Clinical utility of anti-p53 auto-antibody: Systematic review and focus on colorectal cancer

Table 1 Cumulative reported frequencies of anti-p53 auto-antibody (anti-p53) in controls and individual cancers n (%)

Group	Ref.	Anti-p53 positive	Summary of study and tumour type
	Park et al[107]	4/79 (5)	Comparative study with lung cancer
Healthy/Benign	Wu et al[133]	9/879 (1)	Case-control study of anti-p53 in various cancers
	Kulić et al[134]	1/20 (5)	Comparative study with breast carcinoma
	Suppiah et al[130]	0/28 (0)	Comparative study with colorectal carcinoma
	Cai et al[125]	0/30 (0)	Comparative study with oesophageal carcinoma
	Atta et al[135]	5/29 (17.2); 13/26 (50)1	Comparative study with hepatocellular carcinoma
	Mattioni et al[136]	0/64 (0)	Comparative study with gastric carcinoma
	Akere et al[137]	4/45 (8.9)	Comparative study with hepatocellular carcinoma
	Müller et al[123]	0/57 (0); 0/379 (0)2	Single study of anti-p53 in various cancers
	Chang et al[85]	0/40 (0)	Comparative study with colorectal carcinoma
	Fonseca et al[95]	0/15 (0)	Comparative study with glioma
	Shimada et al[82]	10/205 (6.3); 13/189 (7)3	Multi-institutional study of anti-p53 in various cancers
	Neri et al[138]	0/51 (0)	Comparative study with lung carcinoma
	Numa et al[139]	0/9 (0)	Comparative study with uterine, ovarian, cervical carcinoma
	Mack et al[140]	1/46 (2.2)	Comparative study with SCLC
	Chow et al[141]	1/28 (3.6)	Comparative study with head and neck carcinoma
	Moch et al[142]	2/130 (1.5)	Comparative study with skin carcinoma (SCC/BCC)
	Hofele et al[143]	0/80 (0)	Comparative study with oral SCC
	Hagiwara et al[144]	0/13 (0)	Comparative study with oesophageal carcinoma
	Ralhan et al[145]	4/50 (8)	Comparative study with lung carcinoma
	Bielicki et al[111]	0/28 (0)	Comparative study with colorectal carcinoma
	Soussi[90]	35/2404 (1.5)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	102/4924 (2.1)
	Blanchard et al[146]	24/97 (28)	Correlates with decreased overall and disease free survival
Oesophageal	Wu et al[133]	4/29 (13.8)	Case-control study of anti-p53 in various cancers
	Cai et al[125]	18/46 (39.1)	Correlates with advanced histological grade, stage, lymph node metastases and decreased tumour response following radiotherapy
	Müller et al[123]	10/50 (20)	No correlation with stage or prognosis
	Bergström et al[147]	31/42 (73.8)	No correlation with clinico-pathological parameters, tumour size or survival
	Shimada et al[82]	90/301 (29.9)	Multi-institutional study of anti-p53 in various cancers
	Kozłowski et al[148]	20/75 (26.6)	No correlation with stage, lymph node metastases or size.
	Shimada et al[99]	14/35 (40)	Correlates with tumour p53 protein expression but not clinico-pathological parameters
	Hagiwara et al[144]	13/46 (28)	Correlates with increased stage and tumour p53 protein expression but not prognosis
	Ralhan et al[145]	36/60 (60)	Correlates with tumour p53 protein expression and missense mutations but not clinico-pathological parameters.
	Soussi[90]	85/274 (31)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	345/1055 (32.7)
Head/Neck16	Wu et al[133]	1/20 (5.0)	Case-control study of anti-p53 in various cancers
	Shimada et al[82]	10/31 (32.3)	Multi-institutional study of anti-p53 in various cancers
	Chow et al[141]	23/75 (31)	Correlates with nodal metastases but not prognosis
	Total	34/126 (27.0)
Oral	Wu et al[133]	5/15 (33.3)	Case-control study of anti-p53 in various cancers
	Hofele et al[143]	19/102 (18.6)4; 12/24 (50)5	Correlates with poor prognosis
	Castelli et al[149]	3/61 (18.7); 9/13 (69.2)3	Serum anti-p53 is useful as a screening tool in pre-malignant lesions
	Soussi[90]	309/1062 (29.1)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	348/1219 (28.5)
Ovary	Wu et al[133]	5/12 (41.6)	Case-control study of anti-p53 in various cancers
	Qiu et al[150]	36/92 (39.1)	Correlates with p53 expression, not clinico-pathological parameters
	Shimada et al[82]	2/27 (7.4)	Multi-institutional study of anti-p53 in various cancers
	Numa et al[139]	8/30 (27)	Correlates with p53 tumour expression and poor prognosis
	Abendstein et al[151]	28/113 (25); 21/113 (19)6	Correlation between serum and ascitic anti-p53. No correlation with stage or grade. Anti-p53 in ascites associated with poor prognosis
	Soussi[90]	140/635 (22)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	219/909 (24.1)
Colorectal(detailed results in Table 2)	Wu et al[133]	11/66 (16.7)	Case-control study of anti-p53 in various cancers
	Suppiah et al[130]	20/92 (21.7)	No correlation with stage or prognosis
	Nozoe et al[97]	17/36 (47.2)	Correlates with advanced lymph node status and stage
	Müller et al[123]	63/197 (32)7; 7/46 (15.2)8	No correlation with stage or prognosis
	Chang et al[85]	47/167 (28.1)	p53 mutation, not anti-p53, correlates with poor prognosis
	Lechpammer et al[88]	40/220 (18.2)	? Correlation with stage or prognosis in Dukes’ A/B1
	Shimada et al[82]	46/192 (23.9)	Multi-institutional study of anti-p53 in various cancers
	Forslund et al[84]	24/88 (27.3)	Correlates with p53 mutation
	Tang et al[89]	130/998 (13)	Correlates with advanced lymph node involvement but not prognosis
	Broll et al[152]	20/130 (15.4)	No correlation with stage or prognosis
	Takeda et al[98]	17/27 (63)	95% negative sero-conversion within 3 wk post-surgery
	Shiota et al[112]	18/71 (25.4)	Correlates with advanced stage and poor prognosis
	Bielicki et al[111]	30/145 (20.7)	? Correlation with Dukes’ A →B
	Soussi[90]	307/1244 (24.7)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	797/3719 (21.4)
HCC	Wu et al[133]	15/93 (16.1)	Case-control study of anti-p53 in various cancers
	Atta et al[135]	28/41 (68.3)	Correlates with advanced stage and shorter survival.
	Akere et al[137]	5/41 (12.2)	Correlates with increased Okuda stage
	Müller et al[123]	19/80 (23.8)	Non-significant trend towards poor prognosis
	Charuruks et al[153]	26/141 (18.4)	Correlates with stage but not tumour p53 protein expression
	Tangkijvanich et al[154]	16/121 (13.2)17	Preliminary report of Charuruks et al (2001). No correlation with severity, stage or prognosis. Survival too short for survival analysis (3 mo vs 4 mo)
	Sitruk et al[155]	19/159 (12)	Correlates with multinodular, infiltrative tumour but not survival
	Soussi[90]	82/387 (1.2)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	210/1063 (19.8)
	Wu et al[133]	0/11 (0)	Case-control study of anti-p53 in various cancers
Bladder	Müller et al[123]	3/24 (12.5)	No correlation with prognosis
	Watanabe et al[156]	17/63 (27)9	Correlates with higher grade, stage, lymph node metastases and tumour p53 protein expression, but not prognosis
	Gumus et al[157]	14/80 (17.5)	Correlates with tumour p53 protein expression and poor prognosis.
	Gumus et al[158]	25/76 (33)	Negative sero-conversion post-treatment (35%, 8/23) associated with good prognosis.
	Shimada et al[82]	4/33 (12.1)	Multi-institutional study of anti-p53 in various cancers
	Morita et al[159]	12/100 (12)	Correlates with stage, and p53 protein expression but not prognosis
	Wunderlich et al[160]	4/32 (12.5)	Correlates with tumour protein p53 expression but not stage.
	Soussi[90]	8/29 (27.6)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	70/385 (18.2)
Lung	Park et al[107]	28/82 (34.1)	Sensitivity study with other markers for lung cancer
	Wu et al[133]	13/95 (13.7)	Case-control study of anti-p53 in various cancers
	Bergqvist et al[161]	14/84 (16.6)	No correlation with tumour volume. Correlates with survival in adenocarcinoma, but not SCC
	Bergqvist et al[162]	12/58 (20.7)	No correlation with tumour volume or lymph node metastases
	Neri et al[138]	2/30 (6.7)10; 8/48(16.7)11	No correlation with stage, histology or prognosis. Non-significant increased survival in LC but not MM
	Cioffi et al[163]	35/109 (32.1)	Low sensitivity, but high specificity (100%) and accuracy (69%). Only 14% agreement with other tumour markers (CEA/TPA, CYFRA21-1, NSE.)
	Zalcman et al[126]	20/97 (20.6)	Correlates with poor prognosis in limited stage SCLC, but not all SCLC
	Mack et al[140]	4/35 (11.1)12; NSCLC 13/99 (13.3)13	Correlates with stage and prognosis in NSCLC but not SCLC
	Shimada et al[82]	18/125 (14.4)	Multi-institutional study of anti-p53 in various cancers
	Soussi[90]	219/1282 (17.1)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	373/2049 (18.2)
Cervix	Shimada et al[82]	10/53 (18.9)	Multi-institutional study of anti-p53 in various cancers
	Numa et al[139]	12/86 (14)	No correlation with tumour p53 protein expression or prognosis
	Total	22/139 (15.8)
	Wu et al[133]	7/43 (16.3)	Case-control study of anti-p53 in various cancers
Gastric	Qiu et al[150]	19/61 (31.1)	Correlates with tumour size but not prognosis.
	Mattioni et al[136]	17/111 (15.3)	Correlates with tumour p53 protein expression, prognosis and survival
	Lawniczak et al[164]	16/71 (22.5)	Correlates with tumour type and age, but not stage or prognosis
	Müller et al[123]	14/122 (11.5)	No correlation with prognosis
	Shimada et al[82]	13/123 (10.6)	Multi-institutional study of anti-p53 in various cancers
	Nakajima et al[165]	13/81 (16)	Correlates with lymph node metastases but not stage or prognosis
	Maehara et al[166]	23/120 (19.2)	Correlates with increased stage and tumour p53 protein expression but not prognosis
	Soussi et al[90]	105/727 (14.1)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	227/1459 (15.6)
	Nozoe et al[167]	15/42 (35)	Correlates with grade 3 and triple negative cancer
Breast	Wu et al[133]	9/25 (16)	Case-control study of anti-p53 in various cancers
	Kulić et al[134]	21/61 (35)	Correlates with decreased 5 year survival
	Müller et al[123]	17/50 (34)	Non-significant trend towards poor prognosis
	Gao et al[168]	31/144 (21.5)	Correlates with stage, lymph node metastases, ER negative, c-erb-2 and tumour p53 protein expression
	Shimada et al[82]	13/71 (18.3)	Multi-institutional study of anti-p53 in various cancers
	Volkmann et al[169]	18/165 (10.9)	Poor concordance between recombinant/native p53 ELISA, immunoblot and immunofluorescence
	Metcalfe et al[87]	155/1006 (15.4)	No correlation with stage and prognosis
	Soussi[90]	296/2006 (14.8)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	539/3467 (15.5)
Uterus	Wu et al[133]	1/13 (7.7)	Case-control study of anti-p53 in various cancers
	Shimada et al[82]	5/22 (22.7)	Multi-institutional study of anti-p53 in various cancers
	Numa et al[139]	5/41 (12)	No correlation with tumour p53 expression/prognosis (see Cervix, Ovary)
	Total	11/79 (13.9)
Pancreas	Wu et al[133]	0/17 (0)	Case-control study of anti-p53 in various cancers
	Müller et al[123]	5/22 (22.7)	Increase sensitivity in conjunction with CA19-9. No correlation with prognosis.
	Shimada et al[82]	3/28 (10.7)	Multi-institutional study of anti-p53 in various cancers
	Ohshio et al[170]	19/82 (23.2)	No correlation with tumour p53 expression or prognosis
	Soussi[90]	60/650 (9.2)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	87/799 (10.9)
Lymphoma	Messmer et al[171]	19/120 (15.8)	Associated with 17p deletions
	Wu et al[133]	0/18 (0)	Literature review of anti-p53 in various cancers (1979-1999)
	Soussi[90]	19/248 (14.3)	Case-control study of anti-p53 in various cancers
	Total	38/386 (9.8)
Biliary tract16	Wu et al[133]	1/8 (6.3)	Correlates with tumour p53 protein expression but not stage
	Limpaiboon et al[172]	6/49 (12.2)	Multi-institutional study of anti-p53 in various cancers
	Shimada et al[82]	1/6 (16.7)	Correlates with tumour p53 mutation
	Tangkijvanich et al[173]	6/82 (7.3)
	Total	14/145 (9.7)
Haematological	Wu et al[133]	8/33 (25)	Case-control study of anti-p53 in various cancers
	Shimada et al[82]	32/364 (6.3)14	Multi-institutional study of anti-p53 in various cancers
	Soussi[90]	14/428 (3.3)15	Literature review of anti-p53 in various cancers (1979-1999)
	Total	54/825 (6.5)
Glioma	Wu et al[133]	1/24 (4.2)	Case-control study of anti-p53 in various cancers
	Fonseca et al[95]	5/24 (20.8)	No correlation with p53 protein but increased in patients < 16 years
	Shimada et al[82]	2/31 (6.5)	Multi-institutional study of anti-p53 in various cancers
	Soussi[90]	6/144 (4.2)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	14/223 (6.3)
Prostate	Wu et al[133]	1/8 (12.5)	Case-control study of anti-p53 in various cancers
	Shimada et al[82]	4/23 (17.4)	Multi-institutional study of anti-p53 in various cancers
	Soussi[90]	4/148 (2.7)	Literature review of anti-p53 in various cancers (1979-1999)
	Total	9/179 (5.0)
Skin	Moch et al[142]	3/105 (2.9)	No difference between controls and patients. Increased in aggressive SCC (8%) vs slow-growing BCC (1.5%)
Testicular	Soussi[90]	0/144 (0)	Literature review of anti-p53 in various cancers (1979-1999)
Melanoma	Soussi[90]	0/58 (0)	Literature review of anti-p53 in various cancers (1979-1999)
Total		3419/18595 (18.4)	All cancers (1979-2012)

Cancer types are listed in order of decreasing anti-p53 auto-antibody frequency. The reported studies within each cancer type are listed in reverse chronology. Squamous cell carcinoma (SCC); basal cell carcinoma (BCC), hepatocellular carcinoma (HCC), carcino-embryonic antigen (CEA). Tissue polypeptide antigen (TPA), CYFRA21-1, Neurone-specific enolase (NSE), Oestrogen receptor (ER), c-erb-2.

¹Cirrhosis;

²Benign disease;

³Oral pre-malignant lesions-excluded from calculation;

⁴Primary carcinoma;

⁵Secondary/recurrent carcinoma;

⁶Ascitic titre, not included in calculation of serum titres;

⁷Colon;

⁸Rectum;

⁹Upper renal tract tumours, excluded from anti-p53 titres in bladder carcinoma;

¹⁰Malignant mesothelioma (MM);

¹¹Lung carcinoma (LC);

¹²Small cell lung carcinoma (SCLC);

¹³Non-small cell lung carcinoma (NSCLC);

¹⁴Myeloma;

¹⁵Leukaemia;

¹⁶Tumour type not specified;

¹⁷Excluded as is preliminary report of the same cohort (duplicate) reported in Charuruks et al.

Table 2 Anti-p53 auto-antibody in all published colorectal cancer studies and key findings n (%)

Ref.	Method and manufacturer	Samples	Follow-up	Key findings
Suppiah et al[130]	ELISA (p53 ELISAPLUS, Calbiochem, Darmstadt, Germany)	20/92 (21.7); 0/20 (0)10/8 (0)2	Median 97 mo	No correlation with tumour stage, differentiation or location. Multivariate analysis show only Stage (Dukes’ and TNM) to be independent prognostic factors
Nozoe et al[97]	ELISA (Pharmacell, France)	17/36 (47.2)	Not stated	Anti-p53-ab (+) associated with greater lymphatic invasion (94.1%; 16/17 vs 68.4%; 13/19), nodal involvement (70/6%; 12/17 vs 17.6%; 3/17) and advanced stage (P = 0.02). Anti-p53 frequency higher in p53 protein expressing tumours (74%; 14/19 vs 18%; 3/17). Only 3 patients with Dukes’ A CRC, all sero-negative
Muller et al[123]	Immunoblot	Colon 63/197 (32); Rectum 7/46 (15.2); 0/57 (0)1 0/379 (0)2	CRC patients enrolled into trial with 5 year follow-up	No correlation with clinico-pathological parameters or prognosis. Trend toward higher anti-p53 sero-positivity in N2/3 disease, poor differentiation and metastases. There were no patients with Dukes’ A in this study. Anti-p53 independent of CEA and CA19-9 with 16% information gain. This is the only study to report negative to positive sero-conversion (3.6%, 11/303)
Chang et al[85]	ELISA (p53-AK, Dianova, Hamburg, Germany)	47/167 (28.1); 0/40 (0)1	Median 36.3 mo (4-58)	Anti-p53 correlates with p53 mutation (43% vs 18%) but not tumour p53 expression, clinico-pathological features or prognosis. p53 mutations, advanced stage and pre-operative CEA > 5 ng/mL were independent prognostic factors (in that order). p53 mutation strongly associated with advanced stage and poor differentiation
Lechpammer et al[88]	ELISA (ELISAPLUS Oncogene Research Products, Cambridge, United States)	40/220 (18.2); 0/42 (0)1	40 patients up to 20 wk; 8 patients up to 48 wk	Anti-p53 had higher tumour p53 expression (70% vs 52%). Anti-p53 frequency shows highest increase in Dukes’ A (0%, 0/28) →Dukes’ B: (24%, 21/87) but no increase in progression to Dukes’ C (18%,19/105). No correlation with overall tumour grade or metastases. Anti-p53 reflects tumour load following surgery, during chemotherapy and with disease recurrence
Shimada et al[82]	ELISA (Anti-p53 EIA Kit II, MESACUP anti-p53 Test; MBL; Nagoya, Japan)	46/192 (23.9); 10/205 (4.9)1; 13/189 (6.9)2	Not reported	Validation study for MESACUP ELISA using prevalence of anti-p53 in various cancers. Good intra- and inter-assay coefficient of variation of 1.85-2.37% and 0.3-3.2% respectively. Demonstrates stability of anti-p53 titres at room temperature for 7 d and following 10 freeze-thaw cycles. No comment on correlation with clinico-pathological parameters or prognosis
Forslund et al[84]	ELISA (Dianova, Hamburg, Germany)	24/88 (27)	Not reported	Cross-sectional study on relationship between p53 mutations and anti-p53 presence. Frequency of p53 mutation higher in anti-p53 sero-positive group (92%, 22/64 vs 34%, 22/64) Correlation with clinico-pathological and survival parameters not reported
Tang et al[89]	ELISA (Calbiochem-Novabiochem, Darmstadt, Germany)	130/998 (13);2/211 (1)3	Not reported	Anti-p53 sero-positivity increases in progression from N2→N3 (2.9%-10.6%); but not N0→N1 (11.7%-12.3%), N1→N2 (12.3%-10.6%) or M0→M1 (12%-17%). No correlation with CEA, overall TNM stage or metastases. Anti-p53 associated with shorter survival in uni- but not multi-variate analysis. Largest study on anti-p53 in CRC
Broll et al[152]	ELISA (p53-autoantikorpfer ELISA, Dianova, Hamburg, Germany)	20/130 (15); 0/44 (0)1	Median 25.5 mo	Anti-p53 positive predictive value of 100%, but accuracy 37% and negative predictive value 29% due to poor sensitivity (15%). Anti-p53 correlated with p53 expression (P < 0.05), but not TNM stage, grade or location (exact numbers not shown). Approximately 70% of series Stage I/II CRC
Takeda et al[98]	Anti-p53 EIA(PharmaCell, Paris, France)	17/27 (63); 1/38 (2.6)3	Up to 2 yearsMedian not reported	Anti-p53 correlates with p53 protein expression and independent of CEA and CA-19-9. Sero-conversion in 94% (16/17) within 3 wk of endoscopic resection. No correlation with clinico-pathological parameters or prognosis/recurrence as all patients had early superficial CRC (23 mucosal, 4 submucosal invasion). This study reports exceptionally high anti-p53, especially considering very early CRC
Takeda et al[174]	ELISA (anti-p53-EIA kit, Pharmacell, Paris, France)	40 patients with anti-p53 ab from previous studies	Up to 29 mo	No correlation between post-operative anti-p53 sero-positivity and histological (depth, lymphatic or venous invasion) or clinico-pathological features of lymph node or liver metastases. High (96%; 27/28) sero-conversion in patients with complete tumour resection. No sero-conversion in patients with residual disease.
Shiota et al[112]	ELISA (GIF, Munster, Germany)	18/71 (25); 1/18 (6)3	Not stated, median survival 56 mo anti p53 ab negative	Anti-p53 correlates with TNM stage (Stage I-IIIb: 9%, 4/45 vs IV: 56%, 14/25), Dukes’ stage (A-C: 9%, 4/45 vs D: 56%, 14/25), CEA, CA19-9 and tumour p53 protein expression. Anti-p53 associated with shorted survival (56 mo vs 20 mo) and is weak poor prognostic indicator. Anti-p53 prognostic significance secondary to other factors, including weak factors e.g., CEA and CA19-9. Only small number of Stage I-IIIb patients
Bielicki et al[111]	ELISA (Dianova, Hamburg, Germany)	30/145 (21); 0/20 (0)2; 0/8 (0)3	Not stated. Cross sectional study	No correlation with Dukes’ Stage (A/B: 22%, 16/73 vs C/D 19% 14/72), size, location, CEA. Highest increase in anti-p53 frequency from Dukes’ A (0%, 0/6) to Dukes B1 (28%, 5/18) but no further difference in progression to Dukes’ C (19%, 7/36). Only 6 Dukes’ A patients in study, all sero-negative
Soussi[90]	ELISA/WB/IP	307/1244 (24.7)	ELISA/WB/IP	Review combining all studies with different methodologies from 1979-1999. Range of sero-positivity (12.5%-68% in 11 studies)
Total (1999-2009)		479/2409 (19.9)		All modern studies (1999 onwards) using commercial ELISA only, with one exception using Immunoblot (Muller et al, 2006)
Review Total (1979-2009)		786/3653 (21.5)		All studies on anti-p53 in CRC (1979-2009)

Studies prior to 1999 used different methodology and not included (see above). Enzyme-linked immunosorbent assay (ELISA); Western blotting (WB); Immunoprecipitation (IP)

¹healthy,

²benign disease,

³adenoma. The study by Muller et al was included despite using immunoblot technique as it was a recent study with relatively large sample size. CRC: Colorectal cancer; CEA: Carcino-embryonic antigen.

Table 3 Prevalence of anti-p53 auto-antibody and carcino-embryonic antigen in studies reporting the presence of both tumour markers in colorectal cancer n (%)

	CEA	Anti-p53
Tang et al[89]	408/943 (43.3)	130/998 (13.0)
Shibata et al[96]	23/47 (48.9)	32/47 (68.0)
Bielicki et al[111]	46/148 (31.1)	29/148 (19.6)
Hammel et al[175]	20/54 (37.0)	14/54 (25.9)
Overall	497/1192 (41.7)	204/1247 (16.4)

CEA: Carcino-embryonic antigen.

Table 4 Combined carcino-embryonic antigen and anti-p53 auto-antibody rates from all studies reporting the presence of both markers (n = 1192) n (%)

	CEA normal	CEA elevated
Anti-p53 ab present	112 (9.4)	90 (7.6)
Anti-p53 ab absent	584 (48.9)	406 (34.1)

CEA: Carcino-embryonic antigen.

Table 5 Anti-p53 auto-antibody and sero-conversion in colorectal cancer

Ref.	Patients, method	Follow-up	Findings
Müller et al[123]	303 patients, 197 colon, 46 rectal	Median 6 mo	All cancers: 3.6% (11/303) sero(-)→ (+); 3.6% (11/303) sero(+)→ (-); Total 7.2% (22/303) sero-conversion.
			Colon cancer: 3% (4/137) sero(-)→ (+); 3.6% (5/137) sero(+)→ (-); Total 6.6% (9/137) sero-conversion.
			Rectal cancer: 6.5% (2/31) sero(-)→ (+); 3.2% (2/31) sero(+)→ (-); Total 12.9% (4/31) sero-conversion
Lechpammer et al[88]	Immunoblot 32 , ELISA (Oncogene, Research Products, Cambridge, United States)	Up to 20 wk; 8 patients-48 wk	Non-significant decrease at 4 wk (pre-first cycle chemo) and significant decrease at 12 wk post-surgery
			Significant decreases during chemotherapy and 2 patients with anti-p53 increase at 12 wk (during chemotherapy) developed recurrence
			8 patients with extended follow-up: 7/8 had decreased anti-p53 with no recurrence. 1/8 anti-p53 decrease post-surgery/chemotherapy but increased at 12 wk corresponding with liver metastases. Anti-p53 fluctuates in response to tumour load but does not disappear. Anti-p53 levels reflects tumour load even during chemotherapy
Takeda et al[174]	30 CUR A, 5 CUR B, 5 CUR C, anti-p53 EIA, Pharmacell	Median 26 mo (13-144)	CUR A (n = 30): 28/30 sero(+)→(-) in 6 mo; 2 no sero-conversion: 1 recurrence
			CUR B (n = 5): 2 sero(+)→(-) no recurrence. 3 no sero-conversion, 2 had metastases
			CUR C: No sero-conversion
			Correlation between post-operative negative conversion and operative curability
Takeda et al[98]	17 mucosal/submucosal, ELISA (anti-p53 EIA, Pharmacell, France)	Up to 2 years	94%, 16/17 sero(+)→(-) within 3 wk post-surgery
			No recurrences as early stage tumours and hence not able to comment on anti-p53 and recurrence rates
Polge et al[128]	10, ELISA (Dianova, Hamburg, Germany)	Up to 6 mo	8 followed-up: 5/8 remained sero(+) post-operatively. All developed metastases
			3/8 decreased anti-p53 titres. No metastases or recurrence.
			Anti-p53 titres decreased within 1 mo of surgery/chemotherapy but no sero-conversion to anti-p53(-)
Angelopoulou et al[81]	6, “In house” immunofluorometric assay	Up to 17 mo	Anti p53 decreases with surgery/chemotherapy but persists at low levels
			Anti-p53 increases with recurrence
			Anti-p53 reflects tumour load more sensitively than CEA (n = 5) and in non-CEA producing tumour (n = 1)
Hammel et al[175]	12, “In house” ELISA	Up to 20 mo	Anti-p53 in 5/8 patients decrease by > 25% within 1 mo.
			At 1 year, 3 with normal anti-p53 levels and 3 with substantial decrease in anti-p53 remain disease-free
			2 patients with post-operative increased anti-p53: 1 developed recurrence and 1 developed metastases
			Anti-p53 decreased again following surgery in both patients. CEA and CA19-9 were normal in both cases

Sero(-): Sero-negative; Sero(+): Sero-positive; CUR A: No residual tumour macroscopically; CUR B: No residual tumour but not as evaluable as CUR A; CUR C: Definite residual tumour; CEA: Carcino-embryonic antigen.

Table 6 Anti-p53 auto-antibody and prognosis in colorectal cancer

Ref.	n (%)	Follow-up	Findings
Suppiah et al[130]	20/92 (21.7)	Median 97 mo	No difference in overall survival (62 mo vs 60 mo) or disease-free survival (73 mo vs 82 mo)
Müller et al[123]	70/243 (28.8)	5-year trial protocol	No survival difference with anti-p53 in CRC and other cancers. Trend towards decreased survival in anti-p53 positive patients with HCC and breast carcinoma
Tang et al[89]	130/998 (13)	Recruitment 1995-2000	Anti-p53 associated with decreased survival in univariate analysis but not multivariate analysis. Anti-p53 associated with advanced nodal disease (Stage N2→N3) and metastases (M1)
Chang et al[85]	147/167 (28)	Median 36.3 mo (22-85)	p53 mutation associated with poor differentiation and advanced stage. Multivariate analysis shows p53 mutation most significant survival predictor, followed by CRC stage. No prognostic significance of p53 protein expression or anti-p53
Shiota et al[112]	18/71 (25)	Not stated	Anti-p53 associated with shorter overall survival (20 mo vs 56 mo) but highly significant association with metastases (M1). Cox regression showed prognostic significance with liver metastases, TNM stage, Dukes stage, Ca19-9 and anti-p53 (in that order)
Kressner et al[131]	59/184 (32.1)	Median 6 years	Anti-p53 associated with decreased survival in univariate, but not multivariate analysis. Anti-p53 is independent prognostic indicator in Dukes’ A-C with curative surgery (i.e., when metastases excluded)
Houbiers et al[132]	65/255 (25.5)	36 mo	Anti-p53 associated with reduced overall (75% vs 88%) and disease-free survival (56% vs 64%) at 3 years in subgroup analysis of Dukes’ A and B1. No difference in overall survival (61% vs 68%) or disease-free survival (51% vs 58%) when all stages included

CRC: Colorectal cancer; HCC: Hepatocellular carcinoma.