologic types are more often associated with MSI-H status; however, not all of tumors with such morphologies are MSI-H tumors. Therefore, immunohistochemical (IHC) stains for MLH1, MSH2, MSH6 and PMS2 or PCR-based MSI testing is required to determine microsatellite instability.

IHC, MSI testing or both can be used as an initial screening test for Lynch syndrome. Approximately 90% of MSI-H CRCs show loss of nuclear staining for one or more of the 4 mismatch repair (MMR) proteins. Loss of MLH1 or MSH2 is consistently accompanied by loss of PMS2 or MSH6, respectively. (Figure 1). However, loss of PMS2 or MSH6 does not always result in degradation of MLH1 or MSH2. While lack of nuclear MLH1 and PMS2 expression can be seen in Lynch syndrome-associated and sporadic MSI-H CRCs, loss of nuclear expression of PMS2, MSH2+MSH6, or MSH6 always indicates Lynch syndrome. Any CRCs with loss of MLH1 and PMS2 by IHC should be tested for BRAF (V600E) mutation and/or MLH1 promotor methylation. For patients with tumor(s) showing loss of MLH1/PMS2 with no BRAF mutation and no MLH1 promotor hypermethylation or loss of any other proteins (MSH2, MSH6, or PMS2), germline evaluation should be carried out for the genes corresponding to the absent proteins. For tumors with loss of MSH2, a test for EPCAM deletions should be included as EPCAM deletions can cause somatic hypermethylation of MSH2, which is responsible for approximately 1% of Lynch syndrome-related CRCs.

PCR-based MSI testing has a similar sensitivity (about 90%). A fluorescence-based PCR assay from Promega has been used by most laboratories in the US, which includes five mononucleotide repeat markers (microsatellite markers) and two pentanucleotide repeat markers (identity markers). Both normal tissue (can be replaced by blood sample) and tumor are needed for comparison. For tumors showing MSI-H by MSI testing, testing for the BRAF mutation and/or MLH1 promotor methylation assay should also be followed. IHC for the MMR proteins can be used to determine which protein(s) are affected.

Using either MSI or IHC testing alone can result in false-negative results in about 5-10% MSI-H CRCs. A low tumor cellularity is one of the main reasons for false negative MSI testing results. Generally a tumor cellularity of >20% is needed for MSI testing.  In some cases, to reach a tumor cellularity of >20%, tumor enrichment by microdissection or macrodissection is needed. The IHC testing can be falsely negative in treated cancers.  And it may not be reliable in small biopsies. In addition, some mutations in the MMR genes do not result in the absence of a detectable protein product (but non-functional). Therefore, for patients who are highly suspicious for Lynch syndrome (like this case), if one test is negative, another test should be processed.

Both metastatic and primary tumor can be used for the IHC or MSI testing.

13376

Dung T. Le, MD
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 19, 2016 8:20 AM

Course Faculty Response

At the patient’s initial presentation at the age of 27 with and MSI-H stage II colon cancer, adjuvant therapy was not indicated.  Her tumor did not exhibit any high risk features.  Poor differentiation is often seen with an MSI-H tumor.  Stage II, MSI-H tumors have a good prognosis and do not benefit from 5-FU adjuvant therapy (Sargent 2010, NCCN 2016).  For this reason, MMR or MSI testing should be performed for all patients with stage II disease.  Furthermore, Lynch syndrome tumor screening should be performed for all patients diagnosed with colorectal cancer at age < 70 and those > 70 who meet the Besthesda Guidelines.

The guidelines also suggest consideration of screening of patients with endometrial cancer who are age < 50 or with a family history of endometrial or colorectal cancer.  This patient’s young age, personal history of 2 colon primaries, endometrial cancer, and strong family history should have triggered a referral to genetics for further evaluation and also triggered further investigation into the testing of the liver metastases which was reported as having intact mismatch repair proteins.

13381

ASCO University
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 19, 2016 8:23 AM

Patient Case Update

The patient was treated with FOLFOX and bevacizumab and responded for 4 months before her CEA began to rise and a CT confirmed progressive lymphadenopathy and growth in her liver lesion.  Her oncologist started her on FOLFIRI and bevacizumab and --at the patient’s request-- sent her liver tumor for next generation sequencing.  She was referred to a clinical trial testing a programmed death-1 (PD-1) inhibitor in colorectal cancers with mismatch repair deficiency.  However, she was turned away because of the history of intact mismatch repair proteins in her liver tumor specimen.  Her oncologist persisted and sent the tissue for PCR-based MSI testing.

How tissue was acquired/preserved: Formalin fixed paraffin embedded liver biopsy was used for next generation sequencing and PCR-based MSI testing. A blood sample in an EDTA (purple top) tube was used as a normal control for MSI testing.

Testing Platform/Molecular Profiling: Additional molecular testing order for Next generation sequencing and MSI testing :  

Pre-analytics: Review of histopathology; marking of slide for high-cellularity tumor (>20%); recuts of block for unstained sections for microdissection for tumor enrichment; DNA extraction and quality control checks.

Analysis: Mutational testing in Clinical Laboratory Improvement Amendments (CLIA)-accredited laboratory with laboratory-developed 31-gene “hot spot” next generation sequencing solid tumor panel. Fluorescence-based PCR assay from Promega for MSI testing.

13386

ASCO University
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 19, 2016 8:24 AM

Discussion Questions

1.    What is the recommendation for MMR testing in patients with stage IV colorectal cancer?
2.    What is the current evidence for PD-1 inhibition in MMR deficient colorectal cancer?
3.    Is PD-L1 expression a predictive marker for response to anti-PD-1 therapy in metastatic colorectal cancer?

Gary Spitzer, MD
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 21, 2016 3:35 PM

if tested on the initial tumor, negative and no criteria to suspect lynch , would not test.

however given the fact that many patients do not fit lynch screening criteria, and , if not previously tested, if Braf positive  would only test for MMR if as stated eligible to get immunotherapy

if braf negative would test for MMR  and if positive reflex to germline testing , given the other tumors that could occur in a patient like this and complicate management . given the duration of responses to  immunotherapy, secondary tumors could become an issue 

PD-1 inhibition is very active in MSI tumors but not in MSS tumors , these tumors generally express PD-1 and the MSS do not 

presumably, this explains the lack of correlation of  response and survival to PD-1 expression 

 

13391

Anis Toumeh, MD
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 19, 2016 10:33 AM

The data for using chekpoint inhibitors to target PD1 are encouraging. The immune-related response rate reported by Le et al. NEJM 2015 is very encouraging at 40% in pateints received 2 or more prior therapies. To my knowledge, for now, testing for MMR in the metastatic setting is done for clinical trials purposes. 

13406

Chanjuan Shi
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 23, 2016 8:29 AM

Course Faculty Response

In non-small cell lung cancers and melanomas, PD-L1 expression by IHC has been used as a predictive marker for response to anti-PD-1 therapy. In metastatic CRCs, MSI status can be used as a predictive marker for the response; however, the predictive role of PD-L1 expression by IHC appears to be limited (Le, 2015). A clinical trial showed that MMR proficient CRCs demonstrated minimal response to anti-PD-1 therapy, whereas the immune-related objective response rate and progression-free survival rate were 40% and 78%, respectively, for MMR deficient CRCs. In addition, some MMR deficient, but PD-L1 negative CRCs responded to anti-PD-1 therapy. Furthermore, in MSI-H CRCs, PD-L1 is often expressed by inflammatory cells (including macrophages and lymphocytes) at the invasion front, rather by tumor cells (Figure 2). An accurate evaluation of PD-L1 expression might require resection specimens, which may not be available in patients with metastatic disease. Therefore, for the time being, MSI status but not PD-L1 expression is used to identify CRCs that would respond to anti-PD-1 therapy.

13411

Dung T. Le, MD
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 23, 2016 8:33 AM

Course Faculty Response

In addition to the previously mentioned criteria for MMR or MSI testing, the guidelines now also recommend testing be performed for all patients with metastatic disease.  This only leaves a couple of groups of patients that may not undergo routine testing (Stage I or III patients who are age > 70 and do not meet the Besthesda guidelines) (Umar, 2004).

MMR deficient tumors are being targeted for immunotherapy because the DNA repair deficiencies lead to accumulation of hundreds to thousands of mutations that could lead to neoantigens that can be recognized by the immune system.  The endogenous lymphocyte infiltration and PD-L1 expression on immune infiltrating cells have been clues as to their potential immunogenicity.

Inhibition of the PD-1/PD-L1 axis is showing to be a promising therapeutic target for MSI-H tumors.  While the incidence of MSI-H status in stage IV disease is < 5%, the promise of the therapy for these subsets of patients lies in the potential longevity of response.

In a study testing PD-1 inhibition in MMR deficient CRC, MMR proficient CRC, and MMR deficient non-CRC solid tumors, the response rates were 40% (N=10), 0% (N=25), and 71% (N=7) respectively.  The duration of response, progression-free survival, and overall survival were not reached in the MMR deficient cohorts (Le, 2015).  While not yet published, there is now data on over 80 patients in the MMR deficient cohorts of this study.  

A subsequent study testing PD-1 inhibition with and without cytotoxic T-lymphocyte antigen (CTLA-4) inhibition in MMR deficient CRC is reporting response rates of 26% (N=47)and 33% (N=27).  The responses again are proving to be durable (Ngeow, 2016).

Registration trials are underway.

13436

Chanjuan Shi
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 28, 2016 10:34 AM

Course Faculty Summary

  • MSI-H tumors share some pathologic features including prominent intratumoral lymphocytes and Crohn’s like lymphoid reaction.  Certain histologic types, including mucinous, signet ring cells and medullary carcinomas, are more associated with MSI-H CRCs.
     
  • Using either MSI or IHC testing alone can cause false-negative results in about 5-10% MSI-H CRCs. Pathologists should be aware of the limitations of both tests.
     
  • MSI status may be a better predictive marker than PD-L1 expression in CRCs (especially in metastatic CRCs) for response to anti-PD1 therapy.

 

13441

Dung T. Le, MD
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 28, 2016 10:36 AM

Course Faculty Summary

  • Clinical and pathologic features may suggest mismatch repair deficiency:  female, young age, right sided tumors, poor differentiation, medullary features, signet cell features, mucinous histology, multiple primaries, family history, BRAF mutation, lymphocytic infiltrate, and increased mutation number.
     
  • In the correct clinical context, both MMR IHC and MSI PCR may be warranted as neither is 100% sensitive.
     
  • Widespread testing for MMR deficiency is now recommended for identification of Lynch syndrome patients, and for treatment decisions in stage II and IV disease.  Studies in stage III disease are also being proposed (NCCN).
     
  • Immunotherapy will likely become a treatment option for more patients as more testing is implemented and studies are completed.
     
  • While not addressed by this particular case, sporadic cases often associated with BRAF mutation or MLH1 hypermethylation can also benefit from immunotherapy.

13446

ASCO University
Re: Colorectal Cancer (September 2016): Molecular Oncology Tumor Boards
Sep 28, 2016 10:39 AM

Thank you to Drs. Le and Shi for leading the discussion of this case and also to all of those who contributed to the conversation! The forum is now closed to further comments and participants can claim CME credit and 1 ABIM MOC point through ASCO University.

Please check back in mid-October for a new case in this series related to lung cancer.  
 
Have an interesting case in mind? Submit your hypothetical patient cases for consideration in an upcoming Molecular Oncology Tumor Board discussion forum.


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