Clinical guidelines – Non CUH WGS referrals

Please see NHS England's test selection (opens in a new tab) or the national genomic test directory (opens in a new tab) for up to date guidance.

Common indications for WGS include:

• Paediatric & teenage/young adult (TYA) tumours (any tumour <25 years of age)
• Sarcoma (any site, any age)
• Triple negative breast cancer, glioma, and high grade ovarian carcinoma (any age)
• Failed standard of care treatment or diagnostic testing (ie. refractory to current standard maximal treatment or diagnosis not possible using non-WGS techniques; any tumour type, including carcinoma of unknown primary, any age)

The decision to proceed to WGS should ideally be made at MDT in conjunction with radiological, clinical and pathological input, but in certain situations it may be appropriate for the treating clinician to make this decision independently.

Further details on eligibility for molecular tests in cancer can be found in the national genomic test directory.

An additional specialised consent process is required for WGS, particularly with reference to the implications of any germline variants that may be discovered.

Consent must be received prior to any biopsy of a tumour for WGS in which tissue (or extra tissue) is taken for the sole purpose of WGS.

The consent process requires completion of a training course “Patient Choice: Discussing Whole Genome Sequencing with Patients” (opens in a new tab).

Each patient’s (and family member where relevant) choices are captured via a nationally standardised Record of Discussion (RoD) form and additional forms where relevant. For WGS to be carried out a RoD must be received by the GLH for each individual undergoing WGS. The patient choice conversation can be performed remotely and the form submitted without a patient signature – please tick the “Remote consent” section of the RoD form to indicate this.

The completed and signed consent form (record of discussion) can either be e-mailed to the lab (with e-mail subject title “Cancer WGS consent”) or posted to the lab with specimen (see address below).

To perform WGS, FRESH TISSUE IS REQUIRED.

This can be taken either:

1. as part of the diagnostic procedure up front (e.g., in paediatric, sarcoma or TYA cases)
2. as part of the treatment (e.g. in a diagnostic/therapeutic resection such as a hysterectomy), or
3. purely to obtain tissue for WGS (e.g., in a case where only formalin fixed tumour is currently available).

To perform WGS, an adequate quantity of tissue is required. To yield adequate quantities of DNA, the following guidelines are offered:

• For core biopsies - tissue totalling ~40mm long (for 16g needle) or 80mm (18g)
• For excisions/excisional biopsies - a fragment of tissue measuring 5 x 5 x 2mm

Please note that these are MINIMUM recommendations.

Detailed guidance on tissue sampling is available in the “Sample Handling Guidance for Whole Genome Sequencing of Solid Tumour Samples” document below, videos are also available (opens in a new tab).

Send the specimen fresh to the histopathology laboratory as soon as possible (ideally within 30 minutes) and ensure it is handled by trained laboratory staff on receipt. If delivery is delayed, store the specimen temporarily at 4°C. Do not place the tissue in any solution or fixative.

Tissue can also be sent in RNAlater (opens in a new tab) (a solution that stabilises nucleic acids at room temperature). An RNAlater/tissue volume ratio of at least 5:1 is needed and tissue fragments must be no more than 5mm in thickness to allow sufficient penetration of the solution.

Note: If a combined diagnostic/WGS procedure is being done using RNAlater and it is either a) being performed out of hours or b) being performed at a centre without a histopathology department, two separate tissue samples are needed. One formalin sample for pathology analysis and an RNAlater sample for WGS. These must be provided in separate specimen pots by the submitting clinician (see guidance below).

To order RNAlater tubes

Email cuh.add-tr.freshtissuehisto@nhs.net or phone 01223 348213 (ask for Section Leader). Please provide a responsible contact and delivery address. We will send 6 × 5 mL RNAlater universals.

Submitting RNAlater samples to histopathology

If the histopathology lab that performs routine histopathology for your Trust is able to perform frozen-section tumour-purity assessment, this should be done using the guidance provided in the next section and then sent to East Genomics.

If the histopathology lab your Trust uses is unable to perform frozen-section tumour-purity assessment, samples should be placed into the appropriate solutions and sent to the CUH histopathology lab with a request form at:

CUH Histopathology
Goods-In 1 Discovery Drive
Trumpington
Cambridge
CB2 0AX

The histopathology team will perform frozen-section review at CUH and then send the sample to East Genomics for WGS.

Key steps are summarised as follows:

On reception in the histopathology laboratory:

• If material is also required for histopathological diagnosis, this should be separated from the fresh specimen, fixed and processed separately.
• In combined diagnostic/WGS procedures (especially with core biopsies) tissue for formalin fixation for routine diagnosis must be prioritised before sampling for WGS (as a guideline for cores, at least two good cores should be fixed).
• The fresh specimen should then be frozen, ideally into at least two separate vials, or placed into RNAlater.

The next step is histopathological assessment:

• Frozen sections should be cut and histologically assessed
• Tumour sampling should be confirmed
• The tumour specimen submitted should be at least 30% tumour cellularity, with <20% necrosis*
• It is acceptable to submit multiple vials of the same tumour to increase yield

*On occasions, it may be appropriate to submit specimens not meeting these criteria, please contact the laboratory to discuss such cases.

See the “Sample Handling Guidance for Whole Genome Sequencing of Solid Tumour Samples" document earlier in this page for further detials.

NOTE: Please ensure there is sufficient fixed material for routine diagnostics before sending the remaining tissue for WGS.

Why we need it

A germline sample provides essential reference point for WGS analysis. The germline sample reflects the patient's healthy genome to compare against the tumour (somatic) genome. Comparing the germline and biopsy samples allows identification of variants that are specific to the tumour, which are relevant to diagnosis, prognosis and treatment. Variants in the germline sample also indicate inherited variants that contribute to cancer risk, and can provide insights into how a patient will respond to certain treatments.

Sample collection

The preferred germline sample is peripheral blood. Required volumes are indicated in the table below. In exceptional circumstances, a saliva sample (using the Oragene collection tube) may be used as a source of germline DNA.

A germline blood sample can be taken at the same time as the biopsy/surgery and sent to our labs together with the tumour sample. Alternatively, the sample can be taken at a different time and sent separately. If sending separately, the blood sample must be accompanied by its own fully completed WGS test order form.

Detailed guidance on germline sampling is available via the FutureNHS (opens in a new tab) NHS Genomic Medicine Service workspace at (see “GLH Sample Handling Guidance for Germline Samples” and “Guidance for Whole Genome Sequencing of Haematological Malignancies”).

The WGS requesting form (opens in a new tab) is available (see Appendix for details on navigating the auto populated form).

A blank form (“NHS Genomic Medicine Service test order form – Cancer” (opens in a new tab)) is also available.

Please complete all fields. We will not be able to submit the sample without the required details.

If there is more than one tumour (distinct tissue mass) to be tested, please complete separate referral forms for each tumour and provide a different identifying description to distinguish them (e.g. 3 o'clock and 12 o'clock).

Please note that frozen tumour specimen must be kept frozen during transport, ideally on dry ice.

Important note: Tissue that has been frozen upfront without prior fixation in RNAlater cannot subsequently be placed in RNAlater for transport purposes. Tissue that has been frozen upfront must be sent frozen as described above.

RNAlater specimens may be sent cooled (preferred) or ambient. Ensure vials are labelled “RNAlater” in addition to PID.

For longer-term storage before dispatch, incubate overnight at 4 °C to permit solution penetration, then store at –20 °C/–80 °C as needed. (RNAlater supports ≤1 week at RT and ≤1 month at 4 °C.)

Genomics England stipulate that the results of WGS studies should be discussed at a GTAB.

A quorate GTAB will include an oncologist, a histopathologist, a clinical scientist and if applicable a clinical geneticist.

The GLH clinical scientist/molecular pathology team (which may be based in Cambridge or, in some cases, at your local centre) will present the findings to the GTAB colleagues for discussion.

Following the discussion, and the completion of any additional work (e.g. further characterisation or confirmation using an orthogonal test of pertinent variants) a report will be issued.

Access further information and guidance.

If a standard discussion pathway has not been established previously, the GLH will e-mail the responsible clinician inviting them to a suitable GTAB forum for discussion of results.